MXPA01010148A - Recombinant and mutant adenoviruses derived of bovine adenovirus type 1. - Google Patents

Abstract

The present invention provides viral vectors, in one embodiment, the present invention provides mutant and recombinant bovine adenovirus having a deletion and / or insertion of DNA in region 4 of the early gene (E4), in another embodiment, the present invention provides a mutant and recombinant bovine adenovirus virus 1 having a deletion and / or insertion of DNA in the early gene region 3 (E3), the present invention also contemplates the use of viral vectors for vaccination, gene therapy or other applications according to be suitable

Description

ADENOVIRUS RECOMBINES BEFORE AND MUTANTS DERIVED FROM BOVINE ADENOVIRUS TYPE 1 FIELD OF THE INVENTION The present invention relates to viral vectors for the vaccination of animals. In particular, the present invention pertains to viral vectors that have insertion sites for the introduction of foreign DNA.

BACKGROUND OF THE INVENTION Adenovirus can cause enteric or respiratory infections in humans as well as in domestic and laboratory animals. Adenovirus insertion genes have been completed. In the human adenovirus genome (HuAd) there are two important regions: E1 and E3 in which foreign genes can be inserted to generate recombinant adenoviruses. This application of genetic engineering has resulted in several attempts to prepare adenovirus expression systems to obtain vaccines. Examples of such investigation include the description of the patent of E.U.A. No. 4,510,245 of a major adenovirus late promoter for expression in a yeast host; the patent of E.U.A. No. 4,920,209 of a live recombinant adenovirus type 7 with a gene encoding hepatitis B surface antigens; European Patent No. 389,286 of a non-defective human adenovirus recombinant expression system 5 in human cells; and published international application number WO 91/1525 of a viable non-pathogenic immunogenic canine adenovirus in a cell. However, because they are more suitable for entering a host cell, an indigenous adenovirus vector must be more suitable for use as a live recombinant virus vaccine in different animal species as compared to an adenovirus of human origin. For example, expression vectors based on bovine adenovirus for bovine adenovirus 3 (BAV-3) have been reported (see U.S. Patent No. 5,820,868). Bovine adenoviruses (BAV) comprise at least nine serotypes divided into two subgroups. These subgroups have been characterized based on enzyme-linked immunoassays (ELISA), serological studies with immunofluorescence assays, virus neutralization tests, immunoelectron microscopy and their host specificity and clinical syndromes. The viruses of subgroup 1 include BAV 1, 2, 3 and 9 and grow relatively well in established bovine cells compared to subgroup 2 viruses which include BAV 4, 5, 6, 7 and 8.

BAV-3 was isolated for the first time in 1965 and is the best characterized of the BAV genotypes and contains a genome of approximately 35 kilobases. The positions of the hexon and proteinase genes in the BAV-3 genome have been identified and sequenced. The genes of the bovine adenovirus 1 genome (BAV-1) have also been identified and sequenced. However, the localization and sequences of other genes such as certain regions of early genes in the BAV genome have not been reported. The continuous identification of suitable viruses and gene insertion sites are useful for the development of new vaccines. The selection of: (i) a suitable virus and (ii) the particular portion of the genome for use as an insertion site to create a vector for the expression of a foreign gene, however, presents an important challenge. In particular, the insertion site must be non-essential for the viable replication of the virus, as well as its functioning in tissue culture n vivo. In addition, the insertion site must be able to accept new genetic material, and at the same time ensure that the virus continues to replicate. What is needed is the identification of a novel virus and gene insertion sites for the creation of new viral vectors.

BRIEF DESCRIPTION OF THE INVENTION In one embodiment, the present invention provides recombinant viruses. Although not limited to a particular use, these recombinant viruses can be used to generate vaccines. Although not limited to a particular virus, in one embodiment the present invention provides a recombinant virus comprising a foreign DNA sequence inserted within the E4 gene region of a bovine adenovirus. In a preferred embodiment, the insertion is in a non-essential place. In another embodiment, the present invention provides a recombinant virus comprising a foreign DNA sequence inserted into the E3 gene region of a bovine adenovirus 1. In a preferred embodiment, the insertion is to a non-essential site. Although it does not limit its ability to replicate, in a preferred embodiment, the recombinant virus is replication competent. Likewise, although it is not limited to the foreign DNA to be inserted, in a preferred embodiment, the foreign DNA encodes a polypeptide and is from a virus or bacterium that is selected from the group consisting of bovine rotavirus, bovine coronavirus, bovine herpes virus type 1, bovine respiratory syncytial virus, bovine influenza virus type 3 (BPI-3), bovine diarrhea virus, bovine rhinotracheitis virus, bovine parainfluenza virus type 3, Pasteurella haemolytica, Pasteurella multocida and / or Haemophilus somnus In another preferred embodiment, the foreign DNA encodes a cytokine. In a further preferred embodiment, the polypeptide comprises more than ten amino acids and is antigenic. Finally, in a particularly preferred embodiment, the foreign DNA sequence is under the control of a promoter that is located towards the 5 * end of the foreign DNA sequence. The present invention also contemplates mutant viruses. Although not limited to a particular mutant virus, in one embodiment, the mutant virus comprises a deletion of at least a portion of the E4 gene region of a bovine adenovirus. In a preferred embodiment, the deletion is from a non-essential site. In another embodiment, the virus comprises a deletion of at least a portion of the E3 gene region of a bovine adenovirus 1. In a preferred embodiment, the mutant virus is capable of replication. In a further preferred embodiment, at least one open reading frame of the relevant gene region of the bovine adenovirus is completely deleted. In still another embodiment, the present invention provides a method for preparing a recombinant virus comprising inserting at least one exigenous gene or a fragment of a gene encoding at least one antigen in the genome of a virus, wherein the gene or The gene fragment has been inserted into region 4 of the early gene of a bovine adenovirus or has been inserted into region 3 of the early bovine adenovirus 1 gene. In a preferred embodiment, the method includes inserting at least a part of the genome of a virus into a bacterial plasmid, transforming the bacterium with the plasmid and incubating the bacterium at about 25 ° C. In another embodiment, the present invention provides vaccines. Although not limited to a particular vaccine, in one embodiment, the vaccines comprise the recombinant viruses described above. The present invention also contemplates methods of vaccination which include, but are not limited to the introduction of the vaccines described above to an animal.

DEFINITIONS The term "animal" refers to organisms in the animal kingdom. Thus, this term includes humans as well as other organisms. Preferably, the term refers to vertebrates. More preferably, the term refers to bovine animals. A "vector" is a replicon such as a plasmid, phage, cosmid, or virus, to which another DNA sequence has been attached so as to carry out the expression of the bound DNA sequence. For purposes of this invention, a "host cell" is a cell used to propagate a vector and its insertion. Infection of the cell can be carried out by methods well known to those skilled in the art, for example as set forth in Transfection of BAV-1 DNA in the following. A "coding sequence" of DNA is a DNA sequence which is transcribed and translated into a polypeptide in vivo when placed under the control of appropriate regulatory sequences. The limits of the coding sequence are determined by a start codon in the 5 '(amino) terminal part and a translation stop codon in the 3' (carboxyl) terminal part. A coding sequence can include, but is not limited to prokaryotic sequences, cDNA from eukaryotic mRNA, genomic DNA sequences from eukaryotic (eg mammalian) DNA, viral DNA, even synthetic DNA sequences. A polyadenylation signal and a transcription termination sequence can be located 3 'to the coding sequence. A "promoter sequence" is a DNA regulatory region capable of binding RNA polymerase or an auxiliary protein and initiating transcription of the coding sequence towards the 3 'direction. For the purpose of defining the present invention, the promoter sequence is in close proximity to the terminal 5 'part of the translation start codon (ATG) of a coding sequence and extends towards the 5' direction to include the minimum number of bases or elements necessary to facilitate transcription at detectable concentrations above the background. Within the promoter sequence will be found a transcription start site, as well as protein binding domains (consensus sequences) responsible for the binding of polymeric RNA. Eukaryotic promoters will often, but not always, contain "TATA" boxes and "CAAT" boxes, conserved sequences found in the promoter region of many eukaryotic organisms. A coding sequence is "operably linked to" or "under the control of" a promoter or control sequence in a cell when the polymeric RNA will interact with the promoter sequence directly or indirectly and will result in the transcription of a coding sequence into mRNA, which then it is translated into the polypeptide encoded by the coding sequence. A "double-stranded DNA molecule" refers to a polymeric form of deoxyribonucleotides (adenine, guanine, thymine or coticine) in its normal double-stranded helix. This term refers only to the primary and secondary structure of the molecule and is not limited to any of its particular tertiary forms. Thus, this term includes, for example, double-stranded DNA that is found in linear DNA molecules (eg, DNA restriction fragments of viruses, plasmids and chromosomes), as well as circular DNA and concatemerized forms. A "foreign DNA sequence" is a fragment of DNA that has been or will bind to another DNA molecule using recombinant techniques in which the particular DNA segment is not found in association with another DNA molecule in nature. The source of foreign DNA may or may not be a form of a separate organism in comparison in which it is placed. The foreign DNA can also be a synthetic sequence that has different codons from the native gene. Examples of recombinant techniques include, but are not limited to the use of restriction enzymes and ligases to divide DNA. An "insertion site" is a restriction site in a DNA molecule into which foreign DNA can be inserted. For purposes of this invention, a "homology vector" is a plasmid constructed to insert a foreign DNA sequence at a specific site in the genome of an adenovirus. The term "open reading frame" or "ORF" is defined as a genetic coding region for a particular gene that, when expressed, can produce a complete and specific polypeptide chain protein. A cell has been "transformed" with exogenous DNA when such exogenous DNA has been introduced into the cell membrane. The exogenous DNA may or may not be integrated (covalently linked) to chromosomal DNA constituting the genome of the cell. In prokaryotes and yeasts, for example, the exogenous DNA can be maintained in an episomal element such as a plasmid. The stably transformed cell is one in which the exogenous DNA has been integrated into the 0 chromosome so that it is inherited to daughter cells through chromosome replication. For mammalian cells, this stability is demonstrated by the ability of the cell to establish cell lines or clones made up of a population of daughter cells containing the exogenous DNA. A "replication competent virus" is a virus whose genetic material contains all of the DNA or RNA sequences necessary for replication as found in the wild-type organism. Therefore, a virus competent for replication does not require a second virus or a cell line to supply a certain defect or loss of the virus in order for it to replicate. A "non-essential site in the adenovirus genome" means a region in the adenovirus genome, the polypeptide product or the regulatory sequence of which is not necessary for infection or viral replication. Two polypeptide sequences are "substantially homologous" when at least about 80% (preferably at least about 90%, and more preferably at least about 95%) of the amino acids coincide over a defined length of the molecule. Two DNA sequences are "substantially homologous" when they are identical or do not differ in more than 40% of the nucleotides, more preferably about 20% of the nucleotides, and more preferably about 10% of the nucleotides.

A virus that has had a foreign DNA sequence inserted into its genome is a "recombinant virus" while a virus that has had a portion of its genome removed by intentional deletion (eg, by genetic engineering) is a "mutant virus" . The term "polypeptide" is used in its broadest sense, ie, any polymer of amino acids (dipeptide or greater) linked through the peptide bonds. Therefore, the term "polypeptide" includes proteins, oligopeptides, protein fragments, analogs, muteins, fusion proteins, etc. The term "antigenic" refers to the ability of a molecule that contains one or more epitopes to stimulate an animal or the immune system of a human by producing a specific response of humoral and / or cellular antigen. An "antigen" is an antigenic polypeptide. A "immunological response" to a composition or vaccine is the development in the host of a cellular immune response and / or antibody-mediated response to the composition or vaccine of interest. Usually, such a response consists of the subject producing antibodies, B cells, T helper cells, suppressor T cells and / or cytotoxic T cells specifically directed to an antigen or antigens included in the composition or vaccine of interest. The term "immunogenic polypeptide" and "immunogenic amino acid sequence" refers to a polypeptide or amino acid sequence, respectively, which induces antibodies that neutralize viral infectivity, and / or mediate antibody-complement cytotoxicity or cytotoxicity of antibody-dependent cells to provide protection from an immunized host. As used herein, an "immunogenic polypeptide" includes the full length (or near full length) sequence of the desired protein or an immunogenic fragment thereof. By "immunogenic fragment" is meant a fragment of a polypeptide which includes one or more epitopes and therefore induces antibodies that neutralize viral infectivity, and / or mediates the cytotoxicity of antibody-complement or antibody-dependent cells to provide protection of an immunized host. Such fragments will usually be at least about 5 amino acids in length, and preferably at least about 10 to 15 amino acids in length. There is no critical upper limit to the length of the fragment, which may comprise the almost complete length of the protein sequence, or even a fusion protein comprising fragments of two or more of the antigens. By "infectious" it is meant that it has the capacity to provide the viral genome inside the cells. A "substantially pure" protein will be free of other proteins, preferably at least 10% homogeneous, more preferably 60% homogeneous and much more preferably at least 95% homogenous.

BRIEF DESCRIPTION OF THE DRAWING FIGURE Figure 1 is a diagram of the genomic DNA of BAV-1 showing the relative size of various regions, in kilobase pairs. The fragments have letters in order of decreasing size.

DETAILED DESCRIPTION OF THE INVENTION After the description, reference is made to various publications, patents and patent applications. The descriptions of these publications, patents and patent applications are described herein by reference. The methods and compositions of the present invention involve modifying DNA sequences from various prokaryotic and eukaryotic sources by gene insertion, gene deletion, single or multiple base changes and subsequent insertions of these modified sequences into the genome of an adenovirus. An example includes inserting parts of an adenoviral DNA into plasmids in bacteria, reconstructing the viral DNA while it is in this state, so that the DNA contains deletions of certain sequences and / or addition by adding foreign DNA sequences either in place of deletions or on sites removed by deletions. Generally, the construction of the foreign gene is cloned in an adenovirus nucleotide sequence which represents only a part of the entire adenovirus genome, which may have one or more appropriate deletions. This chimeric DNA sequence is usually present in a plasmid which allows successful cloning to produce many copies of the sequence. The construction of the cloned foreign gene can then be included in the whole viral genome, for example by in vivo recombination after a DNA-mediated cotransfection technique. Multiple copies of a coding sequence or more than one of the coding sequences in the viral genome can be inserted so that the recombinant virus can express more than one foreign protein or multiple copies of the same protein. The foreign gene may have additions, deletions or substitutions to improve the expression and / or immunological effects of the expressed protein. For successful expression of the gene to occur, it can be inserted into an expression vector together with a suitable promoter including enhancer elements and polyadenylation sequences. Many of the eukaryotic and polyadenylation promoter sequences which provide for successful expression of foreign genes in mammalian cells and the manner of construction of expression cassettes are known in the art, for example in the U.A. number 5,151, 267. The promoter is selected to provide optimal expression of immunogenic protein which in turn satisfactorily leads to humoral, cell-mediated and mucosal immune responses, according to known criteria. The polypeptide encoded by the foreign DNA sequence is produced by expression in vivo in a recombinant cell infected with virus. The polypeptide can be immunogenic. More than one foreign gene can be inserted into the viral genome to obtain a successful production of more than one effective protein. Therefore, one utility of the use of a mutant adenovirus or the addition of a foreign DNA sequence in the genome of an adenovirus is to vaccinate an animal. For example, a mutant virus can be introduced into an animal to induce an immune response to the mutant virus. Alternatively, a recombinant adenovirus having a foreign DNA sequence inserted into its genome that encodes a polypeptide can also serve to induce an immune response in an animal to the foreign DNA sequence, the polypeptide encoded by the foreign DNA sequence and / or the adenovirus itself. Such a virus can also be used to introduce foreign DNA and its products into a host animal to alleviate a defective genomic condition in the host animal or to improve the genomic condition of the host animal. Although the present invention is not limited to the use of particular viral vectors, in preferred embodiments, the present invention utilizes bovine adenovirus expression vector systems. In particularly preferred embodiments, the present invention comprises a bovine adenovirus in which part or all of the region of the E4 gene is deleted and / or into which foreign DNA is introduced. Alternatively, the system comprises bovine adenovirus 1 (BAV-1), in which part or all of the regions of the E3 and / or E4 genes have been deleted and / or within which foreign DNA has been introduced. The present invention is not limited by foreign genes or coding sequences (viral, prokaryotic and eukaryotic) that are inserted within the bovine adenovirus nucleotide sequence, according to the present invention. Typically, the foreign DNA sequence of interest is derived from pathogens that in cattle cause diseases that have an economic impact on livestock or the dairy industry. Genes can be derived from organisms for which vaccines exist, and because of the novel advantages of vector-forming technology, vaccines derived from adenoviruses will be superior. In addition, the gene of interest can be derived from pathogens for which there is currently no vaccine but where there is a requirement for the control of the disease. Usually, the gene of interest codes for immunogenic polypeptides of the pathogen and may represent surface proteins, secreted proteins and structural proteins. The present invention is not limited to the particular organisms from which the foreign DNA sequence is obtained for the insertion of the gene into the bovine adenovirus genome. In preferred embodiments, the foreign DNA is bovine rotavirus, bovine coronavirus, bovine herpes virus type 1, bovine respiratory syncytial virus, bovine influenza virus type 3 (BPI-3), bovine diarrhea virus, bovine rhinotracheitis virus, virus for bovine influenza type 3, Pasteurella haemolytica, Pasteurella multocida and / or Haemophilus somnus. In another preferred embodiment, the foreign DNA encodes a cytokine. The present invention is also not limited to the use of a particular DNA sequence of such an organism. Frequently the selection of the foreign DNA sequence to be inserted into the adenovirus genome is based on the protein it encodes. Preferably, the foreign DNA sequence encodes an immunogenic polypeptide. The preferred immunogenic polypeptide to be expressed by the virus systems of the present invention contains antigens that encode full length (or near full length) sequences. Alternatively, shorter sequences that are immunogenic (i.e., encoding for u or more epitopes) may be used. The shorter sequence can code for a neutralizing epitope, which is defined as an epitope capable of inducing antibodies to neutralize the infectivity of the virus in an in vitro assay. Preferably, the peptide must code for a protective epitope that is capable of generating a protective immune response in the host; that is, an immune response mediated by antibody and / or mediated by cells that protects an immunized host from infection. In some cases, the gene for a particular antigen may contain a large number of introns or it may be an RNA virus. In these cases, a complementary DNA copy (cDNA) can be used. It is also possible to use fragments of nucleotide sequences of genes instead of the complete sequence as found in the wild-type organism. When available, synthetic genes or fragments thereof can also be used. However, the present invention can be used with a wide variety of genes and / or fragments, and is not limited to those indicated herein. Thus, the antigens encoded by foreign DNA sequences used in the present invention may be native or recombinant immunogenic polypeptides or fragments. They can be partial sequences, full-length sequences or even fusions (for example, having leader sequences appropriate for the recombinant host and / or the sequence of additional antigen for another pathogen). The present invention is also not limited by the ability of the resulting recombinant or mutant virus to replicate. In a preferred embodiment, the mutant and recombinant virus of the present invention are capable of replication. In this way, a complementary cell line is not necessary to make adequate supplies of the virus. As stated in the above, the present invention contemplates the administration of recombinant viruses and mutants of the present invention to vaccinate an animal. The present invention is not limited by the nature of administration to an animal. For example, the antigens used in the present invention, particularly when they comprise cholineric oligopeptides, can be conjugated to a vaccine carrier. Vaccine carriers are well known in the art: for example, bovine serum albumin (BSA), human serum albumin (HSA) and keyhole limpet hemocyanin (KLH). A preferred carrier protein, rolavirus VP6, is described in EPO publication number 0259149. Vaccines of the present invention that transpose foreign genes or fragments can also be administered orally in a suitable oral carrier such as an enteric coated dosage form. Oral formulations include commonly used excipients such as, for example, pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, saccharin sodium cellulose, magnesium carbonate, etc. Oral vaccine compositions may take the form of solutions (e.g. water), suspensions, tablets, pills, capsules, sustained release formulations or powders containing from about 10% to about 95% of the active ingredient, preferably about 25% to about 70% An oral vaccine may be preferable to increase mucosal immunity in combination with systemic immunity, which plays an important role in protecting against pathogens that infect the gastrointestinal tract. In addition, the vaccine can be formulated in a suppository. For suppositories, the vaccine composition includes traditional binders and carriers, such as polyalkylene glycols or triglycerides. Such suppositories can be formed of mixtures containing the active ingredient in a concentration of about 0.5% to about 10% (w / w), preferably from about 1% to about 2%. The protocols for administering the vaccine composition (s) of the present invention to animals are within the skill of the art in view of the present disclosure. Those skilled in the art will select a concentration of the vaccine composition in an effective dose to induce an immune response mediated by antibodies and / or T cells for the antigenic fragment. The administration protocol can also be important. For example, a primary inoculation may preferably be followed by subsequent booster inoculations, if needed. It may also be preferred, although optionally, to administer a second booster immunization to the animal several weeks to several months after the initial immunization. To ensure sustained high concentrations of protection against disease, it may be useful to re-administer a booster immunization to the animals, at regular intervals, for example once every several years. Alternatively, an initial dose may be administered orally followed by subsequent inoculations, or vice versa. Preferred vaccination protocols can be established through systematic or routine vaccination protocol experiments. The dosage for all routes of administration of a recombinant virus vaccine in vivo depends on several factors including the size of the patient, the nature of the infection against which protection is needed, the type of carrier and other factors, which are it can be easily determined by those usually skilled in the art. By way of non-limiting example, 2 can be used a dosage of between 103 plaque forming units (pfu) and 108 pfu. The present invention also includes a method for providing gene therapy to an animal in need thereof to control a gene deficiency. In one embodiment, the methods comprise administering to the mammal recombinant bovine adenovirus alive containing a foreign nucleotide sequence which codes for a non-defective form of a gene. The foreign nucleotide sequence is incorporated into the mammalian genome or maintained independently to provide for the expression of the required gene in the target organ or tissue. These kinds of techniques have recently been used by those skilled in the art to replace a defective gene or a portion thereof. For example, the patent of E.U.A. No. 5,399,346 to Anderson et al., describes techniques for gene therapies. In addition, examples of nucleotide sequences of foreign genes or portions thereof that can be incorporated for use in conventional gene therapy include, but are not limited to, a transmembrane conductance regulatory gene of cystic fibrosis, the human minidistrofin gene, the gene for alpha 1 antitrypsin and others. The methods for constructing, selecting and purifying recombinant adenovirus are detailed in the following materials, methods and examples below. The following serves to illustrate some preferred modalities and aspects of the present invention and are not considered as limiting the scope thereof.

