CN1369506A - Human Protein for promoting transform of 3T3 cell and its coding sequence - Google Patents
Human Protein for promoting transform of 3T3 cell and its coding sequence Download PDFInfo
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Abstract
A novel human protein with the function of promoting 3T3 cell transform, the polynucleotide for coding the peptide, the process for preparing said polypeptide by recombination, the antagon of said polypeptide and its medical action, and the application of said polynucleotide are disclosed.
Description
The invention belongs to biological technical field, specifically, the present invention relates to new coding and have the proteic polynucleotide of people that promote 3T3 cell transformation function, and the polypeptide of this polynucleotide encoding.The invention still further relates to the purposes and the preparation of these polynucleotide and polypeptide.
The research of people's gene group is international focus at present, removes human chromosome DNA large scale sequencing, outside the method for expressed sequence order-checking (EST), also lacks the screening that begins from function and has the high-throughout method of functional gene.
Cancer is one of principal disease of harm humans health.In order to treat effectively and prophylaxis of tumours, people more and more pay close attention to genetic treatment of tumor at present.Therefore, this area presses for development research people albumen and the agonist/inhibitor thereof relevant with growth of cancer cells.
The purpose of this invention is to provide people's protein polypeptide that new the having of a class promote 3T3 cell transformation function with and fragment, analogue and derivative.
Another object of the present invention provides the polynucleotide of these polypeptide of coding.
Another object of the present invention provides the method for these polypeptide of production and the purposes of this polypeptide and encoding sequence.
In a first aspect of the present invention, novel isolated protein polypeptide with promotion 3T3 cell transformation function is provided, it comprises the polypeptide of the aminoacid sequence with the group of being selected from down: SEQ ID NO:2,5,8,11,14,17,20,23; Or its conservative property variation polypeptide or its active fragments or its reactive derivative.
Preferably, this polypeptide is the polypeptide with aminoacid sequence of the group of being selected from down: SEQ ID NO:2,5,8,11,14,17,20,23.
In a second aspect of the present invention, a kind of isolating polynucleotide are provided, it comprises a nucleotide sequence, and this nucleotide sequence is shown at least 85% homogeny with a kind of nucleotides sequence that is selected from down group: the polynucleotide with the protein polypeptide that promotes 3T3 cell transformation function that (a) coding is above-mentioned; (b) with polynucleotide (a) complementary polynucleotide.Preferably, the polypeptide of this polynucleotide encoding has the aminoacid sequence of the group of being selected from down: SEQ ID NO:2,5,8,11,14,17,20,23.More preferably, the sequence of these polynucleotide is selected from down group: SEQ ID NO:3,6,9,12,15,18,21,24 coding region sequence or full length sequence.
In a third aspect of the present invention, the carrier that contains above-mentioned polynucleotide is provided, and has been transformed or host cell of transduceing or the host cell that is directly transformed or transduce by above-mentioned polynucleotide by this carrier.
In a fourth aspect of the present invention, provide preparation to have the preparation method of the polypeptide of the protein-active that promotes 3T3 cell transformation function, this method comprises: (a) being fit to express under the proteic condition with promotion 3T3 cell transformation function, cultivate the above-mentioned host cell that is transformed or transduce; (b) from culture, isolate polypeptide with the protein-active that promotes 3T3 cell transformation function.
In a fifth aspect of the present invention, provide and the above-mentioned protein polypeptide specificity bonded antibody that promotes 3T3 cell transformation function that has.The nucleic acid molecule that can be used for detecting also is provided, and it contains, and continuous 10 Nucleotide are to full length nucleotide in the above-mentioned polynucleotide, and preferably it contains the about 10-800 of a successive Nucleotide.
In a sixth aspect of the present invention, a kind of pharmaceutical composition is provided, it contains the protein polypeptide and the pharmaceutically acceptable carrier with promotion 3T3 cell transformation function of the present invention of safe and effective amount.These pharmaceutical compositions can be used for promoting the growth of cell.The present invention also provides a kind of pharmaceutical composition, it contain safe and effective amount at antagonist (as antibody) and the pharmaceutically acceptable carrier with the protein polypeptide that promotes 3T3 cell transformation function of the present invention.This pharmaceutical composition can be treated illnesss such as cancer and cellular abnormality propagation.
Others of the present invention are because disclosing of the technology of this paper is conspicuous to those skilled in the art.
The 3T3 cell is a kind of l cell (J.Cell.Biol., 17:299,1963).In the cancer research field, often foreign gene (especially people's gene) is introduced the 3T3 cell, observe its situation that influences to the growth of 3T3 cell.It has been generally acknowledged that, to 3T3 cell growth (or vicious transformation) influential gene is cancer related gene, wherein to 3T3 cell growth or transform that inhibiting gene is arranged is cancer suppressor gene mostly, and to the growth of 3T3 cell or transform (former) oncogene that has the gene of promoter action to be mostly.
The present invention adopts large-scale cDNA clone transfection mouse embryo fibroblasts 3T3, has on the basis that promotes the growth effect in acquisition, proves new gene through order-checking, further obtains full length cDNA clone.DNA transfection evidence, of the present invention have the egg of promotion 3T3 cell transformation function from the 3T3 cell being had the effect that promotes that the clone forms, its promotion rate 〉=50%.
As used herein, " isolating " is meant that material separates (if natural substance, primal environment promptly is a natural surroundings) from its primal environment.Do not have separation and purification as polynucleotide under the native state in the active somatic cell and polypeptide, but same polynucleotide or polypeptide as from native state with in other materials that exist separately, then for separation and purification.
As used herein, " isolating albumen or polypeptide with promotion 3T3 cell transformation function " is meant to have and promotes the protein polypeptide of 3T3 cell transformation function to be substantially free of natural relative other albumen, lipid, carbohydrate or other material.Those skilled in the art can have the albumen that promotes 3T3 cell transformation function with the purified technology of protein purifying of standard.Basically pure polypeptide can produce single master tape on non-reduced polyacrylamide gel.
Polypeptide of the present invention can be recombinant polypeptide, natural polypeptides, synthetic polypeptide, preferred recombinant polypeptide.Polypeptide of the present invention can be the product of natural purifying, or the product of chemosynthesis, or uses recombinant technology to produce from protokaryon or eucaryon host (for example, bacterium, yeast, higher plant, insect and mammalian cell).The host used according to the recombinant production scheme, polypeptide of the present invention can be glycosylated, maybe can be nonglycosylated.Polypeptide of the present invention also can comprise or not comprise initial methionine residues.