Preparation of bovine adenovirus concentrate (BAV-1) Adenovirus concentrates are prepared by infecting tissue culture cells, Madin-Darby bovine kidney cells (MDBK) at a multiplicity of 0.01 pfu / cell infection in modified Eagle's medium. Dulbecco (DMEM) containing 2 mM glutamine, 100 units / ml penicillin, 100 units / ml streptomycin. These components are obtained from Sigma (St. Louis, MO) or an equivalent provider, and then referred to as complete DME medium) plus 1% fetal bovine serum. After the cytopathic effect is completed, the medium and the cells are harvested. After one or two cycles of freezing (-70 ° C) and reheating (37 ° C), aliquots are taken from the infected cells at a concentration of 1 ml and stored frozen at -70 ° C. Preparation of bovine adenovirus DNA All manipulations of bovine adenovirus are performed using strain 10 (ATCC VR-313). For the preparation of BAV-1 viral DNA from the cytoplasm of infected cells, MDBK cells are infected at a multiplicity of infection (MOI) sufficient to cause an extensive cytopathic effect before cellular overgrowth. All incubations are carried out at 37 ° C in a humidified incubator with 5% CO2 in air. The best yields of DNA are obtained when harvesting monolayers which are infected to the maximum, but which show incomplete cell lysis (usually 5-7 days). The infected cells are harvested by scraping the cells in the medium using a cell scraper (Costar brand). The cell suspension was centrifuged at 3000 rpm for 10 minutes at 5 ° C in a GS-3 rotor (Sorvall Instruments, Newton, CT). The resulting pellet is resuspended in cold PBS (20 ml / Roller Bottle) and subjected to centrifugation for 10 minutes at 3000 rpm cold. After decanting the PBS, the cell pellet is resuspended in 5 ml / revolving bottle of TE buffer (10 mM Tris, pH 7.5 and 1 mM EDTA) and expanded on ice for 15 minutes. NP40 (Nonidet P-40. ™, Sigma, St. Louis, MO) is added to a final concentration of 0.5% and kept on ice for another 15 minutes. The sample is centrifuged for 10 minutes at 3000 rpm cold to settle the nuclei and remove cellular debris. The supernatant fluid is carefully transferred to a 30 ml Corex centrifuge tube. SDS (sodium dodecyl sulfate, 20% concentrate) is added to the sample at final concentrations of 1%. 200 μl of K-proteinase is added at 10 mg / ml (Boehringer Mannheim, Indianapolis, IN) by rotating sample bottle, mix and incubate at 45 ° C for 1 -2 hours. After this period, an equal volume of phenol saturated with water is added to the sample and mixed by swirling. The sample is rotated in a clinical centrifuge for 5 minutes at 3000 rpm to separate the phases. NaAc is added to the aqueous phase to a final concentration of 0.3M (concentrated solution 3, pH 5.2) and the nucleic acid is precipitated at -70 ° C for 30 minutes after the addition of 2.5 volumes of cold absolute ethanol. The DNA in the sample is pelleted by centrifugation for 20 minutes at 8000 rpm in a HB-4 rotor at 4 ° C. The supernatant is carefully removed and the DNA pellet washed once with 25 ml of 80% ethanol. The DNA pellet is briefly dried in vacuo (2-3 minutes), and resuspended in 2 ml of rotating bottle of infected TE buffer cells (20 mM Tris, pH 7.5, 1 mM EDTA), 10 I of RNaseA at 10 mg / ml (Sigma, St. Louis, MO) is added and incubated at 37 ° C for one hour. 0.5 ml of 5N NaCl and 0.75 ml of 30% PEG are added and precipitated at 4 ° C overnight. The DNA in the sample is pelleted by centrifugation for 20 minutes at 8000 rpm in a HB-4 rotor at 4 ° C. The pellet is resuspended in 2 ml of TES buffer (20 mM Tris, pH 7.5, 1 mM EDTA and 0.2% SDS) and extracted with an equal volume of phenol saturated with water. The sample is centrifuged in a clinical centrifuge for 5 minutes at 3000 rpm to separate the phases. NaAc is added to the aqueous phase to a final concentration of 0.3M (3M concentrated solution, pH 5.2) and the nucleic acid is precipitated at -70 ° C for 30 minutes after the addition of 2.5 volumes of cold absolute ethanol. The DNA in the sample is pelleted by centrifugation for 20 minutes at 8000 rpm in an HB-400 rotor at 5 ° C. The supernatant is carefully removed and the DNA pellet washed once with 25 ml of 80% ethanol. The DNA pellet is briefly dried in vacuo (2-3 minutes) and resuspended in 200 μl / rotor vial of infected TE buffer cells (10 mM Tris, pH 7.5, 1 mM EDTA). All viral DNA is stored at approximately 4 ° C. Molecular biology techniques Techniques for the manipulation of bacteria and DNA, including procedures such as restriction endonuclease digestion, gel electrophoresis, DNA extraction from gels, ligation, phosphorylation with kinase, phosphatase treatment, crop growth bacterial, transformation of bacteria with DNA and other molecular biological methods are described by Maniatis et al. (T. Maniatis et al., Molecular Cloning: A Laboratory Manual, Cold Spring Harbor, N.Y. (1 982)) and Sambrook et al. (J. Sambrook, et al., Molecular Cloning: A Laboratory Manual Second Edition, Cold Spring Harbor Press, Cold Spring Harbor, N.Y. (1989)). The polymerase chain reaction (PCR) is used to induce convenient restriction sites for the manipulation of several DNAs. The procedures used are also described by Innis et al (M. A. Innis et al., PCR Protocols; A Guide to Methods and Applications, pp. 84-91, Academic Press, Inc., San Diego, CA (1990)). In general, the amplified fragments are of a size less than 2000 base pairs and the critical regions of the amplified fragments are confirmed by DNA sequencing. Except where indicated, these techniques are used with minor variations. DNA sequencing DNA sequencing is performed on an Applied Biosystems 373A automated sequencer (with an XL enhancement) per manufacturer's instructions. The subclones are elaborated to facilitate the sequencing. Internal primers are synthesized on an ABI 392 DNA synthesizer or obtained commercially (Genosys Biotechnology, I nc., The Woodlands, TX). Larger DNA sequences are constructed using consecutive superposition primers. The sequence through the junctions of large genomic subclones is determined directly using a full-length genomic clone as a template. The assembly, manipulation and comparison of the sequences is done with DNAstar programs. Comparisons with GenBank are made using the NCBI BLAST programs ((Altschul, Stephen F., Thomas L. Madden, Alexander A. Scháffer, Zhang Jinghui, Zheng Zhang, Webb Miller, and David J. Lipman (1997), "Gapped BLAST and PSI-BLAST: a new generation of protein data search programs ", Nucleic A cids Res. 25: 3389.3402) Construction of recombinant BAV-1 genomes in E. coli Recombinant BAV-1 genomes are constructed by homologous recombination according to with the method of C. Chartier et al (996) J. of Virology 70: 805-481 0. A small, easily manipulated plasmid is constructed, containing approximately 1,000 base pairs, each of the left and right ends of the BAV-1 genome The homologous recombination between this vector and the BAV-1 genomic DNA results in a plasmid that contains the entire BAV-1 genome (adenoviral main structure vector) .This genomic plasmid BAV-1 can be used to generate recombinant genomes by li plasmid nealization and recombination with homology DNA engineered to contain foreign DNA bleached by DNA derived from the desired BAV-1 insertion site. Note that to linearize the adenoviral main structure vector, a rare enzyme within the region analogous to the flanking sequences of BAV-1 must be located. We mapped the restriction sites of such an enzyme. Pvul cuts the BAV-1 genome in two positions, one in the D fragment BamHl and one in the C Bam fragment (see Figure 1). The adenoviral backbone vector contains a third Pvul site within the antibiotic resistance gene of the plasmid. The Pvul site within the C BamHI fragment is suitable for gene insertion sites within the E3 and E4 regions. Therefore, a partial Pvul digestion of the adenoviral backbone vector will provide a subpopulation of linearized molecules at the Pvul site in the C BamHI fragment. These molecules can be recombined with the homology DNA to generate a viable plasmid. The linearized molecules in the other two sites are not able to recombine to generate viable plasmids. The high competition of E. coli bacteria cells BJ5183 recBC sbcBC (D. Hanahan (1983) J. Mol. Biol .. 166: 557-580) is desired to obtain efficient recombination. Typically, 10 ng of a restriction fragment containing foreign DNA flanked by the appropriate BAV insertion sequences (homology DNA) is mixed with 1 ng of the linearized adenoviral backbone vector in a total volume of 10 I. 50 I of competent BJ5183 cells are added. After 15 min, on ice, at 5 min at 37 ° C and 15 min on ice, 200 I of LB are added and the cells are seeded on plates, on agar containing LB + 80 g / ml carbenicillin, after a hour at 37 ° C.

A low temperature (25-27 °) is essential for small-scale crop growth (to examine carbenicillin-resistant colonies) and subsequent large-scale cultures (for isolation of large amounts of plasmid DNA). The CarbR colonies are first grown in cultures of 4-5 ml at 25-27 ° C for two days. Small scale DNAs are prepared using the boiling method (J. Sambrook, et al., Molecular Cloning: A Laboratory Handbook Second Edition, Cold Spring Harbor Press, Cold Spring Harbor, NY (1989)) and analyzed by analysis. of DNA restriction. To purify the DNA by removing any concatemer of vector DNA, the DNA of the correct clones are transformed back into DH10B cells (Life Technologies). The analysis of bacterial colonies is repeated by DNA restriction analysis. The glycerol concentrates are prepared from the correct clones and stored at -70 ° C. Large amounts of plasmid DNA are prepared using the Qiagen Plasmid Kit (Qioagen Inc.) or the large-scale boiling method from 250 ml of culture which is inoculated with glycerol concentrate and grown at 37 ° C during one day. Transfection of BAV-1 DNA Approximately 1.5 x 1 05 cells / ml are plated in the plate (MDBK) in plates of 6 cm 24 h before transfection, time in which they reach 50-70% of confluence. For transfection, the Lipofection method is used, according to the manufacturer's instructions (Lipofectibn, Life Technologies, Rockville, MD). A transfection mixture is prepared by adding several (4-15) g of BAV-1 viral DNA or linearized genome plasmid DNA and 50 I of Lipofectin reagent to 200 I of serum-free medium, according to the manufacturer's instruction . After incubation at room temperature for 15-30 min, the transfection mixture is added to the cells. After 4-6 h at 37 ° C, the medium containing the transfection mixture is removed and 5 ml of growth medium is added. The cytopathic effect becomes evident in the following 7-10 days. The transfected virus concentrate is harvested by scraping cells in the culture and stored at -70 ° C. Purification in plates of recombinant constructs Monolayers of MDBK cells in 6 cm or 10 cm plates are infected with transfection concentrate, coated with nutrient agarose medium and incubated for 5-10 days at 37 ° C. Once the plates are developed, the plates are taken alone and well isolated on MDBK cells. After 5-10 days, when 80-90% of the cytopathic effect has been reached, the infected cells are harvested (concentrate P1) and stored at -70 ° C. This procedure is repeated once more with the concentrate P1. Cloning of the gene for glycoprotein 53 (g53) of bovine viral diarrhea virus (BVDV) The bovine viral diarrhea g53 gene is cloned by a PCR cloning procedure essentially as described by Katz et al., (Journal of Virology 64: 1 808-1 81 1 (1990)) for the HA gene of human influenza. Viral RNA prepared from the BVD virus of the Singer strain is grown in DMBK cells and priomer is converted to cDNA using an oligonucleotide primer specific for the target gene. The cDNA is then used as a template for cloning by polymerase chain reaction (PCR) (MA I nnis et al., PCR Protocols: A Guide to Methods and Applications, 84-91, Academic Press, Inc., San Diego (1990 )) of the target region. The PCR primers are designed to incorporate restriction sites which allow the cloning of the amplified coding regions into vectors containing the appropriate signals for expression in BAV-1. A pair of oligonucleotides are required for the coding region. The coding region for the g53 gene (amino acids 1-394) from the BVDV Singer strain (MS Collett et al., Journal of Virology, 65, 200-208 (1988)) is cloned using the following primers: 5'- CTTGGATCCTCATCCATACTGAGTCCCTGAGGCCTTCTGTTC-3 '[SEQUENCE OF N UMBER I DENTIFICATION: 1] for primed cDNA and combined with 5'-CATAGATCTTGTGGTGCTGTCCGACTTCGCA-3' [SEQUENCE OF I NDENTIFICATION N UMBER: 2] for PCR.

Samples of Uses and protein standards are run on a polyacrylamide gel according to the procedure of Laemnli, Nature, 227, 680-685 (1 970)). After gel electrophoresis, the proteins are transferred and processed according to Sambrook, et al., Molecular Cloning A Laboratory Handbook Second Edition, Cold Spring Harbor Press, Cold Spring Harbor, New York (1989). The primary antibody is a mouse monoclonal antibody (Mab 6.12.2 from Dubovi, Cornell University, Ithaca, New York) diluted 1: 1 00 with dry milk without 5% fat and trisodium chloride, and sodium azide (TSA: 6.61 g of Tris-HCl, 0.97 g of Tris-base, 9.0 of NaCl and 2.0 g of sodium azide per liter of H20). The secondary antibody is a goat anti-mouse antibody conjugated with alkaline phosphatase, diluted 1: 1000 with TSA. Plasmid 990-1 1 Plasmid 990-1 1 contains the left and right ends of the BAV-1 genome. These sequences are cloned by standard PCR methods (MA Innis, et al., PCR Protocols: A Guide to Methods and Applications, pp. 84-91, Academic Press, Inc., San Diego, CA (1990) using primers based In the sequences determined for the EcoRI A and SamHI F fragments respectively, the primers (1) 5'-3 'GGCCTTAATTAACATCATCAATAATATACGGAACAC [SEQUENCE OF I DENTIFICATION NUMBER: 5] and (2) 5'-3' GGAAGATCTTGAGCATGCAGAGCAATTCACGCCGGGTAT [IDENTIFICATION SEQUENCE] are used. NUMBER: 6] to PCR the left end of BAV-1 Since three repeating elements within this region share the same 5 'end sequences (ie, primer 1), fragments of BAV-1 DNA are amplified. 1760 bp, 1340 bp and 920 bp by PCR The 920 bp DNA fragment is cloned into the blunt PCR vector (Invitrogen, Carlsbad, CA) Primers (1) and (3) 5'-GGCAATGAGATCTTTTGGATGACAAGCTGAGCTACGCG-3 are used. '[SEQUENCE OF IDENTIFICATION NUMBER: 7] for the PCR of the end of BAV-1, the 740 bp and 1160 bp PCR products are amplified and the DNA of the 1160 bp fragment is cloned into the PCR-blunt vector (Invitrogen, Carlsbad, CA). The plasmid 990-11 is then constructed by cloning the BAV-1 end fragments into the polylinker of plasmid pPolyll (R. Lathe, J.L. Vilotte, and A.J. Ciar, Gene, 57: 193-201, 1997). The end fragments are cloned as a single PacI fragment containing a unique BglW site in its internal junction. Only the BamH \ and EcoRI sites are retained from the polylinker. Plasmid 990-50 (adenoviral backbone vector) Plasmid 990-50 is constructed according to the method described above (Construction of Recombinant BAV-1 Genomes in E. coli). Briefly, the cotransformation of the linearized plasmid with BglW 990-11 and the BAV-1 genomic DNA regenerates a stable circular plasmid containing the entire BAV-1 genome. In this plasmid, the Paci sites flank the inserted genomic sequences of BAV-1.