The present invention also comprises having the proteic fragment of people, derivative and the analogue that promotes 3T3 cell transformation function.As used herein, term " fragment ", " derivative " are meant with " analogue " and keep natural identical biological function or the active polypeptide of people's albumen that promotes 3T3 cell transformation function that have of the present invention basically.Polypeptide fragment of the present invention, derivative or analogue can be that (i) has one or more conservative or substituted polypeptide of non-conservation amino-acid residue (preferred conservative amino acid residue), and the amino-acid residue of such replacement can be also can not encoded by genetic code, or (ii) in one or more amino-acid residues, has a polypeptide of substituted radical, or (iii) mature polypeptide and another compound (such as the compound that prolongs the polypeptide transformation period, polyoxyethylene glycol for example) merge formed polypeptide, or (iv) additional aminoacid sequence is fused to this peptide sequence and the polypeptide that forms (as leader sequence or secretion sequence or be used for the sequence or the proteinogen sequence of this polypeptide of purifying).According to the instruction of this paper, these fragments, derivative and analogue belong to the known scope of those skilled in the art.
Polynucleotide of the present invention can be dna form or rna form.Dna form comprises the DNA of cDNA, genomic dna or synthetic.DNA can be strand or double-stranded.DNA can be coding strand or noncoding strand.Be example with PP12719 albumen (in this application, its clone numbering is adopted in proteinic name), the coding region sequence of encoding mature polypeptide can be identical with the coding region sequence shown in the SEQ ID NO:3 or the varient of degeneracy.As used herein, " varient of degeneracy " is meant that in the present invention coding has the protein of SEQ ID NO:2, but with the differentiated nucleotide sequence of coding region sequence shown in the SEQ ID NO:3.Be example with PP13181 albumen (in this application, its clone numbering is adopted in proteinic name) again, the coding region sequence of encoding mature polypeptide can be identical with the coding region sequence shown in the SEQ ID NO:6 or the varient of degeneracy.Have the albumen that promotes 3T3 cell transformation function for other, the rest may be inferred.
The polynucleotide of encoding mature polypeptide comprise: the encoding sequence of an encoding mature polypeptide; The encoding sequence of mature polypeptide and various additional code sequence; Encoding sequence of mature polypeptide (with optional additional code sequence) and non-coding sequence.
Term " polynucleotide of coded polypeptide " can be the polynucleotide that comprise this polypeptide of encoding, and also can be the polynucleotide that also comprise additional code and/or non-coding sequence.
The invention still further relates to the varient of above-mentioned polynucleotide, its coding has the polypeptide of identical aminoacid sequence or fragment, analogue and the derivative of polypeptide with the present invention.The varient of these polynucleotide can be the allelic variant of natural generation or the varient that non-natural takes place.These nucleotide diversity bodies comprise and replace varient, deletion mutation body and insert varient.As known in the art, allelic variant is the replacement form of polynucleotide, and it may be replacement, disappearance or the insertion of one or more Nucleotide, but can be from not changing the function of its encoded polypeptides in fact.
The invention still further relates to and above-mentioned sequence hybridization and two sequences between have at least 50%, preferably at least 70%, the polynucleotide of at least 80% homogeny more preferably.The present invention be more particularly directed under stringent condition and the interfertile polynucleotide of polynucleotide of the present invention.In the present invention, " stringent condition " is meant: (1) than hybridization under low ionic strength and the comparatively high temps and wash-out, as 0.2 * SSC, and 0.1%SDS, 60 ℃; Or (2) hybridization the time is added with denaturing agent, as 50% (v/v) methane amide, 0.1% calf serum/0.1%Ficoll, 42 ℃ etc.; Or (3) only at the homogeny between the two sequences at least more than 95%, be more preferably 97% and just hybridize when above.And the polypeptide of interfertile polynucleotide encoding has identical biological function and activity with the mature polypeptide shown in the SEQ IDNO:2 (is example with PP12719 albumen).
The invention still further relates to nucleic acid fragment with above-mentioned sequence hybridization.As used herein, the length of " nucleic acid fragment " contains 15 Nucleotide at least, better is at least 30 Nucleotide, is more preferably at least 50 Nucleotide, preferably more than at least 100 Nucleotide.The amplification technique (as PCR) that nucleic acid fragment can be used for nucleic acid has the proteic polynucleotide that promotes 3T3 cell transformation function to determine and/or to separate coding.
Polypeptide among the present invention and polynucleotide preferably provide with isolating form, more preferably are purified to homogeneous.
Dna sequence dna of the present invention can obtain with several method.For example, with hybridization technique DNA isolation well known in the art.These technology including, but not limited to: 1) with probe and genome or the hybridization of cDNA library to detect homology nucleotide sequence and 2) antibody screening of expression library to be to detect the dna fragmentation of the clone with common structure feature.
Coding has the proteic specific DNA fragment sequence that promotes 3T3 cell transformation function and produces also and can obtain with following method: 1) separate double chain DNA sequence from genomic dna; 2) the chemical synthesising DNA sequence is to obtain the double-stranded DNA of required polypeptide.
When the whole aminoacid sequence of the polypeptide product of needs was known, the direct chemical of dna sequence dna is synthetic to be the method for often selecting for use.When if required amino acid whose whole sequence is not known, the direct chemical of dna sequence dna is synthetic to be impossible, and the method for selecting for use is the separation of cDNA sequence.The standard method that separates interested cDNA is from the donorcells separating mRNA of this gene of high expression level and carries out reverse transcription, forms plasmid or phage cDNA library.Extract the existing multiple proven technique of method of mRNA, test kit also can obtain (Qiagene) from commercial channels.And the construction cDNA library also is usual method (Sambrook, et al., Molecular Cloning, A Laboratory Manual, Cold Spring HarborLaboratory.New York, 1989).Also can obtain the cDNA library of commercial offers, as the different cDNA library of Clontech company.When being used in combination the polymeric enzyme reaction technology, even few expression product also can be cloned.
Available ordinary method is screened gene of the present invention from these cDNA libraries.These methods include, but is not limited to; (1) DNA-DNA or DNA-RNA hybridization; (2) function of marker gene occurs or forfeiture; (3) measure level with the proteic transcript that promotes 3T3 cell transformation function; (4), detect the protein product of genetic expression by immunological technique or mensuration biologic activity.Aforesaid method can singly be used, but also several different methods combined utilization.
In (1) kind method, hybridizing used probe is and any a part of homology of polynucleotide of the present invention that at least 15 Nucleotide of its length better are at least 30 Nucleotide, are more preferably at least 50 Nucleotide, preferably at least 100 Nucleotide.In addition, the length of probe within 2kb, preferably is within the 1kb usually.Probe used herein is the dna sequence dna of chemosynthesis on the basis of gene DNA sequence information of the present invention normally.Gene of the present invention itself or fragment are certainly as probe.The mark of dna probe can be used radio isotope, fluorescein or enzyme (as alkaline phosphatase) etc.
In (4) kind method, detect protein product and can use immunological technique such as Western blotting, radioimmunoprecipitation, enzyme-linked immunosorbent assay (ELISA) etc. with the protein gene expression that promotes 3T3 cell transformation function.