Since Pac is absent from the genomic DNA of BAV-1, digestion with this enzyme allows precise cutting of the full-length BAV-1 genome of plasmid 990-50. Plasmid 996-80D Plasmid 996-80D contains DNA encompassing approximately 5945 base pairs of the right end of the BAV-1 genome from which the "G" and "H" fragments of EcoRI have been deleted and replaced with a Sma site. \ synthetic The plasmid is constructed for the purpose of suppressing a portion of the E4 region of BAV-1. It can also be used to insert foreign DNA into recombinant BAV-1 genomes. It contains a unique Smal restriction enzyme site in which foreign DNA can be inserted. The plasmid can be constructed using standard recombinant DNA techniques (see above Molecular Biological Techniques) by binding restriction fragments from the following sources with the indicated synthetic DNA sequences. The plasmid vector is derived from a HindWl or Pvu restriction fragment of approximately 2774 base pairs from pSP64 (Promega Corporation, Madison, Wl). The synthetic linker sequence 5'-CTGTAGATCTGCGGCCGCGTTTAAACGTCGACAAGCTTCCC-3 '[IDENTIFICATION SEQUENCE NUMBER: 8] is linked to the PvuW site of pSP64 (Promega Corporation, Madison, Wl). Fragment 1 is approximately 1693 base pairs of the Psfl to EcoRI subfragment of fragment "C" BamHI BAV-1 (positions 28241 to 29933 of IDENTIFICATION SEQUENCE NUMBER: 3] The synthetic linker sequence 5'-AATTCGAGCTCGCCCGGGCGAGCTCGA-3 ' [IDENTIFICATION SEQUENCE NUMBER: 9] is ligated to fragment 1 by retaining the EcoRI sites at both ends of the linker sequence.Fragment 2 is about 48 base pairs of the restriction subfragment EcoRI to BamH I of fragment "C" SamH I of BAV-1 (positions 31732 to 31 779 of the SEQUENCE OF I DENTI FICATION NUMBER: 3.) Fragment 3 is about 2406 base pairs of fragment "F" SamHI of BAV-1 (positions 31 780 to 34185 of the NUME RO IDENTIFICATION SEQUENCE: 3] The synthetic linker sequence 5'- GACTCTAGGGGCGGGGAGTTTAAACGCGGCCGCAGATCTAGCT -3 '[SEQUENCE OF I DENTIATION NUMBER: 10] is ligated between fragment 3 and the Hind site of pSP64 (Promega Corporation, Madison, Wl.) N tese that BAV-1 sequences can be cut in this piásmido via the restriction sites Not \ that are located in the flanking synthetic linker sequences. Piásmido 1004-73.1 6.14 Piásmido 1004-73.1 6.14 contains a recombinant BAV-1 genome from which the "G" and "H" fragments of EcoRI have been deleted and replaced by the synthetic Smal site (5'-GAATTCGAGCTCGCCCGGGCGAGCTCGAATTC -3 ') [SEQUENCE OF IDENTIFICATION NUMBER: 1 1]. This plasmid can be used to generate recombinant bovine adenovirus vectors with their pressure and insertion of genes in the E4 region. The plasmid can be constructed according to the above method (Construction of recombinant BAV-1 genomes in E. coli). The homology DNA is derived from the Not \ insert of plasmid 996-80D and the adenoviral backbone vector plasmid 990-50 is linearized by partial digestion with Pvu \. Plasmid 1004-07.16 Plasmid 1004-07.1 6 is constructed by inserting a g53 gene BVDV, engineered to be under the control of the immediate early promoter of human cytomegalovirus (I nvitrogen, Carlsbad, CA), within the single Smal site of plasmid 996-80D. The g53 gene of BVDV is isolated according to the above method (Cloning of the g53 gene of bovine viral diarrhea virus). Plasmid 1004-40 Plasmid 1004-40 contains a recombinant BAV-1 genome from which the G and H fragments of EcoRI have been deleted. The gene for glycoprotein 53 (g53) (amino acids 1-394) of the bovine viral diarrhea virus (BVDV) under the control of the HCMV immediate early promoter is inserted into the suppressed region. The plasmid can be constructed according to the above method (Construction of the recombinant BAV-1 genomes in E. coli). The homology DNA is derived from the Not \ insert of plasmid 1 004-1 7.16 and the plasmid 990-50 of the adenoviral backbone vector is linearized by partial digestion with Pvu \. Plasmid 101 8-14.2 Plasmid 1018-14.2 contains DNA that flanks the region E3 of BAV-1, from which a specific region of this sequence flanked by the Sa / I and SamH I sites has been deleted (positions 25664 to 26840 of the SEQUENCE OF I NDENTIFICATION OF NU MERO: 3). The plasmid is constructed for the purpose of suppressing the corresponding portion of the E3 region of BAV-1. It can also be used to insert foreign DNA into the recombinant BAV-1 genomes. It contains a unique HindA U restriction enzyme site into which foreign DNA can be inserted. The plasmid can be constructed using standard recombinant DNA techniques (see above Molecular Biological Techniques) by binding restriction fragments from the following sources with the indicated synthetic DNA sequences. The plasmid vector is derived from a restriction fragment Hind / \ \ \ at PvuW of approximately 2774 base pairs from pSP64 (Promega Corporation, Madison, Wl). The synthetic linker sequence 5'-CTGTAGATCTGCGGCCGCGTTTAAACG-3 '[SEQUENCE OF IDENTIFICATION NUMBER: 12] is ligated to the PvuW site of pSP64 (Promega Corporation, Madison, Wl). Fragment 1 is about 2665 base pairs from subfragment Sa / I to Sa / I (positions 22999 to 25663 of IDENTIFICATION SEQUENCE NUMBER: 3) of the Sa HI BAV-1 fragment of BAV-1. Fragment 1 is linked to the synthetic sequence towards the 5 'end by retaining the Sa / I site at the junction. Fragment 1 contains a unique Aval site (positions 25317 to 25322 of IDENTIFICATION SEQUENCE NUMBER: 3). Fragment 1 is oriented such that the Ava \ unique site is closest (406 base pairs) to fragment 2 as compared to the plasmid vector. The synthetic linker sequence 5'-TCGACAAGCTTCCC-3 '(IDENTIFICATION SEQUENCE NUMBER: 13) is ligated to the second end of fragment 1 again retaining the Sa / I site at the junction. Fragment 2 is about 4223 base pairs from the eamHI to HindAW restriction subfragment of fragment C of eamHI from BAV-1 (positions 26851 to 31973 of SEQUENCE IDENTIFICATION NUMBER: 3). Note that the ends of both fragments are blunt ended by treatment with T4 polymerase. The synthetic linker sequence 5'-CCCGGGAGTTTAAACGCGGCCGCAGATCTAGCT-3 '[IDENTIFICATION SEQUENCE NUMBER: 14] is ligated between fragment 2 and the HindAW site of pSP64 (Promega Corporation, Madison, Wl). Note that the HindAW site is not retained. The BAV-1 sequences can be cut out of this plasmid via the Not? Restriction sites that are located in the flanking synthetic linker sequences. Plasmid 1018-75 Plasmid 1018-75 contains a recombinant BAV-1 genome from which the specific region of BamH fragment "B" has been deleted (positions 25664 to 26840 of SEQUENCE OF IDENTIFICATION NUMBER: 3). This plasmid can be used to generate recombinant bovine adenovirus vectors with their pressures and insertions of genes in the E3 region. The plasmid can be constructed according to the above method (Construction of recombinant BAV-1 genomes in E. coli). The homology DNA is derived from the Not \ insert of plasmid 1018-14.2 and the adenoviral backbone vector plasmid 990-50 is linearized by partial digestion with Pvu. Plasmid 1018-23C1 5 Plasmid 1018-23C1 5 is constructed by inserting the g53 gene from BVDV, engineered to be under the control of the human cytomegalovirus immediate early promoter (I nvitrogen, Carlsbad, CA), within the unique HindAW site of plasmid 1018-14.2. The gDV gene of BVDV is isolated according to the above method (Cloning of the bovine viral diarrhea virus g53 gene) Plasmid 1018-42 Plasmid 1018-42 contains a recombinant BAV-1 genome from which a recombinant BAV-1 has been deleted. specific region of fragment "B" of SamHI (positions 25664 to 26840 of SEQUENCE OF IDENTIFICATION NUMBER: 3). The gene for bovine viral diarrhea virus (BVDV) glycoprotein 53 (g53) (amino acids 1-394) under the control of the HCMV immediate early promoter is inserted into the suppressed region. The plasmid can be constructed according to the above method (construction of the recombinant BAV-1 genomes in E. coli). The homologous DNA is derived from the Not? Insertion of plasmid 1 01 8-23C1 5 and the plasmid 990-50 of the adenoviral backbone vector is linearized by partial digestion with Pvu ?. Plasmid 1018-45 Plasmid 1 01 8-45 contains DNA flanking the E3 region of BAV-1, from which a specific region of this sequence flanked by the EcoRI and SamH I sites has been primed (positions 25765 to 26850 of the N UME RO IDENTIFICATION SEQUENCE: 3). The plasmid is constructed for the purpose of suppressing the corresponding portion of the E3 region of BAV-1. It can also be used to insert foreign DNA into the recombinant BAV-1 genomes. It contains a unique HindA W restriction enzyme site into which foreign DNA can be inserted. The plasmid can be constructed using standard recombinant DNA techniques (see above molecular biology techniques) by joining restriction fragments from the following sources with the synthetic DNA sequences indicated. The plasmid vector is derived from a restriction fragment HindA W to PvuW of approximately 2774 base pairs from pSP64 (Promega Corporation, Madison, Wl). The synthetic linker sequence 5'- CTGTAGATCTGCGGCCGCGTTTAAACG-3 '[SEQUENCE OF I DENTIFICATION NUM ER0: 12] is linked to the PvuW site of pSP64 (Promega Corporation, Madison, Wl). Fragment 1 is a subfragment of SacI to EcoR of approximately 1 582 base pairs (positions 24183 to 25764 of SEQUENCE OF IDENTIFICATION NUMBER: 3) of fragment B Bam of BAV-1. Fragment 1 is linked to the synthetic sequence towards the 5 'end. The fragment becomes blunt ended with the T4 polymerase treatment so that the SacI or EcoRI sites are not retained. Fragment 1 contains a unique A site (positions 25317 to 25322 of the SEQUENCE OF IDENTIFICATION NUMBER: 3). Fragment 1 is oriented so that a unique Aval site is closer (406 base pairs) to fragment 2 than the plasmid vector. The synthetic linker sequence 5'-CAAGCTTCCC-3 '[SEQUENCE OF IDENTIFICATION NUMBER: 17] is ligated to the second end of fragment 1 again retaining the Sa / I site at the junction. Fragment 2 is a BamHI to Hindi restriction subfragment of approximately 4223 base pairs of the C SamH I fragment of BAV-1 (positions 26851 to 31073 of SEQUENCE OF I NDENTIFICATION NUMBER: 3). Note that the end of both fragments have blunt ends by treatment with T4 polymerase. The synthetic linker sequence 5'- CCCGGGAGTTTAAACGCGGCCGCAGATCTAGCT-3"[SEQUENCE OF I NDENTIFICATION NUMBER: 14] is ligated between fragment 2 and the Hind site of pSP64 (Promega Corporation, Madison, Wl). retains the Hind site The BAV-1 sequences can be trimmed from this plasmid via Nou restriction sites that are located in the flanking synthetic linker sequences Plasmid 1028-03 Plasmid 1028-03 contains a recombinant BAV-1 genome from which a specific region of fragment "B" of BamH I has been deleted (positions 25765 to 26850 of the SEQUENCE OF IDENTIFICATION NUMBER: 3) This plasmid can be used to generate vectors of recombinant bovine adenovirus with its pressures e gene insertions in the E3 region The plasmid can be constructed according to the above method (construction of the recombinant BAV-1 genomes in E. coli) The homology DNA is derived from the Not \ insert plasmid 1 01 8-45 and the plasmid 990-50 of the adenoviral main structure vector is linearized by partial digestion with Pvu \. Plasmid 1028-77 Plasmid 1028-77 is constructed by inserting a gDV gene from BVDV, engineered to be under the control of the human cytomegalovirus immediate early promoter (I nvitrogen, Carlsbad, CA), within the Hind site. unique of plasmid 1018-45. The BVDV g53 gene is isolated according to the above method (Cloning of the bovine viral diarrhea virus g53 gene) Plasmid 1038-16 Plasmid 1038-16 contains a recombinant BAV-1 genome from which a recombinant BAV-1 genome has been deleted. specific region of fragment "B" of BamH I (positions 25765 to 26850 of SEQUENCE OF I DENTIFICATION NUMBER: 3). The gene for bovine viral diarrhea virus (BVDV) glycoprotein 53 (g53) (amino acids 1-394) under the control of the HCMV immediate early promoter is inserted into the suppressed region. The plasmid can be constructed according to the above method (construction of the recombinant BAV-1 genomes in E. coli). The homology DNA is derived from the Not \ insert of plasmid 1028-77 and the plasmid 990-50 of the adenoviral backbone vector is linearized by partial digestion with Pvu \. Plasmid 1054-83 Plasmid 1054-93 contains DNA derived from the E4 region of BAV-1. The sequence corresponding to positions 33614 to 33725 of the SEQUENCE OF I DENTIFICATION NUMBER: 3) has been deleted and replaced with a synthetic Pst site. The plasmid is constructed for the purpose of suppressing the corresponding portion of the E4 region of BAV-1. It can also be used to insert foreign DNA into the recombinant BAV-1 genomes. Contains a restriction enzyme site Unique Psil into which strange DNA can be inserted. The plasmid can be constructed using standard recombinant DNA techniques (see molecular biology techniques above) by joining restriction fragments from the following sources with the indicated synthetic DNA sequences. Note that fragments derived from BAV-1 are ligated in the orientation indicated by the positions provided for each fragment. The plasmid vector is derived from a Hind \\\ Pvull restriction fragment of approximately 2774 base pairs from pSP64 (Promega Corporation, Madison, Wl). The synthetic linker sequence 5'-CTGTAGATCTGCGGCCGCGTTTAAACGTCGACAAGCTTCCC-3 '[IDENTIFICATION SEQUENCE NUMBER: 8] is linked to the PvuW site of pSP64 (Promega Corporation, Madison, Wl). Fragment 1 is a Psp \ a BamVW subfragment of approximately 3538 base pairs of the "C" SamHI fragment of BAV-1, positions 28241 to 31779 of SEQUENCE OF IDENTIFICATION NUMBER: 3). Fragment 1 is ligated to the 3 'end of the synthetic linker sequence (IDENTIFICATION SEQUENCE NUMBER: 8). Fragment 2 is a PCR fragment of approximately 1832 base pairs containing sequences derived from the BAV-1 genome (positions 31780 to 33613 of SEQUENCE OF IDENTIFICATION NUMBER: 3). Fragment 2 is ligated to fragment 1 so that the SamHI site is retained at the junction. The synthetic linker sequence 5'-CTGCAG-3 '[IDENTIFICATION SEQUENCE NUMBER: 4] is linked to fragment 2. Fragment 3 is a PCR fragment of approximately 460 base pairs containing sequences derived from the genome of BAV-1 ( positions 33725 to 34185 of SEQUENCE OF IDENTIFICATION NUMBER: 3). Fragment 3 is ligated to the 3 'end of the synthetic linker sequence 5'-CTGCAG-3'. The synthetic linker sequence 5'-GACTCTAGGGGCGGGGAGTTTAAACGCGGCCGCAGATCTAGCT-3 '[IDENTIFICATION SEQUENCE NUMBER: 10] is ligated between fragment 3 and the Hind site of pSP64 (Promega Corporation, Madison, Wl). Note that BAV-1 sequences can be cut out of this plasmid via the? / Ofl restriction sites that are located in the flanking synthetic linker sequences. Plasmid 1055-38 Plasmid 1055-38 contains DNA derived from the E4 region of BAV-1. The sequence coding for nORF13 (see Table 1) has been deleted and replaced with a synthetic Psii site. The plasmid is constructed for the purpose of suppressing a portion of the E4 region of BAV-1. It can also be used to insert foreign DNA into the recombinant BAV-1 genomes. It contains a unique Psü restriction enzyme site into which foreign DNA can be inserted. The plasmid can be constructed using standard recombinant DNA techniques (see above, Molecular Biology Techniques) by joining DNA fragments from the following sources with the indicated synthetic DNA sequences. Note that fragments derived from BAV-1 DNA are ligated in the orientation indicated by the positions provided for each fragment. The plasmid vector is derived from a restriction fragment H¡nd \ to PvuW of approximately 2774 base pairs of pSP64 (Promega Corporation, Madison, Wl). The synthetic linker sequence 5'-CTGTAGATCTGCGGCCGCGTTTAAACGTCGACAAGCTTCCC-3 '[IDENTIFICATION SEQUENCE NUMBER: 8] is linked to the PvuW site of pSP64 (Promega Corporation, Madison, Wl). Fragment 1 is a PCR fragment of approximately 1282 base pairs containing sequences derived from the BAV-1 genome (positions 28240 to 29522 of SEQUENCE OF IDENTIFICATION NUMBER: 3). Fragment 1 is ligated to the 3 'end of the synthetic linker sequence indicated above (SEQUENCE OF IDENTIFICATION NUMBER: 8). The synthetic linker sequence 5'-CTGCAG-3 '[IDENTIFICATION SEQUENCE NUMBER: 4] is linked to fragment 1. Fragment 2 is a PCR fragment of approximately 1372 base pairs containing the sequences derived from the BAV-1 genome (positions 30407 to 31779 of SEQUENCE OF IDENTIFICATION NUMBER: 3). Fragment 2 is ligated to the 3 'end of the synthetic linker sequence 5'-CTGCAG-3' [SEQUENCE OF IDENTIFICATION NUMBER: 4]. Fragment 3 is the "F" fragment of BamHI of BAV-1 of approximately 2406 base pairs (positions 31779 to 34185 of SEQUENCE OF IDENTIFICATION NUMBER: 3). Fragment 3 is ligated to the 3 'end of fragment 2. The synthetic linker sequence 5'-GACTCTAGGGGCGGGAGTTTAAACGCGGCCGCAGATCTAGCT-3' [IDENTIFICATION SEQUENCE NUMBER: 10] is ligated between fragment 3 and the Hind site [pSP64] (Promega Corporation, Madison, Wl). Note that the BAV-1 sequences can be cut out of this plasmid via the Nou restriction sites that are located in the flanking synthetic linker sequences. Plasmid 1055-52 Plasmid 1055-52 contains a recombinant BAV-1 genome from which a portion of the E4 region (positions 29522-30407 of the SEQUENCE OF I DENTIFICATION NUMBER: 3) has been deleted and replaced by a synthetic Psü site (5'-CTGCAG-3 ') [SEQUENCE OF IDENTIFICATION NUMBER: 4]. This plasmid can be used to generate recombinant bovine adenovirus vectors with insertions or deletions, or both of genes, in the E4 region. The plasmid can be constructed according to the above method (construction of recombinant BAV-1 genomes in E. coli). The homology DNA is derived from the Noü insertion of plasmid 1 055-38 and the adenoviral backbone vector plasmid 990-50 is linearized by partial digestion with Pvu \. Plasmid 1055-47 Plasmid 1055-47 is constructed by inserting a g53 gene of BVDV into the unique Psü site of plasmid 1055-38. The coding region of BVDV is inserted into the complementary complementary arrangement such that it is transcribed by the promoter of the E4 region located at the far right of the genome. The g53 gene of BVDV is isolated according to the method (Cloning of the g53 gene of bovine viral diarrhea virus). Plasmid 1055-56 Plasmid 1055-56 contains a recombinant BAV-1 genome from which the sequence of BAV-1 from positions 29522 to 30407 has been deleted [SEQUENCE OF I NDENTIFICATION NUMBER: 3). The gene for g53 of BVDV (amino acids 1-394) is inserted into the suppressed region. The plasmid can be constructed according to the above method (Construction of the recombinant BAV-1 genomes in E. coli). The homology DNA is derived from the Not \ insert of plasmid 1055-47 and the adenoviral backbone vector plasmid 990-50 is linearized by partial digestion with Pvu \. Plasmid 1055-93 Plasmid 1055-93 contains DNA derived from the E4 region of BAV-1. The sequence coding for nORF1 3 (see Table 1) has been deleted and replaced with a synthetic site Pst \. The plasmid is constructed for the purpose of suppressing a portion of the E4 region of BAV-1. It can also be used to insert foreign DNA into the recombinant BAV-1 genomes. It contains a unique Psfl restriction enzyme site within which foreign DNA can be inserted. The plasmid can be constructed using standard recombinant DNA techniques (see above, molecular biology techniques) by joining DNA fragments from the following sources with the indicated synthetic DNA sequences. Note that fragments derived from BAV-1 DNA are ligated in the orientation indicated by the positions provided for each fragment. The plasmid vector is derived from a restriction fragment Hind \\\ to PvuW of approximately 2774 base pairs from pSP64 (Promega Corporation, Madison, Wl). The synthetic linker sequence 5'-CTGTAGATCTGCGGCCGCGTTTAAACGTCGACAAGCTTCCC-3 '[IDENTIFICATION SEQUENCE NUMBER: 8] is linked to the PvuW site of pSP64 (Promega Corporation, Madison, Wl). Fragment 1 is a PCR fragment of approximately 1282 base pairs containing sequences derived from the genome BAV-1 (positions 28240 to 29522 of SEQUENCE OF IDENTIFICATION NUMBER: 3). Fragment 1 is ligated to the 3 'end of the synthetic linker sequence indicated above (SEQUENCE OF IDENTIFICATION NUMBER: 8). The synthetic linker sequence 5'-CTGCAG-3 '[IDENTIFICATION SEQUENCE NUMBER: 4] is linked to fragment 1. Fragment 2 is a PCR fragment of approximately 1372 base pairs containing the sequences derived from the BAV-1 genome (positions 30403 to 31779 of SEQUENCE OF IDENTIFICATION NUMBER: 3). Fragment 2 is ligated to the 3 'end of the synthetic linker sequence 5'-CTGCAG-3' [SEQUENCE OF IDENTIFICATION NUMBER: 4]. Fragment 3 is the "F" fragment of BamHI of BAV-1 of approximately 2406 base pairs (positions 31779 to 34185 of the SEQUENCE OF I NDENTIFICATION NUMBER: 3). Fragment 3 is ligated to the 3 'end of fragment 2. The synthetic linker sequence 5'-GACTCTAGGGGCGGGAGTTTAAACGCGGCCGCAGATCTAGCT-3' [IDENTIFICATION SEQUENCE NUMBER: 10] is ligated between fragment 3 and the Hind site pSP64. Promega Corporation, Madison, Wl). Note that the BAV-1 sequences can be cut out of this plasmid via the Noii restriction sites that are located in the flanking synthetic linker sequences. Plasmid 1064-26 Plasmid 1064-26 contains a recombinant BAV-1 genome from which a portion of the E4 region (positions 33613 to 33725 of the SEQUENCE OF I DENTIFICATION NUMBER: 3) has been deleted and replaced by a Synthetic Psü site (5'-CTGCAG-3 ') [SEQUENCE OF IDENTIFICATION NUMBER: 4]. The plasmid can be constructed according to the above method (construction of recombinant BAV-1 genomes in E. coli). The homology DNA is derived from the Noü insert of plasmid 1054-93 and the adenoviral backbone vector plasmid 990-50 is linearized by partial digestion with Pvul.