The method (Saiki, et al.Science 1985:230:1350-1354) of using round pcr DNA amplification/RNA is optimized for and obtains gene of the present invention.When particularly being difficult to obtain the cDNA of total length from the library, can preferably use RACE method (the terminal rapid amplifying method of RACE-cDNA), the primer that is used for PCR can suitably be selected according to sequence information of the present invention disclosed herein, and available ordinary method is synthetic.Available ordinary method is as the DNA/RNA fragment by gel electrophoresis separation and purifying amplification.
The gene of the present invention that obtains as mentioned above, perhaps the available ordinary method of mensuration of the nucleotide sequence of various dna fragmentations etc. such as dideoxy chain termination (Sanger et al.PNAS, 1977,74:5463-5467).This class nucleotide sequencing is available commercial sequencing kit etc. also.In order to obtain the cDNA sequence of total length, order-checking need be carried out repeatedly.Sometimes need to measure a plurality of clones' cDNA sequence, just can be spliced into the cDNA sequence of total length.
The present invention also relates to comprise the carrier of polynucleotide of the present invention, and with carrier of the present invention or have the host cell that the albumen coded sequence that promotes 3T3 cell transformation function produces through genetically engineered, and the method that produces polypeptide of the present invention through recombinant technology.
Recombinant DNA technology (Science, 1984 by routine; 224:1431), can utilize polymerized nucleoside acid sequence of the present invention to can be used to express or produce the protein polypeptide that promotes 3T3 cell transformation function that has of reorganization.In general following steps are arranged:
(1). have the proteic polynucleotide of people (or varient) that promote 3T3 cell transformation function with coding of the present invention, or transform or the transduction proper host cell with the recombinant expression vector that contains these polynucleotide;
(2). the host cell of in suitable medium, cultivating;
(3). separation, protein purification from substratum or cell.
Among the present invention, the people's albumen polynucleotide sequence with promotion 3T3 cell transformation function can be inserted in the recombinant expression vector.Term " recombinant expression vector " refers to that bacterial plasmid well known in the art, phage, yeast plasmid, vegetable cell virus, mammalian cell virus are as adenovirus, retrovirus or other carriers.The carrier of Shi Yonging includes but not limited in the present invention: and the expression vector based on T7 of in bacterium, expressing (Rosenberg, et al.Gene, 1987,56:125); The pMSXND expression vector of in mammalian cell, expressing (Lee and Nathans, J Bio Chem.263:3521,1988) and at the carrier that derives from baculovirus of expressed in insect cells.In a word, as long as can duplicate in host and stablize, any plasmid and carrier can be used.A key character of expression vector is to contain replication orgin, promotor, marker gene and translation controlling elements usually.
Method well-known to those having ordinary skill in the art can be used to make up contain and has people's encoding histone dna sequence dna of promoting 3T3 cell transformation function and suitable transcribing/translate the expression vector of control signal.These methods comprise (Sambroook, et al) such as extracorporeal recombinant DNA technology, DNA synthetic technology, the interior recombinant technologys of body.Described dna sequence dna can effectively be connected on the suitable promotor in the expression vector, and is synthetic to instruct mRNA.The representative example of these promotors has: colibacillary lac or trp promotor; Lambda particles phage P
LPromotor; Eukaryotic promoter comprises CMV immediate early promoter, early stage and late period SV40 promotor and some other known may command gene expression promoter in protokaryon or eukaryotic cell or its virus.Expression vector also comprises ribosome bind site and the transcription terminator that translation initiation is used.
In addition, expression vector preferably comprises one or more selected markers, to be provided for selecting the phenotypic character of transformed host cells, cultivate Tetrahydrofolate dehydrogenase, neomycin resistance and the green fluorescent protein (GFP) of usefulness as eukaryotic cell, or be used for colibacillary tsiklomitsin or amicillin resistance.
Comprise the carrier of above-mentioned suitable dna sequence dna and suitable promotor or control sequence, can be used to transform appropriate host cell, so that it can marking protein.
Host cell can be a prokaryotic cell prokaryocyte, as bacterial cell; Or eukaryotic cell such as low, as yeast cell; Or higher eucaryotic cells, as mammalian cell.Representative example has: intestinal bacteria, streptomyces; The bacterial cell of Salmonella typhimurium; Fungal cell such as yeast; Vegetable cell; The insect cell of fruit bat S2 or Sf9; The zooblast of CHO, COS or Bowes melanoma cells etc.
When polynucleotide of the present invention are expressed in higher eucaryotic cells, be enhanced if will make to transcribe when in carrier, inserting enhancer sequence.Enhanser is the cis acting factor of DNA, and nearly 10 to 300 base pairs act on promotor transcribing with enhancing gene usually.Can for example be included in the SV40 enhanser of 100 to 270 base pairs of replication origin side in late period one, at the polyoma enhanser of replication origin side in late period one and adenovirus enhanser etc.
Persons skilled in the art all know how to select appropriate carriers, promotor, enhanser and host cell.
Can carry out with routine techniques well known to those skilled in the art with the recombinant DNA transformed host cell.When the host was prokaryotic organism such as intestinal bacteria, the competent cell that can absorb DNA can be used CaCl in exponential growth after date results
2Method is handled, and used step is well-known in this area.Alternative is to use MgCl
2If desired, transforming also the method for available electroporation carries out.When the host is an eukaryote, can select following DNA transfection method for use: coprecipitation of calcium phosphate method, conventional mechanical method such as microinjection, electroporation, liposome packing etc.
The transformant that obtains can be cultivated with ordinary method, expresses the polypeptide of coded by said gene of the present invention.According to used host cell, used substratum can be selected from various conventional substratum in the cultivation.Under the condition that is suitable for the host cell growth, cultivate.After host cell grows into suitable cell density, induce the promotor of selection with suitable method (as temperature transition or chemical induction), cell is cultivated for some time again.
Recombinant polypeptide in the above methods can wrap by in cell, extracellular or on cytolemma, express or be secreted into the extracellular.If desired, can utilize its physics, the separating by various separation methods with other characteristic and the albumen of purification of Recombinant of chemistry.These methods are well-known to those skilled in the art.The example of these methods includes, but are not limited to: conventional renaturation handles, with protein precipitant handle (salt analysis method), centrifugal, the broken bacterium of infiltration, superly handle, the combination of super centrifugal, sieve chromatography (gel-filtration), adsorption chromatography, ion exchange chromatography, high performance liquid chromatography (HPLC) and other various liquid chromatography (LC) technology and these methods.
Having of reorganization promotes the people's albumen or the polypeptide of 3T3 cell transformation function to be of use in many ways.These purposes include, but is not limited to: directly have the disease due to the low or forfeiture of the protein function that promotes 3T3 cell transformation function as pharmacological agent and be used to screen and promote or antagonism has antibody, polypeptide or other part of the protein function that promotes 3T3 cell transformation function.For example, this antibody can be used for treating cancer or cellular abnormality propagation.The peptide molecule that can suppress or stimulate people's protein function that can be used for seeking therapeutic value with recombinant expressed protein screening peptide library of the present invention with promotion 3T3 cell transformation function.
The present invention also provides screening of medicaments to improve (agonist) or check the method that (antagonist) has the proteic medicament of people that promotes 3T3 cell transformation function to identify.Agonist improves and to have the people's albumen that promotes 3T3 cell transformation function biological function such as stimulate cellular proliferation, and antagonist prevention disorder such as the various cancer relevant with cell hyperproliferation with treatment.
Have the proteic antagonist of people that promotes 3T3 cell transformation function and comprise antibody, compound, acceptor disappearance thing and the analogue etc. that filter out.Have the proteic antagonist of people that promotes 3T3 cell transformation function and can and eliminate its function with people's protein binding with promotion 3T3 cell transformation function, or suppress to have the proteic generation of people that promotes 3T3 cell transformation function, or combine with the avtive spot of polypeptide and to make polypeptide can not bring into play biological function.Have and promote the proteic antagonist of people of 3T3 cell transformation function to can be used for therepic use.
In screening during as the compound of antagonist, can add in the bioanalysis mensuration having the albumen that promotes 3T3 cell transformation function, the albumen and the interaction between its acceptor that have promotion 3T3 cell transformation function by the mensuration compounds affect determine whether compound is antagonist.With the same quadrat method of above-mentioned SCREENED COMPOUND, can filter out the acceptor disappearance thing and the analogue of antagonist action.
The proteic antagonist of the present invention can be directly used in disease treatment, for example, and various malignant tumours and cellular abnormality propagation etc.
Polypeptide of the present invention, and fragment, derivative, analogue or their cell can be used as antigen to produce antibody.These antibody can be polyclone or monoclonal antibody.Polyclonal antibody can obtain by the method with this polypeptide direct injection animal.The technology of preparation monoclonal antibody comprises hybridoma technology, three knurl technology, people B-quadroma technology, EBV-hybridoma technology etc.
Can be with polypeptide of the present invention and antagonist and suitable pharmaceutical carrier combination back use.These carriers can be water, glucose, ethanol, salt, damping fluid, glycerine and their combination.Composition comprises the polypeptide or the antagonist of safe and effective amount and carrier and the vehicle that does not influence effect of drugs.These compositions can be used as medicine and are used for disease treatment.
The present invention also provides medicine box or the test kit that contains one or more containers, and one or more medicinal compositions compositions of the present invention are housed in the container.With these containers, can have by the given indicative prompting of government authorities of making, using or selling medicine or biological products, the government authorities that this prompting reflects production, uses or sells permits it to use on human body.In addition, polypeptide of the present invention can be used in combination with other treatment compound.
Pharmaceutical composition can be with mode administration easily, as by in part, intravenously, intraperitoneal, intramuscular, subcutaneous, the nose or the route of administration of intracutaneous.Have the albumen or its specific antibody that promote 3T3 cell transformation function, can come administration by the amount that treats and/or prevents concrete indication effectively.Be applied to having of patient and promote the proteic amount and the dosage range of 3T3 cell transformation function will depend on many factors, as administering mode, person's to be treated healthiness condition and diagnostician's judgement.
Have and promote the proteic polynucleotide of people of 3T3 cell transformation function also to can be used for multiple therapeutic purpose.Gene therapy technology can be used for treating since have that the proteic nothing that promotes 3T3 cell transformation function is expressed or unusual/non-activity have cell development or a metabolic disturbance due to the proteic expression that promotes 3T3 cell transformation function.The gene therapy vector (as virus vector) of reorganization can be designed to express the albumen that promotes 3T3 cell transformation function that has of variation, to suppress the endogenic protein-active that promotes 3T3 cell transformation function that has.For example, a kind of albumen that promotes 3T3 cell transformation function that has of variation can be the albumen with promotion 3T3 cell transformation function that shortens, lacked signal conduction function territory, though can combine with the substrate in downstream, lacks signaling activity.Therefore the gene therapy vector of reorganization can be used for treating and has the protein expression that promotes 3T3 cell transformation function or the disease of active caused by abnormal.Deriving from the expression vector of virus such as retrovirus, adenovirus, adeno-associated virus (AAV), hsv, parvovirus etc. can be used for having and promotes the protein gene of 3T3 cell transformation function to be transferred in the cell.The method that structure carries the recombinant viral vector with the protein gene that promotes 3T3 cell transformation function is found in existing document (Sambrook, et al.).Reorganization has the people's protein gene that promotes 3T3 cell transformation function and can be packaged in the liposome and be transferred in the cell in addition.
Inhibition has the oligonucleotide (comprising sense-rna and DNA) of the people's protein mRNA that promotes 3T3 cell transformation function and ribozyme also within the scope of the invention.Ribozyme is the enzyme sample RNA molecule that a kind of energy specificity is decomposed specific RNA, and its mechanism of action is to carry out the endonuclease effect after ribozyme molecule and the hybridization of complementary target RNA-specific.The RNA of antisense and DNA and ribozyme can obtain with existing any RNA or DNA synthetic technology, as the technology widespread use of solid phase phosphoamide chemical synthesis synthetic oligonucleotide.Antisense rna molecule can be transcribed acquisition by the dna sequence dna of this RNA that encodes in external or body.This dna sequence dna has been incorporated into the downstream of rna polymerase promoter of carrier.In order to increase the stability of nucleic acid molecule, available several different methods is modified it, and as increasing the sequence length of both sides, the connection between the ribonucleoside is used phosphoric acid thioester bond or peptide bond but not phosphodiester bond.
Polynucleotide imports tissue or intracellular method comprises: directly be injected into polynucleotide in the in-vivo tissue; Or external by carrier (as virus, phage or plasmid etc.) earlier with the polynucleotide transfered cell in, again cell is transplanted in the body etc.Because albumen of the present invention has the function that promotes the 3T3 cell transformation, so the antisense sequences of albumen coded sequence of the present invention, can be introduced into cell to suppress the abnormality proliferation (as canceration) of cell.
The present invention also provides at the antibody with the people's proteantigen determinant that promotes 3T3 cell transformation function.These antibody include, but is not limited to: the fragment that polyclonal antibody, monoclonal antibody, chimeric antibody, single-chain antibody, Fab fragment and Fab expression library produce.
The anti-proteic antibody of people with promotion 3T3 cell transformation function can be used in the immunohistochemistry technology, detects the people's albumen that promotes 3T3 cell transformation function that has in the biopsy specimen.
The also available labelled with radioisotope of the protein bound monoclonal antibody of people with having promotion 3T3 cell transformation function injects in the body and can follow the tracks of its position and distribution.This radiolabeled antibody can be used as a kind of atraumatic diagnostic method and is used for the location of tumour cell and has judged whether transfer.
Antibody among the present invention can be used for treating or preventing and have the relevant disease of people's albumen of promotion 3T3 cell transformation function.The antibody that gives suitable dosage can be blocked proteic generation of people or the activity with promotion 3T3 cell transformation function, thus the abnormality proliferation of the growth of anticancer and/or cell.