Plasmid 1066-29 Plasmid 1066-29 is constructed by inserting a g53 gene of BVDV into the unique Psü site of plasmid 1055-93. The coding region of BVDV is inserted in the reverse complementary orientation so that it is transcribed by the promoter of the E4 region that is located at the right end of the genome. The g53 gene of BVDV is isolated according to the above method (Cloning of the g53 gene of bovine viral diarrhea virus). Plasmid 1066-44 Plasmid 1066-44 containing a recombinant BAV-1 genome from which a portion of the E4 region (positions 29523 to 30403 of the SEQUENCE OF I NDENTIFICATION NUMBER: 3) has been deleted and replaced by a Pst \ synthetic site (5'-CTGCAG-3 ') [SEQUENCE OF IDENTIFICATION NUMBER: 4]. This plasmid can be used to generate recombinant bovine adenovirus vectors with insertions or a gene deletion, in the E4 region, or both. The plasmid can be constructed according to the above method (construction of recombinant BAV-1 genomes in E. coli). The homology DNA is derived from the Not? Insert of plasmid 1055-93 and the adenoviral backbone vector plasmid 990-50 is linearized by partial digestion with Pvu.

Plasmid 1066-51 Plasmid 1066-51 contains a recombinant BAV-1 genome from which the sequence of BAV-1 from positions 30403 to 29523 has been deleted [SEQUENCE OF I NDENTIFICATION NUMBER: 3]. The gene for g53 of BVDV (amino acids 1-394) is inserted into the suppressed region. The plasmid can be constructed according to the above method (construction of the recombinant BAV-1 genomes in E. coli). The homology DNA is derived from the Not? Insertion of plasmid 1 066-29 and the plasmid 990-50 from the adenoviral backbone vector is linearized by partial digestion with Pvu ?.

EXAMPLES Example 1: Sequence of BAV-1 By convention, we will refer to the region containing the EcoRI A fragment as the left end of the BAV-1 genome. To complement the previously published restriction map (Maria Benko and B. Harrach, 1990 Hungarian Veterinary Act 38: 281-284) other restriction enzyme sites are defined in the genome of BAV-1 (Pvu \, Sma \). The entire genome is cloned into several large restriction fragments. These include BamH \ "B", "C", "D", "F", EcoRI "A", "E" and Pvu \ "B". The genome is also cloned in its entirety as described above (construction of the recombinant BAV-1 qenomas in E. coli). These clones and 126 different oligonucleotide primers are used according to the method described above (DNA sequencing) to determine a superimposed sequence for the entire BAV-1 genome. This sequence (34,185 base pairs) contains 43 open reading frames (ORFs) initiated in methionine of greater than or equal to 10 amino acids (excluding smaller hosted ORFs). All of the 43 ORFs are compared to the current version of the Genbank protein subset as described in the previous methods (DNA sequencing). Based on the BLAST analysis, 28 of the ORFs (ORF 1 -28) show significant homology with one or more other virus genes. An amount of 15 ORF does not show significant homology with virus genes in the current version of Genbank (nORF 1-15). Table 1 shows that ORFs have a broadly variable homology with the adenovirus genes of several different species.

Table 1 Open reading frame (ORF) of the left end of BAV-1 ORF * position ** Best match% with Genbank *** similarity ORF1 1400, 1867 BAV-2 E1A 49.5% ORF2 2189, 2656 BAV-2 E1 B 58.6% ORF3 2566, 3777 SAV-3 E1 B 42.3% ORF4 3838, 4185 BAV-2 hexanone 36.9% ORF5 4197, 5315 HuAd-7 protein of 69.6% RC maturation ORF6 5285, 8530 BAV-2 polymerase 72.9% RC ORF7 6255, 6680 HuAd-7 protein 59.4% unknown ORF8C 8527, 10185 BAV-2 protein 71.2% terminal ORF9 10376, 11437 CAV-1 Orf9 63.0% ORF10 1 1465, 13174 CAV-2 protein of 57.0% hexon ORF1 1 13235, 14662 SAV-3 protein of 74.6% penton base ORF12 14725, 15207 BAV-2 protein of 37.1% main nucleus ORF13 15267, 16388 BAV-2 protein of 62.2% minor nucleus ORF14 16703, 17113 CAV-1 protein of 60.5% minor capsid ORF15 17509, 20238 CAV-1 protein 2 75.3% Hexone late ORF16 20241, 20864 BAV-2 82.6% endoprotease ORF17 20906,22246 OvAV protein 77.8% RC DNA binding ORF18 22258, 24498 BAV-3 protein 59.0% late 100 kd ORF19 24212, 24796 BAV-3 protein 44.0% delayed 33 kd ORF20 25009, 25680 BAV-1 protein 97.8% hexon ORF21 25673, 26041 BAV-1 protein E3 87.7% of 12.5 kd ORF22 25923, 27287 BAV-1 protein 85.8% unknown ORF23 27483, 29294 HuAd-12 protein 31.2 % fiber ORF24 29311, 29730 HuAd-40 protein 38.2% RC E4 ORF25 30404, 30739 HuAd-12 protein 38.5% RC unknown ORF26 30730, 31464 HuAd-40 protein 28.9% RC E4 30 kd ORF27 31471, 32232 HuAd protein E4 40.8 % RC of 34 kd ORF28 32956, 33384 AvAd dUTPasa 54.7% RC nOrfl 278, 736 nOrf2 697, 1167 nOrf3 5634, 5975 nrf4 RC 10301, 10669 nrf5 RC 12607, 1 3212 nrf6 RC 14246, 14722 nrf7 RC 1 5479, 1 6102 nrf8 RC 1 7878, 1 8288 nrf9 19031, 19621 nrfl O 21464, 21 991 nr1 1 RC 24437, 24820 nOrf12 RC 27800, 28174 nrf13 RC 29523, 30407 nrf14 RC 3221 9, 32557 nr † 15 RC 33438, 33908 * RC, reverse complement Apostles in the SEQUENCE OF I DENTIFICATION NUMBER: 3 *** AvAd, avian adenovirus; HuAd human adenovirus; CAV, canine adenovirus, SAV, porcine adenovirus; OvAd adenovirus ovine. The E3 and E4 gene regions of BAV-1 can be defined by homology with genes from the corresponding regions of human adenoviruses. Evans et al., (Virology 244-1 73-1 85) define the region of the E3 gene of BAV-1 as bound by the sequence of the TATA box at positions 25362 to 25365 and the polyadenylation signal in the positions 27291 to 27296. The analysis of the gene homologies of Table 1 indicates that the E4 region of BAV-1 is linked by the polyadenylation signal at positions 29059 to 29065 and the sequence of the TATA box at positions 34171 to 34174 BAV-1 shows a complex sequence organization at its left and right ends. The genome shows an inverted terminal repeat sequence (ITR) of 578 base pairs. A sequence of 419 base pairs is repeated twice at the left end of the genome. A single inverted copy of this repeated sequence occurs at the far right of the genome. The two repeated sequences of 419 base pairs at the left end of the genome are followed by a 424 bp sequence that appears in an inverted copy towards the 5 'end of the 419 base pair sequence at the far right of the genome. The sequence of the BAV-1 genome is useful for the construction of recombinant BAV-1 viral vectors. For example, this information can be used by analogy to human adenovirus vector systems to predict non-essential regions that can be used as gene insertion sites. The information can also be used to predict intergenic regions, which can also be used as gene insertion sites. Example 2: Method of constructing recombinant BAV-1 viral vectors We have developed a novel method for the generation of recombinant bovine adenovirus vectors. This procedure takes advantage of recombinant viral genomes constructed as bacterial plasmids (see construction methods of recombinant BAV-1 genomes in E. coli). When the DNA derived from these bacterial plasmids is transferred to the appropriate cells (see methods - transfection of BAV-1 DNA). recombinant bovine adenovirus vectors are generated. This procedure is exemplified by the infectivity of plasmid 990-50. The DNA derived from this plasmid is transfected as described above in MDBK cells. Progeny viruses recovered from independent transfection accumulations are amplified in MDBK cells and analyzed for growth characteristics, yields of virus production and DNA restriction patterns. In all cases, plasmid 990-50 derived from adenovirus (S-BAV-002) is not differentiable from wild-type BAV-1. This method can be used to generate bovine adenovirus vectors expressing useful foreign DNA sequences. The method can also be used to suppress genomic sequences of the bovine adenovirus vector. The production of bovine adenovirus vectors displaying the expression cassette of the E2 glycoprotein (g53) of bovine diarrhea virus (BVDV) and deletions in the E4 and E3 regions of BAV-1 respectively, are described as follows ( see examples 4-6).

Example 3: Preparation of the recombinant adenovirus vector S-BAV-003 S-BAV-003 is a BAV-1 virus that has a deletion in the E4 region of the genome. This deletion encompasses the H and G fragments of EcoRI. This deletion removes all or a major portion of the ORF 25-27 and nORF13. A polylinker sequence (GAATTCGAGCTCGCCCGGGCGAGCTCGAATCC) [SEQUENCE OF IDENTIFICATION NUMBER: 15) containing a Smal site is inserted into the deletion. Since Smal is absent from the genomic DNA of BAV-1 (see Figure 1), the introduction of this polylinker sequence generates a useful Smal site that can be used directly to engineer the virus. S-BAV-003 is created by transfection of DNA derived from plasmid 1004-73.16.14 according to the method described above (method of constructing recombinant BAV-1 viral vectors). The resulting viruses are purified according to the above method (plaque purification of the recombinant constructs). The progeny viruses derived from the accumulated independent transfection are amplified in MDBK cells and analyzed to determine the DNA restriction patterns of BamHI, EcoRI and Smal. This analysis indicates that the G and H fragments of EcoRI have been deleted and that the Smal site be introduced into the genome. It has also been shown that S-BAV-003 grows to similar titers as wild type BAV-1.

Example 4: Preparation of recombinant adenovirus vector S-BAV-004 S-BAV-004 is a BAV-1 virus that has a deletion in the E4 region of the genome. This deletion encompasses the H and G fragments of EcoRI. This deletion removes all or a large portion of the ORF 25-27 and nORF13. The gene for bovine viral diarrhea virus (BVDV) glycoprotein 53 (g53) (amino acids 1-394) under the control of the HCMV immediate early promoter is inserted into the suppressed region. S-BAV-004 is generated by transfection of DNA derived from plasmid 1004-40 according to the method described above (method of constructing recombinant BAV-1 viral vectors). The resulting viruses are purified according to the above method (plaque purification of the recombinant constructs). Example 5: Preparation of the recombinant adenovirus vector S-BAV-005 S-BAV-005 is a BAV-1 virus that has a deletion in the E3 region of the genome. The smallest subfragment Sa / I to BamHI of fragment "B" of 8amHI (positions 25664 to 26850 of SEQUENCE OF I DENTIFICATION NUMBER: 3) has been deleted. This deletion removes a larger portion of the ORFs 21 and 22. A polylinker sequence (5'-TCGACAAGCTTCCC-3 ') [SEQUENCE OF IDENTIFICATION NUMBER 16] containing the Hind site \\ is inserted into the deletion. S-BAV-005 is generated by transfection of DNA derived from plasmid 1018-75 according to the method described above (method of constructing recombinant BAV-1 viral vectors). The resulting viruses are purified according to the above method (plaque purification of the recombinant constructs). Example 6: Preparation of the recombinant adenovirus vector S-BAV-006 S-BAV-006 is a BAV-1 virus that has a deletion in the E3 region of the genome. The subfragment Sa / I to smaller fiamHI of fragment B of Bam) (positions 25664 to 26850 of SEQUENCE OF IDENTIFICATION NUMBER: 3) has been deleted. This deletion removes a larger portion of the ORFs 21 and 22. The gene for gDV of BVDV (amino acids 1-394) under the control of the immediate early promoter of HCMV is inserted into the suppressed region. S-BAV-005 is generated by transfection of DNA derived from plasmid 1018-42 according to the method described above (method of construction of recombinant viral vectors BAV-1). The resulting viruses are purified according to the above method (plaque purification of the recombinant constructs). Example 7: Preparation of the recombinant adenovirus vector S-BAV-007 S-BAV-007 is a BAV-1 virus which has a deletion in the E3 region of the genome. The smallest EcoRI to Bam subfragment of fragment "B" of BamHI (positions 25765 to 26850 of SEQUENCE OF IDENTIFICATION NUMBER: 3) has been deleted. This deletion removes a larger portion of the ORFs 21 and 22. A polylinker sequence (5'-TCGACAAGCTTCCC-3 ') [IDENTIFICATION SEQUENCE NUMBER 16] containing the Hind site \\\ has been inserted into the deletion. S-BAV-007 is generated by transfection of DNA derived from plasmid 1018-45 according to the method described above (method of constructing recombinant BAV-1 viral vectors). The resulting viruses are purified according to the above method (plaque purification of the recombinant constructs). Progeny viruses derived from accumulated independent transfection are amplified in MDBK cells and analyzed to determine the DNA restriction patterns for SamHI, EcoRI and Xbal. It is also shown that S-BAV-007 grows at similar titers compared to wild-type BAV-1. Example 8: Preparation of recombinant adenovirus vector S-BAV-014 S-BAV-014 is a BAV-1 virus that has a deletion in the E3 region of the genome. The smallest subfragment of Eco / RI to BamH \ of fragment B of BamYW (positions 25765 to 26850 of SEQUENCE OF IDENTIFICATION NUMBER: 3) has been deleted. This deletion removes a larger portion of the ORFs 21 and 22. The gene for gDV of BVDV (amino acids 1-394) under the control of the HCMV immediate early promoter is inserted into the suppressed region. S-BAV-014 is generated by transfection of DNA derived from plasmid 1038-16 according to the method described above (method of constructing recombinant BAV-1 viral vectors). The resulting viruses are purified according to the above method (plaque purification of the recombinant constructs). The expression of the g53 gene of BVDV is assayed by the Western blot procedure. S-BAV-014 shows the expression of a corrected size protein with specific reactivity for the BVDV g53 antibody. Example 9: Preparation of recombinant adenovirus vector S-BAV-022 S-BAV-022 is a BAV-1 virus that has a deletion in the E4 region of the genome. This deletion encompasses positions 29523-30407 of IDENTIFICATION SEQUENCE NUMBER: 3. A 5'-CTGCAG-3 'linker sequence containing a PstI site is inserted into the deletion. S-BAV-022 is generated by transfection of DNA derived from plasmid 1055-52 according to the method described above (method for constructing viral vectors of recombinant BAV-1). The resulting viruses are purified according to the above method (plaque purification of the recombinant constructs). The progeny viruses derived from the independent transfection concentrates are amplified in MDBK cells and analyzed to determine the DNA restriction patterns for SamHI, EcoRI and X £ >al. It has also been shown that S-BAV-022 grows to similar titers as wild type BAV-1. Example 10: Preparation of recombinant adenovirus vector S-BAV-023 S-BAV-023 is a BAV-1 virus that has a deletion in the E4 region of the genome. This deletion encompasses positions 29523-30407 of IDENTIFICATION SEQUENCE NUMBER: 3. The gene for gDV of BVDV (amino acids 1-394) is inserted into the suppressed region. The gDV gene of BVDV is under the control of the E4 promoter or promoters. S-BAV-023 is generated by transfection of DNA derived from plasmid 1055-56 according to the method described above (method for constructing recombinant BAV-1 viral vectors). The resulting viruses are purified according to the above method (plaque purification of the recombinant constructs). The expression of the g53 gene of BVDV is assayed by the Western blot procedure. S-BAV-006 shows the expression of a correct size protein with specific reactivity to the gDV antibody of BVDV. The expression of the foreign antigen g53 BVDV in S-BAV-023 establishes the utility of the promoter of the E4 region of BAV-1 for the transcription of foreign genes in vector systems. Example 11: Preparation of recombinant adenovirus vector S-BAV-Q25 S-BAV-025 is a BAV-1 virus that has a deletion in the E4 region of the genome. This deletion encompasses positions 33614-33725 of IDENTIFICATION SEQUENCE NUMBER: 3. A linker sequence 5'-CTGCAG-3 'containing a PstI site is inserted into the deletion. S-BAV-025 is generated by transfection of DNA derived from plasmid 1064-26 according to the method described above (method for constructing viral vectors of recombinant BAV-1). The resulting viruses are purified according to the above method (plaque purification of the recombinant constructs). The progeny viruses derived from the accumulated independent transfection are amplified in MDBK cells and analyzed for DNA restriction patterns for SamHI, EcoRI and Psfll. It is also shown that S-BAV-025 grows at similar titers as wild-type BAV-1. Example 12: Preparation of recombinant adenovirus vector S-BAV-026 S-BAV-026 is a BAV-1 virus that has a deletion in the E4 region of the genome. This deletion encompasses positions 29523-30403 of the IDENTIFICATION SEQUENCE NUMBER: 3. A linker sequence 5'-CTGCAG-3 'containing a Pstl site is inserted into the deletion. S-BAV-026 is generated by transfection of DNA derived from plasmid 1 066-44 according to the method described above (method for constructing viral vectors of recombinant BAV-1). The resulting viruses are purified according to the above method (plaque purification of the recombinant constructs). The progeny viruses derived from the independent transfection concentrates are amplified in MDBK cells and analyzed for the DNA restriction standards for Bam tt, EcoRI and Xba It is also shown that S-BAV-026 grows at similar titers as BAV-1 wild type. Example 1 3: Preparation of recombinant adenovirus vector S-BAV-027 S-BAV-027 is a BAV-1 virus that has a deletion in the E4 region of the genome. This deletion encompasses positions 29523-30403 of the NUMBER EFFECTIVE SEQUENCE SECTION: 3. The gene for BVDV g53 (amino acids 1-394) is inserted into the suppressed region. The g53 gene of BVDV is under the control of the E4 promoters. S-BAV-027 is generated by transfection of DNA derived from plasmid 1066-51 according to the method described above (method for constructing recombinant BAV-1 viral vectors). The resulting viruses are purified according to the above method (plaque purification of the recombinant constructs).

Example 14: Vaccine against pasteurellosis The pasteurellosis or bovine respiratory disease complex (BRD) manifests as a result of a combination of infectious diseases of livestock and factors related to additional stress (CA Hjerpe, The Bovine Respiratory Disease Complex. Veterinary Therapy 2: Food Animal Practice, Ed. By J.L. Howard, Philadelphia, WB Saunders Co., 1986, pp 670-680). Respiratory virus infections, increased by pathophysiological effects of stress, alter the susceptibility of livestock to Pasteurella organisms that are normally present in the upper respiratory tract, by many of the mechanisms. The control of viral infections that initiate BRD as well as the control of terminal bacterial pneumonia is essential to avoid the syndrome of the disease (F. Fenner, et al., "Mechanisms of Disease Production: Acute Infections", Veterinary Virology Academic Press, Inc., Orlando, Florida, 1987, pp. 183-202). The major infectious diseases contributing to BRD are: infectious bovine rhinotracheitis virus parainfluenza virus type 3, bovine viral diarrhea virus, bovine respiratory syncytial virus and Pasteurella haemolytica (F. Fenner, et al., "Mechanisms of Disease Production: Acute Infections ", Veterinary Virology, Academic Press, I nc., Orlando Florida, 1987, pp 1 83-202). An extension of this approach is to combine vaccines in a way that controls the distribution of disease pathogens with a single immunization. For this purpose, the mixing of several antigens with vector BAV-1 (IBR, BRSV, PI-3, BVDV and P. Haemolytica) in a single dose of vaccine. In addition, conventionally derived vaccines (destroyed viruses, inactivated bacterins and modified live viruses) can be included as part of the BRD vaccine formulation such that the vaccine components provide more effective. Many modifications and variations of this invention can be made without departing from its spirit and scope, as will be apparent to those skilled in the art. The specific embodiments described herein are offered by way of example only and the invention is limited only by the terms of the appended claims, together with the full scope of equivalents for which such claims are entitled.