Antibody also can be used for designing the immunotoxin at a certain privileged sites in the body.As have the people's albumen high-affinity that promotes 3T3 cell transformation function monoclonal antibody can with bacterium or plant poison (as diphtheria toxin, ricin, abrine etc.) covalent attachment.A kind of usual method is with sulfydryl linking agent such as SPDP, attacks the amino of antibody, by the exchange of disulfide linkage, toxin is incorporated on the antibody, and this hybrid antibody can be used for killing relevant positive cell (as cancer cells).
Available people's albumen or the polypeptide immune animal of the production of polyclonal antibody with promotion 3T3 cell transformation function, as rabbit, mouse, rat etc.Multiple adjuvant can be used for the enhancing immunity reaction, includes but not limited to freund's adjuvant etc.
Have promote 3T3 cell transformation function people's protein monoclonal antibody can with hybridoma technology production (Kohler andMilstein.Nature, 1975,256:495-497).With the variable region bonded chimeric antibody in human constant region and inhuman source can with existing technology production (Morrison et al, PNAS, 1985,81:6851).And the technology of existing manufacture order chain antibody (U.S.Pat No.4946778) also can be used for producing the anti-proteic single-chain antibody of people that promotes 3T3 cell transformation function that has.
Can with have the protein bound peptide molecule of people that promotes 3T3 cell transformation function and can be incorporated into the rondom polypeptide storehouse that solid formation forms by the various amino acid that may make up by screening and obtain.During screening, must promote people's protein molecular of 3T3 cell transformation function to carry out mark to having.
The invention still further relates to quantitatively and detection and localization has the diagnostic testing process of people's protein level of promotion 3T3 cell transformation function.These tests are known in the art, and comprise that FISH measures and radioimmunoassay.That is detected in the test has a protein level that promotes 3T3 cell transformation function, can have the importance of albumen in various diseases that promotes 3T3 cell transformation function with laying down a definition and be used to diagnose to have the disease that the albumen that promotes 3T3 cell transformation function works.
Proteic polynucleotide with promotion 3T3 cell transformation function can be used for having the diagnosis and the treatment of the protein related diseases that promotes 3T3 cell transformation function.Aspect diagnosis, have the proteic polynucleotide that promotes 3T3 cell transformation function can be used for detecting have promote 3T3 cell transformation function proteic expression whether or under morbid state, have an abnormal exprssion that promotes 3T3 cell transformation function.As the protein D NA sequence with promotion 3T3 cell transformation function can be used for that the hybridization of biopsy specimen is had the proteic abnormal expression that promotes 3T3 cell transformation function with judgement.Hybridization technique comprises the Southern blotting, Northern blotting, in situ hybridization etc.These technological methods all are disclosed mature technologies, and relevant test kit all can obtain from commercial channels.Part or all of polynucleotide of the present invention can be used as probe stationary on microarray (Microarray) or DNA chip (being gene chip), is used for analyzing the differential expression analysis and the gene diagnosis of tissue gene.Carry out RNA-polymerase chain reaction (RT-PCR) amplification in vitro with the special primer of albumen and also can detect proteic transcription product with promotion 3T3 cell transformation function with promotion 3T3 cell transformation function.
The sudden change that detection has the protein gene that promotes 3T3 cell transformation function also can be used for diagnosing the relevant disease of albumen with promotion 3T3 cell transformation function.Form with the protein mutation that promotes 3T3 cell transformation function comprises that to have point mutation that the protein D NA sequence that promotes 3T3 cell transformation function compares, transposition, disappearance, reorganization and other any unusual etc. with normal wild type.Available existing technology such as Southern blotting, dna sequence analysis, PCR and in situ hybridization detect sudden change.In addition, sudden change might influence proteic expression, therefore can judge indirectly that with Northern blotting, Western blotting gene has or not sudden change.
Sequence of the present invention identifies it also is valuable to karyomit(e).These sequences can be specifically at certain bar human chromosome particular location and and can with its hybridization.At present, need to identify the concrete site of each gene on the karyomit(e).Yet have only chromosomal marker thing seldom to can be used for the marker chromosomes position now based on actual sequence data (repetition polymorphism).For these sequences are associated with disease related gene.The first step is positioned dna sequence dna of the present invention on the karyomit(e) exactly.
In brief, prepare PCR primer (preferred 15-35bp), sequence can be positioned on the karyomit(e) according to cDNA.Then, these primers are used for the somatocyte hybrid cell that the PCR screening contains each bar human chromosome.Have only those hybrid cells that contain corresponding to the people's gene of primer can produce the fragment of amplification.
The PCR localization method of somatocyte hybrid cell is that DNA is navigated to concrete chromosomal quick method.Use Oligonucleolide primers of the present invention,, can utilize one group to realize inferior location from specific chromosomal fragment or a large amount of genomic clone by similar approach.Other the similar strategy that can be used for chromosomal localization comprises in situ hybridization, uses the karyomit(e) prescreen and the hybridization preliminary election of the airflow classification of mark, thereby makes up the special cDNA storehouse of karyomit(e).
The cDNA clone is carried out fluorescence in situ hybridization (FISH) with Metaphase Chromosome, can in a step, accurately carry out chromosomal localization.The summary of this technology is referring to Verma etc., Human Chromosomes:a Manual of BasicTechniques, Pergamon Press, New York (1988).
In case sequence is positioned to chromosome position accurately, the physical location of this sequence on karyomit(e) just can be associated with the gene map data.These data for example are found in, V.Mckusick, Mendelian Inheritance in Man (can by with the online acquisition of Johns Hopkins University Welch Medical Library).Can pass through linkage analysis then, determine gene and navigated to relation between the disease on the chromosomal region already.
Then, need to measure ill and not cDNA between diseased individuals or genome sequence difference.If observe certain sudden change in some or all of diseased individuals, and this sudden change is not observed in any normal individual, then this sudden change may be the cause of disease of disease.More ill and diseased individuals not is usually directed at first seek the variation of structure in the karyomit(e), as from the horizontal visible of karyomit(e) or use based on detectable disappearance of the PCR of cDNA sequence or transposition.
Pyrenoids thuja acid full length sequence or its fragment with promotion 3T3 cell transformation function of the present invention can obtain with the method for pcr amplification method, recombination method or synthetic usually.For the pcr amplification method, can be disclosed according to the present invention about nucleotide sequence, especially open reading frame sequence designs primer, and with commercially available cDNA storehouse or by the prepared cDNA storehouse of ordinary method well known by persons skilled in the art as template, amplification and must relevant sequence.When sequence is longer, usually needs to carry out twice or pcr amplification repeatedly, and then the fragment that each time amplifies is stitched together by proper order.
In case obtained relevant sequence, just can obtain relevant sequence in large quantity with recombination method.This normally is cloned into carrier with it, changes cell again over to, separates obtaining relevant sequence then from the host cell after the propagation by ordinary method.