SEQUENCE LIST < 110 > Schering-Plow Veterinary Corporation < 120 > Recombinant adenoviruses and novel mutants < 130 > SY0993K PCT < 140 > < 141 > < 150 > US 09 / 289,930 < 151 > 1999-04-09 < 160 > 17 < 170 > Patentln Ver. 2.1 < 210 > 1 < 211 > 42 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: DNA primer < 400 > 1 cttggatcct catccatact gagtccctga ggccttctgt te 42 < 210 > 2 < 211 > 31 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: DNA primer < 400 > 2 catagatett gtggtgctgt ccgacttcgc to 31 < 210 > 3 < 211 > 34185 < 212 > DNA < 213 > Bovine adenovirus type 1 < 400 > 3 catcatcaat aatatacgga acacttttgc gtgatgacgt tgacgtgctg tgcgtaaggg 60 ggcgtgggaa aattgttcaa aggtcgctgg gcgggagttt ctgggagggg cggggagtgt 120 ccgtgtgcgt gcgagcggcg gggcgaggcg ctgagtcagg tgtatttatg atggggtgtt 180 tagggtatca gctgttggac tttgactttc actgtcgtaa atttgccact ttattggagc 240 cctttgcccg cggacgtgga gggatttttc ccaatttatg gccactttta ctgcgatgcc 300 gcacgaaacc tgtctaaagg tctatgccac gtctcccatc atgggcggtc ttttttctct 360 atgcaccact cccagtgagc tatatattac ctgcgcaggt aaagaggtgc cactcttgac 420 atcatcaata atatacggaa cacttttgcg tgatgacgtt gacgtgctgt gcgtaagggg 480 attgttcaaa gcgtgggaaa ggtcgctggg cgggagtttc tgggaggggc ggggagtgtc 540 cgtgtgcgtg cgagcggcgg ggcgaggcgc tgagtcaggt gtatttatga tggggtgttt 600 agggtatcag ctgttggact ttgactttca ctgtcgtaaa tttgccactt tattggagcc 660 ctttgcccgc ggacgtggag ggatttttcc caatttatgg ccacttttac tgcgatgccg 720 cacgaaacct gtctaaaggt ctatgccacg tctcccatca tgggcggtct tttttctcta 780 tgcaccactc ccagtgagct atatattacc tgcgcaggta aagaggtgcc actcttgaca 840 tcatcaataa tatacggaac acttttgcgt gatgacgttg acgtgctgtg cgtaaggggg 900 cgtgggaaaa ttgttcaaag gtcgctgggc gggagtttct gggaggggcg gggagtgtcc 960 gtgtgcgtgc gagcggcggg gcgaggcgct gagtcactgc ccttttgcac tgtcttgtgc 1020 tttgtcacgc ggtttcggtt acgcctgtca ggcgccagaa gccctttcgc ctcgtcacag 1080 accgcgcctt ttcgctctat aaagccattt ctctcctctg ctcgtcattc gcctctgctc 1140 tgtgctgcca ctgagccttc tttctaaact tacactcctt gctgtaggcc gtggtctact 1200 ttgccaagag taagtacatc atggctgaca agctgctctt tgtgcgtgtg tctgactcgg 1260 cttgccacgt ctcccatcat gggcggtctt ttttctctat gcaccactcc cagtgagcta 1320 tatattacct gcgcaggtaa agaggtgcca gaagagagta ctcttgagtc gagttttctc 1380 atctgctcat tcattcacca tgaggcacct aagactcgct tttgatgagc gcttctggat 1440 agccgccgaa ggtttgctgg cggattctcc tgctgatgaa gatgagggat ttcatgagcc 1500 caggacttga tttgtctttg ttgaaattga tgacgcttca gacgtggtta gcttattttt 1560 ccctgaactt gaagttcagc aagacctgcc aacagcggag gaggttgagg acttgttaca 1620 ctgtgaggag actgctgctg acttagaatc tgtttctgac ttaccgcctg tggagtctcc 1680 tgaacctccg gattctcact tttccacatt tgagttggat tatcctgaga tacccggcgt 1740 gaattgctct gcatgctcat ttcatcgtca ggagactgga tctgaggagg ctgtatgctc 1800 gctctgctat atgagaaaaa cggcttatgc tgtatatggt aggttgcttt acatactttt 1860 cttttgatta ctcttgcatt gcaatattag cctaatgtgt tgatttgtgc ttgcagagcc 1920 gctccgccta tgtttctcca ctgttgatga gcaacatgag acaggtgcgc cagtttcgtc 1980 tccgcctgct ggccgcaagc ggcgccacca ggatgacctc atactgttta accataaacg 2040 ctgcgcccag gatgaacctt tggacttgtc cttacccaag cccaatgccc aataaactat 2100 gttaatcagt acctagaaag gtgtggtcac tgcctatata aactagggag cgctgctcag 2160 atgcagcctg gcgctcactt ggactaccat ggacatttct ctgggctttt gcgaaaagct 2220 gtccgatttc caatacttaa gacgggtgct gtactacgcc tcagccagac caggttggtg 2280 gacacgcact ctctgtgggg acagactctc aagtttggtt tataatacaa aaattgagca 2340 ttggaaaaac ttagaggaaa tttttaaacg cgattc AGGT ttctggtcta tgctttctag 2400 tgggcgcagc cttgggtttg aggcgaaagt agttccttgg ctggattttt cgtctccagg 2460 gagaactgtg gccagtttat cgttactaac ttatattgtt gatactttgg ataaacagac 2520 tcagctgagc ccagattaca ttttggactc gatttgcggc ccagtatgtt tcaggctgaa 2580 tcaatcagga aaccttggtt aaatgcagca agccgtgcgg ggtcaggagg gtccaattat 2640 agaggaagtg gattaaaccc agacggtata ccataccagt ctatattgat ggagtttgct 2700 agagatccat tttctactca tgacaaatat gattttgaga cagttcagac ttactttctt 2760 aagccagggg atgatttaga aacagtgatc agccagcatg ctaaaattgc tttagatcct 2820 atgtgattga gaggtagagt acatccagta aagattcgat ctctgtgtta tataattggc 2880 aatggagcta aaataaaaat agcatgtcca gaacactttg gaatagaaat ttatccaaga 2940 gatcacagcc ctggtatagt tggaatgtgg cttgtcacat ttaacaatgt ggtgtttgaa 3000 agggaacgaa gtattcctgg tggcattatt cagagtagga cattcttttt gtgccatggc 3060 tgtaatttct tgggagcatt gggaacagct gtttcggctc tggctggtgg ggaagttagg 3120 gggtgccatt tctttggatg tttcaagtgt gttgatagta gaagtaaatt taaggtgaaa 3180 gttagccact ctgtaattga gacttgtatg gtaggcataa g cgcttcagg gccagtgagt 3240 gtaaagcatt gtcaaggatt gagtgtgtac tgctttttgt ttatgttagg agctggtaag 3300 gttgagggaa acagtgtgat caacccaaac aagttttatg agtctgccct tacagagatg 3360 gtgtcttgct atggcaaaat tgttttgccc ctggccactg ttcatatttc ggcctcgccg 3420 aaacatcagt atccacactt tgaagcaaat gtgctgacaa gatgtaaggt gtttgtgggc 3480 gccaggcaag gcacttttac gccaatgctt tcatcactaa gttatacttc tattgtggct 3540 gaccgcgatg ctttcaaaag cctgaatctg aattatactt ttcaccaaac tactactatt 3600 tggaagcttc tgagtgctgc tgatgctgac tttgaccatg gaactgccag aaagtgcctg 3660 tgtggtgatt tgcatccatg tcctgtgttg aagcagcttg attacacaag ccgggttagg 3720 cctaacccat acgaccactc atgtgattcc agggtgtttt ctgatgacga gaattaaggt 3780 aagccacgcc caccatctat ataagcggga gtaaaagcgt ggggtggtat ttgcacaatg 3840 actgatcaag gtgacattcg tacgtgtttt cttacagcga gactgcccag gtgggcaggt 3900 gttcgccaaa atgtcgtcgg gtcaaatatt tctggtggcg ttgtcgactc cccggagacg 3960 cttctggcat ctagatctaa cgcggcagct gcgatgatga ctttgaggaa catcgcgacc 4020 agcagacagt tggaagagca ggtggagact ttgctggagc agaact tgga tctgacggcc 4080 cagcttaatg ccttgctgat gcgtgtcaac gcgattgaac ggcagctagc tgatatgcag 4140 cgcgacttgg aaccaatcat tcaacaacac aatgcaataa tttgatcaat aaatctttat 4200 ttctttgcat gataatatcg agtccagcgt tgtctgtcag caattacttt gctaattttt 4260 tccaaaatag agtacagttt tttagataca acattgcaca ttggtataag tccttctgat 4320 gggtgtaggt atgaccactg tagggcttca ttttctggac atgtattata aattatccag 4380 tcatagttgg tgttaatttt gtgatagttg aatatgtctt ttagcaggag ggaaattggc 4440 ctttagtgta aatggcagtc ttgatttata aatctattaa gctgtgaagg ctgcatttta 4500 ggagagatga tgtgcagctt tgcttgtatt tttaaatttg atatgttccc agcgtgatct 4560 tttctggggt tcatattgtg caatactacc atgacagagt agcctgtgca ttttggaaac 4620 ttatcatgta gtttagatgg aaatgcgtgg aaaaactttg aaattccttt gtgggctccc 4680 5 agatcctcca tacattcgtc tagtattata gcgattggac cttttgatgc cgctttagca 4740 aatatgtttc tggggtcgct caggtcatag ttgtattcct gcgttaggtc tgaatatgcc 4800 attttggcat atttttatga ctctgtggaa cagcgagcca ctaaggtccc ctgaggcccc 4860 tgccttcaca atgctgtagt gatctgtgtt tcccaagcac ttatctcttg gggg ggtatc 4920 atgtcaattt gcggcactat aaagaaaaca gtttctgggg gcggtgtgat taactgtgag 4980 gaaattatgt tcctaagaag ttgggatttg ccgcagcctg tggggccgta aacaacccct 5040 atgacaggct gcatctgaaa attaatagac ctgcatgccc cttgcgggtt caaataaggt 5100 acgcatttat taagcaactc cctgacacaa acattttctt cagccaaatc taaaagtaaa 5160 ctgtgtccgg ctaatgacat cagttgetgg aaagaggaga acgtgtgaag aggttttagg 5220 ccttcagcaa aaggcatgct ttttaagctg ttgtgcaaga cggtcagtct gtcccatagt 5280 tcatttatat gtcccacggt aatgtcatcc agcatttgtc gccgtttctt ggatttgggt 5340 tgcttttgga gtagggcatt agtcgatgct ggtcgaggtt tacgagggtt ctgtccttcc 5400 10 acggcctcac tgtccgagtc agagttgtct ctgtcactgt gaatggagcg gcgtttgctt 5460 ggtttgtcgc tagagtgcgc ttcaggctca tccggctggt ctgaaagtgg tctgagccgt 5520 gctggctgtc cgctaggtag gagtatcata cactgggcga agatagctca gtggttgcgt 5580 gccctttagc tcttagcttg cgtgaccgca cctttcccca gttagggcag tgtatgcttt 5640 tcaacgcata cagctttggc gccaggaata cagattcagg actgtatgca tcactcttgc 5700 atttttcgca ctgcgtttcg cattctacta gccaggtgat gcttgggcag cttgggtcaa 5760 acactagact tcctccattt tttttaattc tatgtttacc tttttcttgc attaggcgat 5820 gtccttcttc tgtcacgaac aggctgtcgg tgtcaccgta cacggacttg attgtgcgtt 5880 gttccatggg ctttcccctg tcgtcgctgt acaggaactc ggcccactcc gccacaaaca 5940 agctagcaca ctcttgtcca aaagaagcga tgtgagaagc gtaccggttg tttttgataa 6000 gtaaagaatt ttgctcgagg gtgtgtaaac aaatgtcatc gtcatcggtg tccatgaatg 6060 tgattggctt gtaagtgtag gtcacgtgac ccgccgtagg tataaaaggg gcggggtcct 6120 -J cgtcttcctc atttgcttct ggctcgacgt gcggtgcagg tgggtaggct acggtaaatt 6180 ctggcataag ttcagcactt aagttgtcgg tttcaatgaa agaagaggat ttgacactgt 6240 aggtgccagt ggcgatgttt tttgacattt ctgattcaag ctggtcagaa aacactattt 6300 ttttgttatc gagtttagta gcaaagctgc cgtacagagc atttgacaac agtttggcta 6360 tgctgcgcat tgtttggttt ttgtttttgt ctgctttttc tttggcggct atgttcagct 6420 ggacatattc tttagccacg caacgccatt ctggaaatat tgttgttctt tcatctggta 6480 gtatgcgcac tttccagcct ctgttgtgca gggttatcat gtctactgat gtggcaacct 6540 caccgcggag aggctcgttt gtccagcata gcctgcctcc cttcctggag cagaagggcg 6600 ggagctcgtc caggaagagt tcgtctggag ggtcggcgtc cactgtgaag atacctggca 6660 acagcgtgtc atcgaaataa tcaatgcgcg aaccatgcgc gctcaacctt ctgctccagt 6720 ctgatgcagc aactgcgcgc tcgaatgggt tcagcggctg ccccgctgga aatggatgag 6780 tcaatgcgct tgcatacatt ccacagatgt catacacata aataggctgt tccagtatgc 6840 Q cgatgtatgt ggggtagcag cgtcccccac ggatgctttg gcgaacgtaa tcatacatct 6900 cgtttgacgg cgcaagcagg gtgtttgaca tgttggaacg gttaggtttg attgagcgat 6960 a caaaatttg tttgaagatt gcatgggagt ttgagctaat tgttggtctt tgaaaaatgt 7020 tgaatgctgc ttcaggtaag tcaacttctt tctgaatgaa ctgctggtat gagttttgga 7080 gttttctgac cagctcagag gtgactaaaa catcttgggc gcaatattca agcgtttgct 7140 ggatgatatc gtaagccccc acgttttttt ctctccacag cgctttgttg agctcgtatt 7200 ctgctgtgtc cttccagtac cgaaggtgtg ggaaaccatc ctcgtcctgc tggtaagagc 7260 ccaggcgata aaattcgttt accgcttcgt acggacagct tcccttttct actggaagtt 7320 catacgccga tgcggcattt ttcaagcttg tgtgtgtcaa cgcaaatgtg tctctgacca 7380 tgaacttaac gaactgcatt ttgtagtctc ctgctgtcat ttttcccagt tcccagtcct 7440 cctggaaaca caaatgtttt aactttgggt ttggaagtcc aaatgtgatg tcattaaata 7500 aaatctttcc atttcttggc ataaagtttc tacttatttt aaatgctgga aggacctcat 7560 aatcacttga ctctgttgtg gccgccagca cgatttcatc gaaaccgcag atgttatgtc 7620 tatttccata ccaccacata aacttagggt ctccgtttag ttgacacttt ctaagcatct 7680 caaatgtgat atcatctact gcagctaggc catgttgctc tttaagctgt tccagatgtg 7740 ggttgtgagc gagaaggtgc tcccatagta tggcggtcgt gcgccgctgt acggcgtcgc 7800 ggaact tttt aaaagctctt cccacctctc cctttgttgg ggtgatgata tagtaagtgt 7860 acttttcgcg ccacgctgtc cactctaggt ttatggcgac attattcgcg gcttctagca 7920 gctcgctgtc cccagatagg tgcatcacta gcataaatgg caccagttgt ttgccaaagc 7980 gcccgtgcca ggtgtaagtc tccacgtcgt atgtgatgaa atgtcaacgt cagcctttga 8040 acgagcctat tggctgaaat gggataagtt cccaccagtc ggcagattta tgattgacgt 8100 ggtgaaagta aaaatcacgc ctccgcacag aacaagtgtg agaatgtttg taaaagtctc 8160 cacagaaatc acatttttgc gagggtgaaa tttcttttat taaaaacacc ttcccatgtt 8220 tgacgaagaa attaattgaa aaaggtagga tgctttcttc catgattgtg gacttttttg 8280 aaattcttcc tttgttgtag ataaaaacac ctccgtcact cggtagtagg ctttctaaca 8340 ttgcgagggc gtttcgtgga gtgaccgctg cggcgttaaa gtgatcaggc atgtcaaata 8400 gatgaatgtg gaaaaggttt gcaagtgctg gctgtagtgc gctgtggtat ttgatttcga 8460 cgtgggtccc atcctccatt gtcccactgc ttgttagcgt ggcgcgtttg gccactactg 8520 tgcctctcat tgctcttccg gcggcggaag gcgcactgct tcgtttagcg ccggcagtgg 8580 tgacgcaaca acgccgttct ttgttgctgc gcgtatcact cgtcggttta tattctggat 8640 ttccctcagg c cggagaaca ccacaggacc gctcactcga aacctgaaag atatttcgat 8700 agaatcaatt tcagaatcat tggtggccac ctgtcttaga atttctgtta catcgccgct 8760 gttttcgtga tatgctattt ctgccataaa ttgttctatt tcctcctcct cgagctctcc 8820 tctgccagcg cgctcaacgg tggctgccaa atcaacactt attctgttca taatagcaga 8880 aaacgcttgc tcgccgtttt cgttccagac gcgactgtag accagcctgc cgtcctgaga 8940 ccttgctctc atgaccactt gtgccagatt tagcatgacg tatcttccga atgggctcgc 9000 gactctcagt tgatgattta agtagttgag agtggtggcg atgtgctccg ccacgaagaa 9060 atacatcacc catcgcctga gtgtcagctc gttgatatcc ccaagcgctt ctaaacgctg 9120 tataacttcg tagaagttca cagcaaaact gaaaaactgc tgatttctgg ccgcaaccgt 9180 cagctcttct tcaagcaatc tgattgcttc agccactgcc gctctaactt cttcttcaaa 9240 cgtagtctca gggctttctt cctcaacttc cattggcgcc tcttccggtg gtggaggcgg 9300 ctgtcttcgg cgccgtctgc gcatcggaag acggtccacg aactgttcta tcatttctcc 9360 tctggctctt ctcatgcttt ctgtgactgc ccggtttcct tctcttggtc gtagctggaa 9420 agcgcctcca ctcatggctg tgccgtggca actggggagg ctcagtgcgc taataataca 9480 ttttgtcaat atttgcgcag gaacttgttg cagcctcatt gcttcgctga tatcggcaga 9540 ttgatcgctt tcggcgaact tctccacgaa ggcatttaac caatagcagt cgcaaggtaa 9600 gtttaactct tgctcttggg ctagtg GGAG gtggcggcat attagaaagt tgaaatatgc 9660 tgttttgagc ttgcgaatcg atgacagcac cactaagtct ttgcgcccgg cgttttgcac 9720 tctgattctg tcagccagcc cccaggcttg gccctggcat gcccctatgt ccttgtattg 9780 aagtattcca ttcctggagc cgggaacgtc gtttctatcg actgaggtgc gaccaaatcc 9840 ccgcattggt cggataagag ctaggtctgc tacgatgcgc tctgccagaa tagcctgctg 9900 gacggctgtg agcgtttcag aaaagttgtc catgtctatg aagcgatggt atgccccggt 9960 gttcacagtg tatgagcagt ttgccattac tgaccaattc attatctgcg atccaaagct 10020 aagctgttcg gtgtatttta accggctgta tgctctggcg tcaaaaatgt agtcgttgca 10080 tatctgcaac agcttttgat atccaaccaa aaagtgcggc ggtgggtagt tatataacgg 10140 ccagttccta gtagccggct cccgcggcga tagattcatc agcattaggc ggtgatattg 10200 gtagacgtgt cttgacagcc atccgagccc ggctggtgtg acagcagccc ttgcccaatc 10260 ttggacacgg ttccaaatgt tgcgcactgg cctaaacact tcaattgtgt aaacgctctg 10320 gccggtcagg cgcgcgcagt cgatggcgtt ctaaaagaaa taaacaacat gtccaatggg 10380 atttgtgcag atgcatccgg tgctgcgcca ttgccccata gctgaaacca gtaaggcatc 10440 acgtgggccg gcccgttgta ATGA cgaccc cgcgcctacg ccgcccatcc aggagggaga 10500 gggtgttgcg cgactgaacg tggagagccc cgagcaacac ccccgtgttc agcttaagaa 10560 ggatgccgga gaggctttcg tcccacccgc caatgtattc agagaccggg agggcgaaga 10620 atgaggcaca ggaggctcag tgagatttaa agcgggagaa caaatgcatg tccctaagaa 10680 gcgcgtgcta agtgatactg actttgaagt ggatgaggtg tccggggtga gcccagccaa 10740 ggctcatatg gcggcagccg atctgctgac cgcctatcag caaactgtca gagaggaggt 10800 aagacattta caacttccaa ataataatgt tcgaacactg gtggccaggg aagaagtggc 10860 agtggggctc atgcatttgt gggactttgt tgaggcgtac gttgtaaatc catcttccaa 10920 agctttaact gcccagctat ttcttattgt ccaacactgc cgcgacgaag gcattctaaa 10980 ggaatcgctg ttgaacattg cagagccaga gagcaggtgg ctgttagatc taataaatct 11040 gctccaaacg atagtggttc aagaacgggg catgtccatt acagaaaagg tggccgccat 11100 taactattct gtaataactc tcagcaagca ttatgccagg aaagtttata ggactccgtt 11160 tgtccccatt gacaaggaag caaagatcac cactttttac atgcgaattg tggttaaact 11220 gttggtgttg agcgatgact tgggcatgta tcgtaatgag cgcatggagc gggtagtcag 11280 agacga cgctgcccgc gaat tcacagataa agagctgatg ttcagtttgc gtaaagcgct 11340 ggcaggagaa gacgaggtat atgacggcca attagaatct gctgttcaga gcgtgccagg 11400 tatagaatgg gcgcatgagg atgatgacga cgagtagtaa gatgttatct tggttacagc 11460 cgctcccgca cgccatgttt acaccgtgtc tgcggcgcgc aatcctaacg ccttggcgcg 11520 cctgcagtct caagcgtctg gggacgtgga atgggccgat gccattaagc gtataatggc 11580 tttgaccgcc agatacccgg aagcgttcgc tagtcagcca tttgcaaata ggatcagcgc 11640 tattcttgag gcggtggttc cttctagaaa aaatccgact catgaaaaag tgctgtcaat 11700 tgtcaacgcc ttggtagaaa cgggcgctat tcgtcctgat gagggagggc aggtgtacaa 11760 cgctctgctt gagagggtat ctcgatacaa cagtatgaat gttcagacta gtatagacag 11820 gatgtgagaa gcttagtcaa acgtagttgc tcaaaaggaa aggatggttg gagagaacat 11880 ggggtcgatg gtggccctta atgcattttt gtcaactctg ccggccaatg tggagagagg 11940 gcaggaaaat tacacagctt tcataagcgc tttgaggctg ttggtgtctg aagtgcctca 12000 gactgaagta tatcagtctg ggccaaatta ctacctgcag atggcagtca acctctagga 12060 cactgtcaac ctgactagag cttttgaaaa cctgagctct ttgtggggag tgaatgcgcc 12120 agtggccga to cgaagtgcca tatcttccat tctcactcca aacactaggc tgctgcttct 12180 gcttatagcc ccgtttacag acggggttaa gcttcataca catttccaga ttggttacct 12240 tacagggaaa gctgacccta ggctcatatt ctatcgggca catacaatga gacgaaagaa 12300 gattactagc gtaagccggg ctgttggcaa cgaagacgct gcaaacctgc aggccacatt 12360 gaatttccta ctgacaaatc ggcagtacag gatccctaaa gagtactcat tgacgccaga 12420 ggaggagcga atattacgtt ttgtgcagca gtctgtcagc ctgcatatga tgcaagacgg 12480 cagcacacct tctgccgccc ttgatgaaac aagccgtaat tttgaaccta gcttttatgc 12540 gggaaatagg ttattcatta acaagctgat ggattatttt cacagggctg ccgctgtagc 12600 cccaaactat tttatgaacg cggttctaaa tccaaaatgg ctccctcctg aagggttttt 12660 tactggcgtc tttgattttc ctgagggcga tgacggtttt gtgtgggacg atacagatgt 12720 atctgaggtt ggggcgagag gtgccgttcc ggcgctagtg gccaagaaag agggagggga 12780 tgattcagat ctgtccatca cgatcccctc tattcccagg cagttacgca gggcttctgt 12840 tgtgtctgat actagcgaca tgagccgcgg tagggtgcgc agccgcagtc gtgtacgacg 12900 gccggtagac atagacattg ggcgctggct agaggacaaa aacactaatg cgacccgagc 12960 ctc agctgct attaataacg aaatggaaaa tttagtcgac aagatgacta gatggcgcac 13020 gtatgcccag gagcaaatgg aggaagtcag agcgcgctct cccataaaaa tagaacagga 13080 tgatgatgat tggagaaacg acaggttttt gaagtttgaa ggcagtgggg cagtcaatct 13140 gttcagccac ttaaagccaa aaggcatggt gtaacaaaaa aaaaaaaaaa taaagtcact 13200 taccacagac atggtttggt tttgtgattg ctagatgata cgagccaggc cagtggaatc 13260 gcctcctcct tcctatgaga gcgtggtcgg cactatggat ccgctctacg tgcccccgcg 13320 atacttgggt cctactgaag gaagaagcag catccgttac tccctattgc ccccgcttta 13380 tgacaccacc aagctttact caagtcggca ttatcgataa cactcaatta gatatttcgt 13440 tcaaaataac cacagcaatt acctcaccag tgttgtgcaa aacagcgact acacgccgca 13500 acgcaaacta ggaggctagc taaactttga tgataggtcg cggtgggggg cggactttaa 13560 aactattttg catatgaaca tgcccaacgt gactgaattt atgtttagca attcattcag 13620 atgtctgcca ggctaaattg aggtgggtgg caacccaacc tatgagtggt tcactctcac 13680 cattccagag ggcaactact cagacattgc agtcttagac ttgatgaata atgcgatagt 13740 agaaaattat ctgcaggttg gacgccagaa tggagtagcg taggcgtaaa gaagaagaca 1 3800 agaaatttca gtttgacact gattgggcta tgatcctgta acccagcttg taatgccagg 13860 gaaatatact tatttggctt ttcacccaga catcatactc gcccctggct gtgcggtaga 13920 agccgcctaa ctttacaacg tggtattcga acaatctact catttcagga aaaaggcagc 13980 aggatttcaa atagcctatg aagatttggt aggtggtaat attccagctc tccttgacgt 14040 ggacaactat gatgaggcag acccagccac aattaggcct atagaggccg acccgtcagg 14100 ccgctcatac cacgtaggtc aagacccgtc tgctggtccc acattcacgt attataggag 14160 ttggtacgtg gcttacaact acggtgaccc attcgcagca acagactgga gtacgttgct 14220 ggtgacccct gacgttacgt gtggttcaga gcaagtatac tggagtgttc cggacatgta 14280 gtgacgttta tgtagagcct aagctagcca aaacgtggca aattatcctg taattggggc 14340 agagctcatg cccgttcagt cgcgcagtta ttataacgcg caggctgtgt attcgcaaat 14400 gattcaagaa agcactaatc agacactggt ttttaaccgc tttcccgaca accagatttt 14460 ggtgcggccg cccgaatcta ctatcacgtt cgtcagtgaa aacgtgccag cgcagactga 14520 tcacggaacg ctccccatca gaaacagtgt gtctggggtg cagcgagtca ctctgactga 14580 cgctaggcgc agagccagtc cttacgttta caaaagcata gccatagctc agc caaaggt 14640 tctgtccagc aggacgttct aaaatggcga ttttagtgtc cccaagcaac aacacagggt 14700 gggggattgg atgcaaaagc atgtatggcg gcgcccgcac gctatcagca aactttccag 14760 tgctcgtgcg aaagcactac agggccgtcg tggggaagca ggaaagggcg cgttgtcgca 14820 ccaacagttg aggttacaga cgaccctgtg gccgatgtag tcaacgccat tgctggtcag 14880 acacgccgcc gacgcggagc caggcgccgc aggcgcgcta cggcagcggt gcgcgccgct 14940 agagcgttgg tgcgaaatgc acggcgcacg ctagcccgta gggggcgcat gcggagaacc 15000 cggaatccag tggctgacgt ggtgagagca gtggaagcca tcgcacgcgc aaacccacgc 15060 cgtcgaagcg ctaggttgat ggcgcgtgct gccaacgcac cgcctccacg tccgcgcgcg 15120 aggaatatct attgggtgcg agacagtaat ggagtccgcg ttcctgtgac gtcccgccct 15180 ccaagaactg tggggactgt ggtttaataa agcctcgttt gctgcatcac acagcgcgtg 15240 ctttgtgcca cctgttcgtg cttcgcgaaa acgtcaatgt gataaaagaa gagatgcttg 15300 aaatcgtggc gccagagatc tatgcgccta gacgccggcg tagtgttaaa gttgagacaa 15360 taaggtccca aaacgaggat taaaatctaa aaagatgaaa acgcaagtgg aggcgtcctg 15420 gcatggctga catagatgag gtcgaaatac tgggagccac tgctcc tagg cgcccgtatc 15480 agtggcgcgg taggcgcgta cagcgcatat tgcgtccagg aacggccgtg gtgtttacac 15540 tagtcgggaa cgggcgctcg cgagcaagca agcgttcttc cgacgaaatg tttgcggacg 15600 cagatatact ggaacagttt gaaagtggag atggcgagtt tagatacgga aagcgtggcc 15660 ggtctgaggc gctagtgttg gacgcctcta acccaactcc gagcatgcag cctgtaacgc 15720 cgcaggtacc tatcatgaca ccttcggtgg cagctaagcg cggcgctagc gcagtgccca 15780 actggcgcca cggtgcaagt aagaagcgac gcatagacgc agtagcgaca gacgatgtat 15840 ttgtcgctcc ttctccactt agcgagatgg acaccgtaga gccaggcacg gccgtccttc 15900 ttccttctag agcagttaag cgagttagga agagacgcgg agttgaagaa atcaagagcg 15960 atcctatggt tcttgaagaa gtaaaggtta gggatgtaaa accgatcgct cctgggatag 16020 gcgtgcagac aatagacgtg aaagtgccgg cggctcctcc agaaataaag ccaccagtgt 16080 cagtggtgga gaagatggac ataagcacag ctcccgcgtc acgaatcacc tatgggcccg 16140 ccagcaagat atttccacag taccgacagc atccgagtca aatgggattt ccaaaagtag 16200 ttcgcactcg aaggcgcgcg gttaggagga gacgaaggcg ggcggcgccc attggtgttg 16260 aaattacagc cgcgcgaaga cgggcgctag ttg gcgccgcata cttccgc ctgttcgcta 16320 tcacccgtcc ctgcagacgg cgcctcgctc tcaggtcgca atctggcgtt gatcgatcat 16380 gcgaataaat tcctgtggta ctgcgtttag gcacctatct aacgcgatgg ctggcgtccc 16440 gagaatcacg taccgagtcc gcgtgcccgt gcacacacga gtgcggcgaa gtggaagact 16500 ggcgcggcgc gcgcctcgac gaaggggact taagggcggc tttctacccg ctctaatacc 16560 tatcatagcg gcggcaattg gcgctgcgcc cggcattgca tccgtagcaa tacaggccgc 16620 ccgccgcaaa taaagttagt tactgtctcc aagactcatt gttatcttta tttgcgccag 16680 ctgcctgcct gcgcccgtcg ccatggaagg aattaatttc tccgcgttgg ctcccagatg 16740 cccatgctta cgggtcaaga tgatatcgga gcagttggtc acaagctcca tgaacggcgg 16800 agcatttaac tggggaaacc tatggagcgg cgttaagtcg tttggcagct ccattaaaaa 16860 ctggggcaat cgcgcctgga acagtagcac tgggcaggcg ttgcgccaaa agctgaaaga 16920 cagcaacctg caggaaaagg tggtagaggg gttggctagt ggcattcacg gcgctgtaga 16980 tattgctaac caggagattg ctaaggcggt gcagaagcgc ttagagtcta ggccgaccgt 17040 tcaaatagag gatccagatt taatgtcaac agccgaagaa ctggatcgtg gaaaaaccgg 17100 taaagcgcca ctccgtccac gttaaagcca ctgtag AAGA gtgtagcgaa aaaaccgccc 17160 gtccgacgaa gaagagatag tcattcgtac agaggagccg cccagatacg aagacatttt 17220 ccccaataac tccgcggttc caataagcct gcgccctaca gcggttaggc cgtctgctcc 17280 agtagtcact gtaccggcgg cccgccccgt aaccacggaa attgtagaag ttcctccaac 17340 gagacctagc gctcgtccgg cggtggtgcc ttctagaaca acaagaggat ggcaggggac 17400 gctcaacagc atagtgggcc taggtgttcg atcagtaaaa cgaagacgct gtttttaagc 17460 atctcgctgc tctttccaag cgcgccccag tgatacccgg ccgcgaagat ggcgactcca 17520 tcgatgatgc cccagtggtc gtacatgcac atcgccgggc aggatgcctc agagtacctg 17580 tctcccggcc tggtgcagtt cgcgcaggcc acagagacct actttaagct gggtaacaag 17640 tttagaaacc ccactgtggc tccaacgcat gacgtcacca cagagcggtc acagcggctg 17700 cagctgcgat ttgttccagt tgaccgtgaa gacacgcagt acactcacaa gaccagattt 17760 cagttggctg tgggcgacaa gacatggcga ccgagtactt gcacttactt tgacatccgc 17820 ggtactttgg acagaggtcc aagctttaag ccatacagcg gcacggcata caacgctcta 17880 gcccctaagg ggtctatcaa taacactttc gtatccgtgg ctggaaacaa caacgccaaa 17940 aagcccctca gcttttgcgc gtcggcaa ca gtagacggaa ctacgggcgc catccaaata 18000 gacggcgccg ccatcgacaa cacctaccag ccagaacctc aaataggaga ggaatcttgg 18060 ttgtccggca ctgtgaaccc aatcgcgcag gctaccggaa gaatactgaa tacatctact 18120 gatcccctgc catgttacgg gtcttatgcc gctcctacga acattgaggg agcccaaact 18180 atttgataca cttaacaaca agtgaatttt gtggctggag gcgcgcctgg cgccccagac 18240 ttatggaaga gtaggcatga cgtggctctg caaaccccag acacacattt agtgtacaag 18300 gtgccagccg ccaacgtagg caacacggcg gccttagcgc agcaagctgc gccaaacaga 18360 gcaaactata ttggcttcag agacaatttc atcggtctaa tgtactacaa cagcaatgga 18420 aacctagggg ttttggcggg gcaggcttcg caattgaatg ccgtcgtgga cctgcaagac 18480 agaaatacag agttgtctta ccagcttatg ctcgacaacc tgtatgacag aagccggtat 18540 tttagcattt ggaaccaggc tgtagacagc tatgacccgg atgttaggat aatagagaac 18600 cacggagtgg aagatgaatt gccaaactac tgcttcccaa taagcggaat agttcctggc 18660 accacctcta ctagagtcac cagaaacggt ggaaactggg aagccacggc aaacaacgat 18720 ccggcgtatg tcaacaaagg caatttagac tgtatggaaa taaacctcgc ggctaatctg 18780 tggcgcgggt tcctatattc taatgttgcc ctgtacttgc taagttcaca cagacgacct 18840 ccgccaaatg tcacacttcc taacaacacc aatacgtatg catacatgaa cggtcgcgtt 18900 ccagcggctg ggttggttga cacttacgtc aacattggcg ctcggtggtc gttggatgtg 18960 atggataacg tgaacccatt caaccatcac agaaacgcgg gcctgcgcta ccgctctcaa 19020 atggccggta cttctaggca ctgtcatttt cacatccaag ttccgcagaa gtttttcgcc 19080 atcaagaacc ttcttctgct gcctgggacg tacacttacg aatggtcttt cagaaaagac 19140 gttaacatgg ttcttcagag cactcttggg aatgatctgc gtgtggacgg agcctccatc 19200 acaattgaga gcgttaacct gtatgccagc tttttcccaa tggcacacaa taccgcatcc 19260 actcttgaag ccatgctgcg caatgacaca aacgaccaat cgttcatcga ctacctgtct 19320 tcagccaaca tgttgtatcc aattcctgcc aatgccacta acctgccaat ttccatccca 19380 tctcgcaact gggccgcctt ccgcggatgg agcttcacca gaaagaaaca gaattaagca 19440 cccgccttgg gctctccatt cgacccctac ttcacatact caggcactat accatacttg 19500 gacggcacct tttatctcaa tcacaccttc agaagggtgt ctatacagtt tgattcgtcg 19560 gtgcagtggc cgggcaacga ccgcttgctc acaccaaatg agtttgagat taaaaggcta 19620 gtggatggag aggggtacaa tgt agctcag agcaacatga caaaggactg gtttctagtg 19680 caaattacaa cagatgcttg cattggctac cagggctatc tggctataaa atctcccgga 19740 gatcgcacat attcttttct gagaaacttt cagccaatga ctaggcagat agtggaccaa 19800 ccgcgtatca actaacgtgc gaatgtccca atcacccacc agcacaataa ttctggcttt 19860 actggatttg ccagtccagc gctgccgcgt gagggacacc cgtatccagc taactggccg 19920 tatccactga ttagcgctac tgcagtggcc acgcaaacac agcgaaagtt cctatgtgac 19980 aggacgctgt ggcgcattcc attctcgtcc aactttatgt ctatgggatc gcttaccgat 20040 acctgctgta ctggggcaga tgcaaatgct gctcacgcct tagacatgac ctttgaagtg 20100 gacgcgatgg acgagcccac gctgctttat gttttatttg aagtgtttga cgtggttcgc 20160 gttcaccagc ctcacagggg agtcatcgaa actgtctacc tcagaactcc attctctgcc 20220 ctacataagc ggcaacgcca atgggatcca gggaagagga actgcgcgcc attgtgcgcg 20280 acctcggagt tgggccatac ttcctgggga cgttcgacaa acgctttcct ggttttctaa 20340 ataactcaaa gccgagctgc gccatcgtga ataccgcagg tagagaaaca ggcggcgcgc 20400 attggctggc cctggcttgg ttccctaaat ctaaggcttt ttactttttt gatccatttg 20460 tagcaa gattcagtga actg aagcagatat atgagtttga gtatgaaggt ctgctgcgcc 20520 gcagcgcctt ggcggctact ggcgatggct gcataaacct ggttaagagc agtgaatcgg 20580 tacagggtcc gaacagcgcc gcctgtggct tattttgctg catgttttta catgcttttg 20640 ctcactggcc ccacagtcct atgacccaca accccaccat ggacttgttg actggtgtgc 20700 cattatgtca ctaaccataa cctagcgccc agcccacact gcgagaaaat caagtcaagc 20760 tttataagtt tctagcagcc cattctcagt actttcgcac ccatcgcccc caaattgaac 20820 ttttaataaa gagacacctc caaaattgca ctgctggaat ttattttgaa ataaatgatt 20880 tcaacatttg agcagcgtgg tgtgttcaaa ataacgcgtc gtcggcgtct tcctgaccgg 20940 tgggtaggat ggtgttctgc actctgtact ggggaagcca cttaaattct tgcacgacaa 21000 tgggcggttt cgtgccaacc attgaattcc agatttgctt tgcgagctgc agccccatga 21060 ctacatctgt cgagctgatc ttaaagtcgc aattcttctg agggtttgct ttggtattgc 21120 gaaatacagg gttgcagcac tgaaatacaa gcactgcagg gtggtctagg gtggccaaca 21180 ccttagcgtc gtcaatcaag gcgcgatctá tgctgttgag tgcagtcatc gcgaacggcg 21240 tgaccttgca cgtctgcttt ccaagcaggg gtagaggctg atgaccgtag ttacaatcac 21300 ataccag cgg cattaagagc atctcaccag cttttggcat gttgggatac atcgccttta 21360 caaaagcgcc tatctgcttg aaggccatca gcgccttggg gccatctgtg taaaaatacc 21420 cacaagactg agagctaaaa ctgttgattg gagactttag atcatgatag caactcatcg 21480 cgtcgctatt cttgacttga accacgctgc ggccccagcg gttggtgaca atcttcgcgc 21540 gctcaggtgt atccttcaat gctcgctggc cattttcgct gttaatgtcc atctcaatga 21600 tctgctcctt gtttatcatg ggcaagccgt gcaaacaata caatttgtcc tcgtctgcct 21660 cacaacacaa tgtgctccca ccagatgggt tccaatctgc cgccgttata tcggcgccgc 21720 gcagaatgaa atccagcaaa aaacgcgcta tcaccgtctg caggctcttc tgagtagaaa 21780 acgtgagttg gatgaatttt tttcgatcat tcatccacgc ctgggctgct tttttcaggc 21840 gccggaatca actccatggt ggaagcaagg taaggtcttt tatgtccact ttcagtggca 21900 cacagccaaa cgagaataga tccattgcgc gttgccactt ctgctcattt ttgtcaatca 21960 actgacgccc catacgagcg acctgggata gctgcgggtc ttggttcttc ttgcgtccct 22020 ggggcgatct agaagggcct ggctgctcat cgtcggtgtc ggaaattggt ttcgattttt 22080 tacgctgcgg gccatccagt aacgcttcgg cgctctgcgg cgcagcgtcc tcactgacgg 22140 ctttgcggcg tctggcaacg cgctttggct tcggtgtttc gtcaatgaac agcttgccct 22200 cgtcgccgct gctttcagac acatcctcat agtgataccg gctcattttc cttctagatg 22260 gaagaacaca gcggtcagtc cagctccgag ccggcgccga atcacgagcc cgcggagctt 22320 agcttagaag atgctttgtc tccccaaccc gcggttgaaa gcgccgctcc gggttccgag 22380 gatgaaagcg aagctctcaa acactacatt gactccgacg tgctatttaa gcacatcgct 22440 agacagagtc gcatcctcaa agatagcctc gccgaccgct ttgaagtgcc tacagacgcg 22500 gtctagcgta ctagaactaa tgagcgctct ctattttctc catctacccc acccaagaag 22560 gcacctgcga caagaaaacg gccaaaccct cgaatcaatt tctacccaac cttcatgctg 22620 ccagaaacac tggcaacata tcacatattc ttttttaacc acaaaattcc gctgtcgtgt 22680 cgcgctaatc gcagtcgagc cgatgaaaag ttaatgctaa cagaaggaga ctgcatacct 22740 gattttccaa ccacggatcg ggttccaaaa atcttcgaag gtttgggctc agaagagaca 22800 actcactaga gtggcctcca agagaaaaga gacagcgctt tagtagaact gcttaacgac 22860 tcgccgcggc tcgcgattat aaagcgctcc acagcgctga ctcatttcgc atatcccgcc 22920 ataaacatgc cgccaaaagt gatgagttgt gtcatggagg aaatgattgt gaaaaaggcc 22980 gaa cccgtgg gagaagagtc gacacctgac ggtccagaag ggggcgcgcc agttgtcagt 23040 gacgcagaat tggccaagtg gcttggaagt agcgacgcca ccctgctcga agacaggcga 23100 aaactgatga tggccgttgt tctagtaaca gctcagctgg agtgcatgaa aaggtttttt 23160 acttcttctg acatgatcag aaagctaggt gaaacgctac actacacttt caggcacgga 23220 tacgtcaaac aagcctgtaa aatatcaaat gtcgaactac caaacctggt atcatacatg 23280 ggcatacttc atgaaaacag actaggtcag cacgtactgc acaacacact ccgcgatgaa 23340 cagaggcggg actacattag agacaccatc tttctgatgc ttttgtacac atggcagaca 23400 gcgatgggag tgtggcaaca atgtcttgag gtcgaaaaca tcaaagaact aagtaaactg 23460 ctcagacgaa agagacgggc gctttggaca ggctttgatg agcgaacaac cgccggcgac 23520 ctagccgaca taatctttcc gtcaaaactg ctatcgacat tgcaagccgg gctaccggat 23580 tttacaagcc agagcatgat gcaaaatttc cgcagcttca tattagaaag gtctggaata 23640 ttgccagcat tatgcaacgc cataccttca gactttgtgc caatagaata caaagagtgc 23700 ccgcctccgc tatgggcata ctgttatttg ctaaaattgg caaactacct aatgttccac 23760 tctgacgtag cttttaatat ggaaggagag gggctatttg agtgctactg tcgctgtaac 2 3820 ttgtgcaccc ctcaccgctg tcttgcaacc aacactgcct tactaaacga ggtgcaggcc 23880 ttgagcttca attggcagtt aaggccccca aatcctgacg ggtctatgcc tcccacactg 23940 aaattaacgg cgggggcttg gacctcggca tatttgagaa aatttgaacc tgcagactac 24000 cgtcacgatc aaattcgatt ctatgaggac caatcaaaac caccaaaatc cgagccatct 24060 gcctgcatca tcacgcaagc cgccattctc gcccaattac atgacataaa aaaagagcgg 24120 gaaaaattct tgcttaaaaa gggccacggc gtgtacctag accccaaaac aggcgaagag 24180 ctcaacacgc tagagccatc agtctctcac aatgccgcga gccgtcagac cgaccagtct 24240 aaatttgaca aaaccgaagt agccgcgcca cgcggaaaaa gaaccccctc ctccaacgcc 24300 actctggaga agacgaggaa gcattccagg cgaggacgta gaggaggaat gggacgatat 24360 agacagtttg gtcgcggagg agagcgagat ggaggacgag gaattggagg atggcgagac 24420 gagctattaa atcagtctcg gcctccgccg agaaggatca ctcccgccga aaacaaggaa 24480 ggccccaaaa cagcgtagat gggaccaaac tccaacatcg gcccctggta agcagaactc 24540 ggaaaataca gtcggtggga agtcgtggcg tccccacaaa catcacataa ttacggctct 24600 gctggcaagc gggtatgacg tgtccttcgc ccgcagattt atgctttacc gcc acggaat 24660 aaaaatgtaa aaacgttcca tccattacta caattcccaa tgcaggacag aatccccaga 24720 agaagtctgg aaagcgaaca atccagtcag ccagtacatc cgcagagccg gcgacgaccc 24780 aagagctgag agctaaaata ttcccaacgt tgtacgccat attccagcaa agcagaggtg 24840 5 gcggagtatc tctaaagata aagaaccgat ccttaagatc cctcacaaaa agctgccttt 24900 accacaagca ggagagtcag ccttggaaga ctgcagagaa cgccgaggct ctactccaga 24960 agtactgttc cgggctgaga ggctctgcgc cttatatctc agctcagcat gagtaaagac 25020 atccccaccc cttacgtatg gactttccag ccccaattgg ggcaggctgc cggcgcgtca 25080 caagactatt cgactcgcat gaactggcta agtgcaggtc cttcaatgat taaccaggtg 25140 aactctgtcc gagccgaccg aaacagaatc ttattgcgtc aagctgcagt atcggaaacg 25200 cccagactcg tccgcaaccc gccaacgtgg cctgcccaat acctatttca gccaattggc 25260 gcgcctcaga cctttgagct tcccaggaat gagtcattgg aggtggcaat gagtaactcg 25320 ggcatgcaat tagccggggg cgggacgcat cgcactaagg atataaaacc agaagacata 25380 gtgggacgcg gcctagagct gaacagcgac attccgagcg cttcgttttt gcgtcctgac 25440 ggagttttcc agcttgccgg aggtagccgt tcctctttca accc aggact gagtaccttg 25500 ctcacggtac aacctgcttc aagcctgcct aggtccggag gaatcggcga agtgcaattt 25560 10 gtgcacgagt ttgtgccgtc cgtgtacttt cagccttttt caggaccacc tggaacatat 25620 ccagacgaat ttatctacaa ctacgacata gtctcagatt ccgtcgacgg ttatgactga 25680 tacagagtct gatctttcgc tgtttggtgt ctgccggctg cactactccc gctgccagtc 25740 taccaactgc ttctggaagc agggaattct gccaacctac cagtgcattt tagacgcgga 25800 cctccacgcc gactgcgtgc cagactccct gcaagccggc cacagcctgc ggctcgaact 25860 gccacaccgt tttgcctgtt atcaaacctc aaatcacgga ttgcctatcg tgtgctccag 25920 caatgtcaag tcaagcagct tcaaagttac atgctcctgt tccagtactg ggatgcatct 25980 ggcgctcgcc gatgctctct gtgatcttgt taaccattct atggcagatg aagagcgcta 26040 aatcgctgcc gcacaaccac acagcggtaa tccccaggag tgtctgcgtg gccaacgttt 26100 cctgtacagg tgtggtgagt gcgacggcga ccctcacaga cgcccagact accgcatcag 26160 ccagtgggta ccgcgcgcca tgcgtagtat ccatcaactc cagcagcgac acaacgtgtg 26220 tttggaacgg ttggacatac aaagaatttc cgcttgaaat tcaattggac gaacgcttag 26280 •) 5-ccgacacccc tgtggactgt gtggaaggaa ggcgccgcac aacatttgac ctaagagctc 26340 tgtgtcggtt tcgctacacg cctctatatg ttttgaaatt atctatgcga agccatacca 26400 ctgtgcttac cattgtgggg gcgctcattg cgtttccggc tctgcgctcc cctctcgcca 26460 tcagcatgct cccagcagcc atggcagata atggctacca ccaccaccac acctgcgcgc 26520 cgttgctact gaccatatta atgttaatcg ccatgctgtg taaggtcacg aagcccacaa 26580 acaaactttt catcttagct cttcttagtc tagctgtgcc aggaaattgt ttgaaccagt 26640 acagtgtgct agaagggagt ccatgtgaac ttaaatctgc aacaaaacgc tacaccaaag 26700 cctcatggta tcgcgactct gaatctgcgc tgctttctcc tttcgccaca atcagtcaat 26760 cctcagtaac atactcttcc ccatcctcca gaataatcct agcttcaaac ttgtctctaa 26820 tttttacttc tgtaaaacct tcagacaacg gatcctattt tctaagcatc gactatcgcg 26880 aatttatcaa gtatgacctg cttgttagtc aatcaaccta ctaaaattca gccatacaaa 26940 cacagccagg agttaatcac acatgcataa tttctgccac ttgcagccca cacagcgccc 27000 Q agtacaggtc agtgatcaag tggcagaacc acacttacca ttcaaaagcg cttttcacag 27060 ttttcactga gcagttaaat aacaacataa catgcacagt gtcttctcct c ttgaaacta 27120 attccaagtc tttaacagcg tcacaaatgt gtgtttttca caatcctaat gacttcagcc 27180 ctctaatcat tgtaggtgtt ttaactcttg tgttcatagc catatggatc atctctatgt 27240 acgcgtccca ttcatactgt atgaactggt atctttaagt aataaactca gatataaatc 27300 cgtgattatt agacgcagct tctccgggtc ttcttttccc caacaatcta ttacagctcc 27360 ttcgtcaaga ctgacataac gcaacttaag caggtaagct attgcctaaa aactttctaa 27420 acactaccta aggcagcaaa acggcactcc attatacttt atctccattg cagttatccg 27480 caatgaagcg ctcgattccg tcagatttcg atccagtata tccctatgga aaaagacctt 27540 catgccgcca cacttaatat ttttacagcc aagacggttt tcaagaagca ccaaccgcca 27600 caaaatcaca ccctctcact cgtttaattc gacccaataa ctttcagtta cagcggggca 27660 aagtgggagg gggaataact attaaccaaa acggccaatt agaaactact aacgcgacca 27720 cccaccatta cagcagttaa gagtatgcta atggggcgat aagcctaaat accggaaacg 27780 gactggcagt tgactcaact caaaatctga atcgagccca caattcttac cttgccgtgt 27840 cacaatctgg tttaacgctg aacacaggtg atggcctaga ggtggatggt gatgaagtga 27900 aagttaagag cgggcaaggt gtgagtgtag gcactactgg agt tgggata aatgccgcct 27960 ctttccctca catactttgc aatgtattat ccctccttac tacagctcct ttaagtgtat 28020 tcttggagta ccagcggctc gagctaggca acggattaca gaccaactga agtgtcaaat 28080 cgctcaacac acagccacta tttacatttt ccaacggggc aatggggctg gcagtcggca 28140 acggaatcca aatagaaaat aacgctgtcg ccatttatgc ccaaccatat ttccaataca 28200 cctgggcctt ccaacagagc cgccttggaa atggcctgca gaccgaaaat aatgcaattg 28260 ccttatattg tcagccgtac ttccaatata cagataacgc actagcgctg cggctagggc 28320 aaggactgca aatctccaac aatcaagtag ctttatatgc tcagtcttac tttcaatata 28380 ccaataatgc attggcattg cgcttagcga acggtttagg cacgtctaac aataacgttg 28440 tggaaaaggc ttgtaaatta actcaagcga ttatttataa ttcaaacaaa ttgcaagtga 28500 atattagatc tccgctaaac tactatggca gttcacacac aattggtcta aatacaggaa 28560 acggtctaac tgttactagt cttggcgcgc taggtggcaa tgtatccgtt aatattggaa 28620 ttttagctca gcgggctatc actgggcagg tgcaggcttc attaggaaac gggctccaaa 28680 tcgcttccag tgccatagaa gtcaaactag gcaacggttt acagtttgac aatggcgcca 28740 tttccctatc agggtcatct cccgcctaca cagact acac tttatggact actccggacc 28800 cctctccaaa cgctaccatc agcgcagaat tagatgccaa gctggtgctt agtatttcaa 28860 aagcaggaag cactgctatc ggcaccatcg gtgtagttgg attgaaaggg cctctattaa 28920 gtttggccga gcaagccatc aatgttgaaa tttactttga caccagcggt aatattattt 28980 ttagcacaag cacgctgaag tcatactggg gatttaggtc tggtgattca tatgatccaa 29040 actccacact taatcctctt tatttaatgc caaatcagac cgcataccct ccagggcgac 29100 aaaccataac ccaaatagcc tcacttgaag tgtacttagg tggggatact accaaacctg 29160 ttcttttaga ggtagctttt aacaccgcaa gtagtggcta ctccctgaag tttacttggc 29220 gaaacttggc cagctatgcc ggacagacct tcgctgtatc ccttggaacc ttcacatata 29280 tcacacaaca ataaataagt tttaacatct ttatttgagt cgtgaatttt gtggcatcac 29340 tcttacagtc attccaccac caccactcca tgcaacctta tacacaagcc tttcaaaatg 29400 cattccagtg ttataacaat cagctttttt atgcaatttt acagcatgtt cataacattc 29460 aaagtcaggg gaagttatag agacaaagcc agcgggcata gactccaaag atggtttcag 29520 gtctaaaagt ttggatgcgt gtccacagtg tggtgaggct gattctccgg aggttctttc 29580 tggagcagat agcacttggg gcagccgcag cggtacttcg tcatcctcac tttgcagatc 29640 ggagtccctc tcgcaatcgc tagagtctcc actccaggaa caagacgaag ttccagactc 29700 actcgtccca ggtgtcgctg gat ctgacat tctgaagcct caagtccttc tagtccacac 29760 aagtcggaca aagaacacct ggcataccac ccaaacggaa catcgatcga cacattaaac 29820 ttcattactc tagaactgcc cgcagttaat aacatatcac tttccgcaca caaatgagcc 29880 gtttccccac ttctaatagg ggccgggtta tgtgagcgaa aaagacagcg aggaattcgc 29940 cgcctacctt cccattcttt cctttcgctt tcgctcatcc tagcccatcg ccgctcaaat 30000 cttaatttat tccttggctc gcaagcgtca tgcccacaat catcatattc acaatcagca 30060 tatttataca tcaagacagg cgtgccggcg cgcaaagcga tagggccttc ttcttccccc 30120 gctctatggt caccaaggca aaatcctgga gaggtgcata gtgacaaatg actgctaaat 30180 gcccaagaaa acacttttag atctggatta aacaatgatg aaagcatccc aactccataa 30240 tagccgtccg gatttctaat ctttacatca aacactatta ttttgtgaga cagttttttc 30300 cttcaagaca attacatctt cagatgaacc accgtcttgt cgctgttaaa gattggccgg 30360 tgtttgcatt tctccccaac ctcgatatta attggaggcg tgctcattct gaaagagatt 30420 ttttttcaat tgaaattttt actactggct ctccaggatt agcataaatc acatagtcta 30480 cccactcata cattttacac actatatttt taatcaaatt tggctcatgg gtgatgtctg 30540 catacagcca tggaac taaa catgaaacac tgaaatcata accggggggt ataaacactt 30600 catgatctaa gcagaagtcg ctctccccat atggcaggaa aaccattttc ttaggtgcca 30660 gcagccacac ttcattatcc gacttaagta tcggagcaag tgcatctgga ccggtgtaga 30720 caactgcatc caaataactt tgcaacacaa atgcgattta gcttaatcaa cgcgtcaagc 30780 gcgttgcttg ttgcgctcat actcgctagg tggcagtgaa agaatgttac atgcactgcc 30840 agcaagcagt aacaccgttc tagttctctc tgcacatgct aatgcggcca cctgtgtcat 30900 ttcatgacag tgtctacaaa ttattacaag atacaccccg gtataaccta tgcaaaacac 30960 cgccccttca aacctaagat tacgaattcg tggcacatcg ccccagtaat taatctttat 31020 aaaaatcaaa tgcaaacccc taaacataat actaccctca tataacagcc taaagcttaa 31080 atctttgtta gtcagtggcc tataaaatgg aaacctaata ttgtattctg tcccacccac 31140 cagctctttc acaatgtcag taataaagcg cctgcgtgac aaggcttcca aagtgctccc 31200 ttcatgcaaa tcatgacaac aaacagacca tttaaagcct ttattccttt tgaacattaa 31260 ttcaatgtct ccaccacatt caacttttaa gcgagtcccc aagcacagat aatctttcaa 31320 tgccaagaaa caatctatat caagaatact tttccatgga attggaattt ctaaattcat 31380 agccaatg gc agagctgact gtggagaatc aacataagaa ataattttgt gctgaaattt 31440 cgaccgagtg gactccgctg tcatctgtaa ttaaagatat ttaacatatt tttgagacat 31500 tacttcataa tggcaggcca gctgcaacag ctctctctgc ctaactacct cttttttgct 31560 tggctgttta gggccagagc agcaacccac agctcttttt aataggcgcc tagttcgttt 31620 tgcacacacc ttggcttgaa tttcagataa attacatgac ctgcaaataa ctattaaaaa 31680 aaaccatcca atttccatac catatacagc ctctccaaaa tttaatttac gaattctctt 31740 tctaaaatta catgtcgcca ctctgatgta tataaggtgg atcccacgaa tgtaaacact 31800 acccatgtac atcaacccct cttattcaca ctggtaaatt caccccctgt aaaaccaaaa 31860 tagctgattg taaaatgtcc cttttaaaac ctcttgaaac agtccagtaa gcactttacg 31920 ggaagacaag cactgaagac tgccagagtt gtcacaatgg caatgtaaat accatcttga 31980 ttgcaatgaa gaaaaatcag gttgaagatc tttagagaac acagaacaca tctctagttc 32040 cacaaatggc aacgccgtca gcttcaaaaa cacaaactca tgatgactta atatatgcat 32100 ggcagttcca ccatggcacc agcacatagc aaaacaggca gcagtgcgcg gtgagcgaac 32160 aaaggctact ggatgactgt ctgttgaatt gcaacaaatt aggccgctgc ctggaacctc 32220 aaaatgctcc atcctcaatc ttaagcagaa gctgtcgcag tttaggttcg acggaactcc 32280 agcctcagca aggagcagaa acctcttcgg gcactccctc gcccctaggg ccgcaattga 32340 tgtaattggc caacacaaaa ccaacgtagt gcacttggcg cattagagtc tggtctgctt 32400 tcctggtgaa atgaattttc gcagttgact gattggaaga gtaaataatt gagattatct 32460 ctgcaacgca ctgatgattt agcgcctcag ttgaaatgct cattgggtaa taatcacgaa 32520 gaatgcgctt cttaaagtga agacgtctgg atgtcattac tgcaataatg caacagtgat 32580 tgttttacat tcttccagac tcacatcctc accgattaat ctaaggtaat catacaaagc 32640 taatcccgca ctcatgtagg aatccgaaac taagaagtct tcatcactgc ctccattagt 32700 cagcaaagcg tccatagagc ttccagcaag gcaaaataat caacatagtt gcctcaagat 32760 ataggagcta ccgtatgatg acgtacagca agttagaacc agagaaatat aaagaagctc 32820 tggcagtcac aaaagccttt gccatagatg aaacagaagc aatgaaagat tctctattct 32880 ccacctgaag gcgagatgta aggcaatggg gaatggaaat ctgcatcatc tccccgtctc 32940 tgccattata taacaggtgg ctccagtaga tccaaatccg ccttcccctc gctcagtgtc 33000 gtcaagtgta tcaacctcct gaacctcagg caatgagatc ttttggatga caagctgagc 33060 tac gcgctgg cctggcgaaa taaggacgtg atgattccca gcaggacgaa tggttaaaga 33120 cacctctccc ctgtagtcgc tgtcaatcac gccagcgccc acatccaagc cttgagtcac 33180 agacaagcca gagcgaggtg caacgcgtcc gtagtgtccc tcaggaatac ggagctttag 33240 gccagtaggc acaagagcgc gagatccagc ctgaatctca acgtaatgcg aagcgcacaa 33300 atcatatcca gccgcaccat tagaagctct tttaggaggc acagccgagt cagacacacg 33360 cacaaagagc agcttgtcag ccatgatgta cttactcttg gcaaagtaga ccacggccta 33420 cagcaaggag tgtaagttta gaaatggcag cacagaaggc tcaggagcag aggcgaatga 33480 cgagcagagg agagaaatgg ctttatagag cgaaaaggcg cggtctgtga cgaggcgaaa 33540 gggcttctgg cgcctgacag aaccgcgtga gcgtaaccga caaagcacaa gacagtgcaa 33600 aagggcagtg actcagcgcc tcgccccgcc gctcgcacgc acacggacac tccccgcccc 33660 tcccagaaac tcccgcccag cgacctttga acaattttcc cacgccccct tacgcacagc 33720 acgtcaacgt catcacgcaa aagtgttccg tatattattg atgatgtcaa gagtggcacc 33780 tctttacctg cgcaggtaat atatagctca ctgggagtgg tgcatagaga aaaaagaccg 33840 cccatgatgg gagacgtggc atagaccttt agacaggttt cgtgcggcat cgcagtaaaa 3 3900 attgggaaaa gtggccataa atccctccac gtccgcgggc aaagggctcc aataaagtgg 33960 caaatttacg acagtgaaag tcaaagtcca ccctaaacac acagctgata cccatcataa 34020 atacacctga ctcagcgcct cgccccgccg ctcgcacgca cacggacact ccccgcccct 34080 cccagaaact cccgcccagc gacctttgaa caattttccc acgccccctt acgcacagca 34140 atcacgcaaa cgtcaacgtc agtgttccgt atattattga 34185 <TGATG; 210 > 4 < 211 > 6 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: Synthetic DNA primer < 400 > 4 ctgcag 6 < 210 > 5 < 211 > 36 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: DNA primer < 400 > 5 ggccttaatt aacatcatca ataatatacg gaacac 36 < 210 > 6 < 211 > 39 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: DNA primer < 400 > 6 ggaagatctt gagcatgcag agcaattcac gccgggtat 39 < 210 > 7 < 211 > 38 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: DNA primer < 400 > 7 ggcaatgaga tcttttggat gacaagctga gctacgcg 38 < 210 > 8 < 211 > 41 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: Synthetic DNA primer < 400 > 8 ctgtagatct gcggccgcgt ttaaacgtcg acaagcttcc c 41 < 211 > 27 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: Synthetic DNA primer < 400 > 9 aattcgagct cgcccgggcg agctcga 27 < 210 > 10 < 211 > 42 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: Synthetic DNA primer < 400 > 10 gactctaggg gcggggagtt taaacgcggc cgcagatcta gc 42 < 210 > 11 < 211 > 32 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of Artificial Sequence: Synthetic Smal Site < 400 > 11 gaattcgagc tcgcccgggc gagctcgaat te 32 < 210 > 12 < 211 > 27 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: Synthetic DNA primer < 400 > 12 ctgtagatct gcggccgcgt ttaaacg 27 < 210 > 13 < 211 > 14 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: Synthetic DNA primer < 400 > 13 tegacaaget tece 14 < 210 > 14 < 211 > 33 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: Synthetic DNA primer < 400 > 14 cccgggagtt taaacgcggc cgcagatcta gct 33 < 210 > 15 < 211 > 32 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: Synthetic DNA primer < 400 > 15 gaattcgagc tcgcccgggc gagctcgaat te 32 < 210 > 16 < 211 > 14 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of Artificial Sequence: Synthetic DNA primer < 400 > 16 tegacaaget tece 14 < 210 > 17 < 211 > 10 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: Synthetic DNA primer < 400 > 17 caagcttccc 10