In addition, also the method for available synthetic is synthesized relevant sequence, especially fragment length more in short-term.Usually, by first synthetic a plurality of small segments, and then connect and to obtain the very long fragment of sequence.
At present, can be fully come the dna sequence dna of code book invention albumen (or its fragment, or derivatives thereof) by chemosynthesis.This dna sequence dna can be introduced then in the various dna moleculars (as carrier) and cell in this area.In addition, also can will suddenly change and introduce in the protein sequence of the present invention by chemosynthesis.
In addition, because the albumen with promotion 3T3 cell transformation function of the present invention has the natural acid sequence that is derived from the people, therefore, compare with the albumen of the same clan that derives from other species, estimate to have higher active and/or lower side effect (for example in the intravital immunogenicity of people lower or do not have) being applied to man-hour.
Below in conjunction with specific embodiment, further set forth the present invention.Should be understood that these embodiment only to be used to the present invention is described and be not used in and limit the scope of the invention.The experimental technique of unreceipted actual conditions in the following example, usually according to people such as normal condition such as Sambrook, molecular cloning: laboratory manual (New York:Cold Spring Harbor LaboratoryPress, 1989) condition described in, or the condition of advising according to manufacturer.Notice that in Nucleotide and amino acid composite sequence, what (1) provided is the position that initial sum stops first Nucleotide of coding, (2) molecular weight unit is dalton.
The acquisition of embodiment 1:cDNA gene and the promoter action that the 3T3 cell clone is formed
PP12719, PP13181, PP13191, PP13479, PP13439, PP13842, PP14673 and PP14776 obtain by making up the human placenta cDNA library with ordinary method.Get the placenta tissue at 3,6,10 monthly ages, (GIBCO BRL company) extracts total RNA by manufacturer's specification sheets with Trizol reagent, extracts mRNA with the mRNA test kit (Pharmacia company) of purifying.Make up the cDNA library of above-mentioned mRNA with pCMV-script TMXR cDNA library construction test kit (Stratagene company).Wherein ThermoScript II is used MMLV-RT-Superscript II (GIBCOBRL) instead, and reverse transcription reaction carries out at 42 ℃.Transform XL 10-Gold recipient cell, obtained 1 * 10
6The cDNA library of cfu/ μ g titre.The first round is picking cDNA clone at random, is probe with high abundance cDNA clone with the cDNA clone who has proved cancer inhibitor cell growth function thereafter, screening by hybridization cDNA library, weak positive and negative clone of picking.With Qiagen 96 orifice plate plasmid extraction test kits, carry out the extraction of plasmid DNA by shop instruction.Plasmid DNA and empty carrier transfection simultaneously 3T3cells.After the 100ng DNA alcohol precipitation drying, add 6 μ l H
2Transfection is treated in the O dissolving.Add 0.74 μ l liposome and 9.3 μ l serum-free mediums in every part of DNA sample, behind the mixing, room temperature was placed 10 minutes.Add 150 μ l serum-free mediums in every pipe, divide equally and add 3 holes and grow in the 3T3 cell of 96 orifice plates, placed 2 hours for 37 ℃, every hole adds 50 μ l serum-free mediums again, 37 ℃ 24 hours.Every hole is changed 100 μ l and is trained liquid entirely, 37 ℃ 24 hours, change the full training liquid 100 μ l that contain G418,37 ℃ 24-48 hour, the limit is observed, the training liquid that G418 concentration does not wait is changed on the limit.After about 2-3 time, there is the clone to form up to the microscopy cell, counting.Find that above clone has the cell clone of promotion formation effect, the result is as shown in the table.
The clone of cDNA clone's transfectional cell (3T3) forms situation
CDNA clones title | CDNA clones number (three repetitions) | Empty carrier clone number (three repetitions) |
????PP12719 ????PP13181 ????PP13191 ????PP13439 ????PP13479 ????PP13842 ????PP14673 ????PP14776 | ????52????51????53 ????44????46????43 ????43????43????40 ????69????56????62 ????44????49????40 ????42????38????24 ????58????57????62 ????46????43????41 | ????27????29????30 ????27????29????30 ????27????29????30 ????27????29????30 ????27????29????30 ????27????29????30 ????27????29????30 ????27????29????30 |
The cDNA clone is adopted two deoxidation cessation method, on the ABI377 automatic dna sequencer, measure the nucleotide sequence of the nearly 500bp of one end.After the analysis, be defined as novel gene cloning, carry out the other end order-checking.For obtaining full length cDNA sequence not yet, the design primer checks order once more, up to obtaining full length sequence (SEQ ID NO:1,4,7,10,13,16,19,22).
Embodiment 2: PCR obtains full-length gene from placenta cDNA:
Get the placenta tissue at 3,6,10 monthly ages, (GIBCO BRL company) extracts total RNA by manufacturer's specification sheets with Trizol reagent, extracts mRNA with the mRNA test kit (Pharmacia company) of purifying.With MMLV-RT-SuperscriptII (GIBCO BRL), ThermoScript II is carried out reverse transcription reaction at 42 ℃, obtains placenta cDNA.Utilize the different primer of commentaries on classics (as shown in the table) of each gene, by 97 ℃ of 3 minutes, 1 circulations; 94 ℃ 30 seconds → 60 ℃ 30 seconds → 72 ℃ 1 minute, totally 35 circulations; 72 ℃ 10 minutes, pcr amplification is carried out in 1 circulation, obtains to contain the amplified production of each protein gene of complete open reading frame sequence.Amplified production is through sequence verification, and the sequence that records with embodiment 1 conforms to, and changes amplified production over to host cell with routine techniques subsequently, obtains recombinant protein (SEQ ID NO:2,5,8,11,14,17,20,23).