Claims (22)

1. NOVELTY OF THE INVENTION CLAIMS 1. A recombinant virus, characterized in that it comprises a foreign DNA sequence inserted in the E4 gene region of a bovine adenovirus. 2. The recombinant virus according to claim 1, further characterized in that the foreign DNA encodes a polypeptide of a virus or bacterium that is selected from the group consisting of bovine rotavirus, bovine coronavirus, bovine herpes virus type 1, virus bovine respiratory syncytial virus, type 3 bovine influenza virus (BPI-3), bovine diarrhea virus, bovine rhinotracheitis virus, bovine parainfluenza virus type 3, Pasteurella haemolytica, Pasteurella multocida and Haemophillus somnus. 3. The recombinant virus, according to claim 2, further characterized in that the polypeptide comprises more than 10 amino acids. 4. The recombinant virus, according to claim 2, further characterized in that the polypeptide is antigenic. 5. The recombinant virus, according to claim 1, further characterized in that the bovine adenovirus is a bovine adenovirus of subgroup 1. The recombinant virus, according to claim 1, further characterized in that the bovine adenovirus is a bovine adenovirus of the subgroup 2. The recombinant virus according to claim 5, further characterized in that the foreign DNA sequence is under the control of a promoter that is located towards the 5 * end of a foreign DNA sequence. 8. A mutant virus, characterized in that it comprises a deletion of at least a portion of the E4 gene region of a bovine adenovirus. 9. The mutant virus, according to claim 8, further characterized in that it also has a foreign DNA sequence inserted in the region of the E4 gene. 10. The mutant virus according to claim 8, further characterized in that at least one open reading frame of the E4 gene region of the bovine adenovirus has been completely deleted. 11. A recombinant virus, characterized in that it comprises a foreign DNA sequence inserted within the E3 gene region of a bovine adenovirus 1. 12. The recombinant virus according to claim 11, further characterized in that the foreign DNA encodes a polypeptide of a virus or bacterium that is selected from the group consisting of bovine rotavirus, bovine coronavirus, bovine herpes virus type 1, respiratory syncytial virus bovine, bovine parainfluenza virus type 3 (BPI-3), bovine diarrhea virus, bovine rhinotracheitis virus, bovine parainfluenza virus type 3, Pasteurella haemolytica, Pasteurella multocida and Haemophillus somnus. 13. The recombinant virus, according to claim 12, further characterized in that the polypeptide comprises more than 10 amino acids. 14. The recombinant virus, according to claim 12, further characterized in that the polypeptide is antigenic. 15. The recombinant virus according to claim 12, further characterized in that the foreign DNA sequence is under the control of a promoter that is located towards the 5 'end of the foreign DNA sequence. 16. A mutant virus, characterized in that it comprises a deletion of at least a portion of the E3 gene region of a bovine adenovirus 1. 17. The mutant virus according to claim 16, further characterized in that it also has a foreign DNA sequence inserted into the region of the E3 gene. 18. The recombinant virus according to claim 16, further characterized in that at least one open reading frame of the E3 gene region of bovine adenovirus 1 has been completely deleted. 19. A vaccine comprising the recombinant virus as claimed in claim 1. 20. The use of a recombinant virus, as claimed in claim 1, for preparing a vaccine for inducing an immune response in an animal. twenty-one . A vaccine, comprising the recombinant virus as claimed in claim 11. 22. The use of the recombinant virus as claimed in claim 1, for preparing a vaccine for inducing an immune response in an animal.