Gene specific primer
Embodiment 3:cDNA cloned sequence is analyzed
Clone's title | Special primer 1 (5 ' → 3 ') | Special primer 2 (5 ' → 3 ') |
??PP12719 ??PP13181 ??PP13191 ??PP13439 ??PP13479 ??PP13842 ??PP14673 ??PP14776 | ????CTGGAAACCCTGAAGCTGAG ????GAGATGCCGTGGACTTCCT ????CCATGGAGAAGGAGCAGTGA ????CCATAGCTGGTGTCCTCCTG ????CACCGTGTCCTTCCCTCC ????CTCAGAGGGATGGGATTGG ????GGTACGTGAAGCTGAAAGCC ????ACTGCCATCCCACTGAAACT | ????CCAATCAGAGCCAACCTAGC ????ACCCGACTCGTAGGTGAGC ????CACCAGGACTGGCTGCCT ????GGGCCAGTTCCATCCTCTTA ????GCCTCTGTGATCCTTTCCCT ????GGCCAAAAAGAAAAGGGAGA ????GATAACAGCTCCTGGAAGGC ????CCTCAAGCAATACACCCACC |
1.?PP12719
A: the nucleotide sequence (SEQ ID NO: 1) Length: 1179
1 GTGGGATTAC AGGCGTGAGC CACCACCACA CCCAGCCCTG GGAATGGAGT CTTGATTCTT
61 CTCTGCCCCT CACTGATTCT TCCAAACTGG AAACCCTGAA GCTGAGAGCC CAGCATGGTT
121 CCTGGCAAAC AGCAGGCACT CAAATATTGA TTGGTTTACT GTATGACTAG TAGAGACCCC
181 AACGAGCAAA ACTGTGGCCT AATAAAATTC TGGCTCCTCT CCCAGACTTC CCCTCCCTTT
241 GAGAAATGCC AGAAGCTTCT TAGGGAGGCT CTTGCCAACC TAGACATCAC AGGCACTCAT
301 GGGGCAGCTC CAGCCTCTTC CTCCTGTCAT CACCATAATG CATCCATATC TACAATATGG
361 CAAATTTCAT ATCCTTCCAA CCTCTTTCCT GCATTATTGA TGGGCTGTGT GCACTTTTTA
421 AAAAATCAAT TAGATCAGGG CGTGGAGCTG GAGTTCAAAG AAGCCTTTAA AAGTCTGCTC
481 TTCTGTTTTG CTGTTTTGAA TAGGCACAGA TAAAGCTTTC CCTCTGGTTT GAATAAGCCA
541 AGCTCAGTGC TAGGTTGGCT CTGATTGGCC AGGACTAGGA AAATGCGGTT AAGATGCAAA
601 CACAAGCAAA TATAACCCAG TATCTCTGTG GCCATTACTA AGCTAAGGCA GCAGGACCTG
661 GAGCCTCCTG CTTTGGAGTG GTTCTTCAAT ACTGCTGCTG CTTACGCGCC GGGAAACTGG
721 GAAGGCTGGT GAGCGAGAAG GCAAGGTAAG GTCTCTGATT TACGGGGGCA TGCCAGTTAA
781 TCCTCCTGAA TGAGGAAGAA ATGAAAGGAA GAGGAGCTTG AGAGTCCCTT GGCTTTGTCT
841 TCTGAGATGA GGCTTTTAAA ATGAACCAGG AGTCTGGCTG GCCAGTTTTG CAAACTTCTT
901 GTTCAGAAGA ACCCCTTAGA GGCAGCTGAA CATATAGGGC ACATTTTCAA AAGCTGGGAA
961 AGACAGACCC CTTTCACCTA TCCCTAGAGA AAGATGGTAT GGAGCAAGGC AGGAGGAGTT
1021 AAAACCAGCT TCCTGGCCCA GAGAGGTGGC TTACACCTGT AATCCCAACA CTTTGGGAAG
1081 CCAGGGCATG AGGATTGTTT GAGCCCAGGA GTTTGTAACT TGTGACCATC CTGGGCAACA
1141 TAGTGAGACC CTGTCTCTAC AAAAAAAAAA AAAAAAAAA
B: Nucleotide sequence (SEQ ID NO: 2) Length: 116
1 MTSRDPNEQN CGLIKFWLLS QTSPPFEKCQ KLLREALANL DITGTHGAAP ASSSCHHHNA
61 SISTIWQISY PSNLFPALLM GCVHFLKNQL DQGVELEFKE AFKSLLFCFA VLNRHR
Nucleotide and amino acid sequence of C. Combination (SEQ ID NO: 3) Clone: PP12719
Start codon: 163 ATG termination codon: 511 TAA Protein Weight: 13057.34
1 GTG GGA TTA CAG GCG TGA GCC ACC ACC ACA CCC AGC CCT GGG AAT GGA 48
49 GTC TTG ATT CTT CTC TGC CCC TCA CTG ATT CTT CCA AAC TGG AAA CCC 96
97 TGA AGC TGA GAG CCC AGC ATG GTT CCT GGC AAA CAG CAG GCA CTC AAA 144
145 TAT TGA TTG GTT TAC TGT ATG ACT AGT AGA GAC CCC AAC GAG CAA AAC 192
1 Met Thr Ser Arg Asp Pro Asn Glu Gln Asn 10
193 TGT GGC CTA ATA AAA TTC TGG CTC CTC TCC CAG ACT TCC CCT CCC TTT 240
11 Cys Gly Leu Ile Lys Phe Trp Leu Leu Ser Gln Thr Ser Pro Pro Phe 26
241 GAG AAA TGC CAG AAG CTT CTT AGG GAG GCT CTT GCC AAC CTA GAC ATC 288
27 Glu Lys Cys Gln Lys Leu Leu Arg Glu Ala Leu Ala Asn Leu Asp Ile 42
289 ACA GGC ACT CAT GGG GCA GCT CCA GCC TCT TCC TCC TGT CAT CAC CAT 336
43 Thr Gly Thr His Gly Ala Ala Pro Ala Ser Ser Ser Cys His His His 58
337 AAT GCA TCC ATA TCT ACA ATA TGG CAA ATT TCA TAT CCT TCC AAC CTC 384
59 Asn Ala Ser Ile Ser Thr Ile Trp Gln Ile Ser Tyr Pro Ser Asn Leu 74
385 TTT CCT GCA TTA TTG ATG GGC TGT GTG CAC TTT TTA AAA AAT CAA TTA 432
75 Phe Pro Ala Leu Leu Met Gly Cys Val His Phe Leu Lys Asn Gln Leu 90
433 GAT CAG GGC GTG GAG CTG GAG TTC AAA GAA GCC TTT AAA AGT CTG CTC 480
91 Asp Gln Gly Val Glu Leu Glu Phe Lys Glu Ala Phe Lys Ser Leu Leu 106
481 TTC TGT TTT GCT GTT TTG AAT AGG CAC AGA TAA AGC TTT CCC TCT GGT 528
107 Phe Cys Phe Ala Val Leu Asn Arg His Arg *** 117
529 TTG AAT AAG CCA AGC TCA GTG CTA GGT TGG CTC TGA TTG GCC AGG ACT 576
577 AGG AAA ATG CGG TTA AGA TGC AAA CAC AAG CAA ATA TAA CCC AGT ATC 624
625 TCT GTG GCC ATT ACT AAG CTA AGG CAG CAG GAC CTG GAG CCT CCT GCT 672
673 TTG GAG TGG TTC TTC AAT ACT GCT GCT GCT TAC GCG CCG GGA AAC TGG 720
721 GAA GGC TGG TGA GCG AGA AGG CAA GGT AAG GTC TCT GAT TTA CGG GGG 768
769 CAT GCC AGT TAA TCC TCC TGA ATG AGG AAG AAA TGA AAG GAA GAG GAG 816
817 CTT GAG AGT CCC TTG GCT TTG TCT TCT GAG ATG AGG CTT TTA AAA TGA 864
865 ACC AGG AGT CTG GCT GGC CAG TTT TGC AAA CTT CTT GTT CAG AAG AAC 912
913 CCC TTA GAG GCA GCT GAA CAT ATA GGG CAC ATT TTC AAA AGC TGG GAA 960
961 AGA CAG ACC CCT TTC ACC TAT CCC TAG AGA AAG ATG GTA TGG AGC AAG 1008
1009 GCA GGA GGA GTT AAA ACC AGC TTC CTG GCC CAG AGA GGT GGC TTA CAC 1056
1057 CTG TAA TCC CAA CAC TTT GGG AAG CCA GGG CAT GAG GAT TGT TTG AGC 1104
1105 CCA GGA GTT TGT AAC TTG TGA CCA TCC TGG GCA ACA TAG TGA GAC CCT 1152
1153 GTC TCT ACA AAA AAA AAA AAA AAA AAA 1179
2. PP13181
A: the nucleotide sequence (SEQ ID NO: 4) Length: 1652
1 GCGGCCCCCC TCTGAGGGCG AGTTCATCGA CTGCTTCCAG AAAATCAAGC TGGCGATTAA
61 CTTGCTGGTG GGTCCGGTGG CCCCAGCCCT GCCCCACTGT CTGTGCTGAG GGGAGGGTGG
121 AGGCCCCGCC CCGCCCCGGC ACCTGCTCAC TTGTTCCCAC CCCCAGGCAA AGCTGCAGAA
181 GCACATCCAG AACCCCAGCG CCGCGGAGCT CGTGCACTTC CTCTTCGGGC CTCTGGACCT
241 GGTGCCTGGG GCCGGGCGGC AGGGGCGCGC AGGGTGGGGG CCCAGAGGCC TCTGCAGCAT
301 CTCCCCGGGG TCGGGGTTGG GGCAGCAGGT GCCCGCCTTG GGCAGCCCGG TTCACGCTGT
361 GTGGCCACTC TCCTGGGGTC CAAAGTCCCT TCCCGAGGGC CAGCCTGTGG AGCTACGGGG
421 GTGCTGGGCC AGGGTCTGTG GGCCTCAGTC CCCTCTGAAC CTCACTGTGC CCCAGATCGT
481 CAACACCTGC AGTGGCCCAG ACATCGCACG CTCCGTCTCC TGCCCACTGC TCTCCCGAGA
541 TGCCGTGGAC TTCCTGCGCG GCCACCTGGT CCCTAAGGAG ATGTCGCTGT GGGAGTCACT
601 GGGAGAGAGC TGGATGCGGC CCCGTTCCGA GTGGCCGCGG GAGCCACAGG TGCCCCTCTA
661 CGTGCCCAAG TTCCACAGCG GCTGGGAGCC TCCTGTGGAT GTGCTGCAGG AGGCCCCCTG
721 GGAGGTGGAG GGGCTGGCGT CTGCCCCCAT CGAGGAGGTG AGTCCAGTGA GCCGACAGTC
781 CATAAGAAAC TCCCAGAAGC ACAGCCCCAC TTCAGAGCCC ACCCCCCCGG GGGATGCCCT
841 ACCACCAGTC AGCTCCCCAC ATACTCACAG GGGCTACCAG CCAACACCAG CCATGGCCAA
901 GTACGTCAAG ATCCTGTATG ACTTCACAGC CCGAAATGCC AACGAGCTAT CGGTGCTCAA
961 GGATGAGGTC CTAGAGGTGC TGGAGGACGG CCGGCAGTGG TGGAAGCTGC GCAGCCGCAG
1021 CGGCCAGGCG GGGTACGTGC CCTGCAACAT CCTAGGCGAG GCGCGACCGG AGGACGCCGG
1081 CGCCCCGTTC GAGCAGGCCG GTCAGAAAAG TACTGGGGCC CCGCCAGCCC GACCCACAAG
1141 CTACCCCCAA GCTTCCCGGG GAACAAAGAC GAGCTCATGC AGCACATGGA CGAGGTCAAC
1201 GACGAGCTCA TCCGGAAAAT CAGCAACATC AGGGCGCAGC CACAGAGGCA CTTCCGCGTG
1261 GAGCGCAGCC AGCCCGTGAG CCAGCCGCTC ACCTACGAGT CGGGTCCGGA CGAGGTCCGC
1321 GCCTGGCTGG AAGCCAAGGC CTTCAGCCCG CGGATCGTGG AGAACCTGGG CATCCTGACC
1381 GGGCCGCAGC TCTTCTCCCT CAACAAGGAG GAGCTGAAGA AAGTGTGCGG CGAGGAGGGC
1441 GTCCGCGTGT ACAGCCAGCT CACCATGCAG AAGGCCTTCC TGGAGAAGCA GCAAAGTGGG
1501 TCGGAGCTGG AAGAACTCAT GAACAAGTTT CATTCCATGA ATCAGAGGAG GGGGGAGGAC
1561 AGCTAGGCCC AGCTGCCTTG GGCTGGGGCC TGCGGAGGGG AAGCCCACCC ACAATGCATG
1621 GAGTATTATT TTTAAAAAAA AAAAAAAAAA AA
...
B: nucleotide sequence, (SEQ, ID, NO:5), length: 232, 1, MSLWESLGES, WMRPRSEWPR, EPQVPLYVPK, FHSGWEPPVD, VLQEAPWEVE, GLASAPIEEV, 61, SPVSRQSIRN, SQKHSPTSEP, TPPGDALPPV, SSPHTHRGYQ, PTPAMAKYVK, ILYDFTARNA, 121, NELSVLKDEV, LEVLEDGRQW, WKLRSRSGQA, GYVPCNILGE, ARPEDAGAPF, EQAGQKSTGA, 181, PPARPTSYPQ, ASRGTKTSSC, STWTRSTTSS, SGKSATSGRS, HRGTSAWSAA, SP, C. nucleotides and amino acid composite sequence, (SEQ, ID, NO:6), clone number:, PP13181, start code: 581, ATG, stop coding: 1277, TGA, protein molecular weight: 25362.69, 1, G, CGG, CCC, CCC, TCT, GAG, GGC, GAG, TTC, ATC, GAC, TGC, TTC, CAG, AAA, ATC, 46, 47, AAG, CTG, GCG, ATT, AAC, TTG, CTG, GTG, GGT, CCG, GTG, GCC, CCA, GCC, CTG, CCC, 94, 95, CAC, TGT, CTG, TGC, TGA, GGG, GAG, GGT, GGA, GGC, CCC, GCC, CCG, CCC, CGG, CAC, 142, 143, CTG, CTC, ACT, TGT, TCC, CAC, CCC, CAG, GCA, AAG, CTG, CAG, AAG, CAC, ATC, CAG, 190, 191, AAC, CCC, AGC, GCC, GCG, GAG, CTC, GTG, CAC, TTC, CTC, TTC, GGG, CCT, CTG, GAC, 238, 239, CTG, GTG, CCT, GGG, GCC, GGG, CGG, CAG, GGG, CGC, GCA, GGG, TGG, GGG, CCC, AGA, 286, 287, GGC, CTC, TGC, AGC, ATC, TCC, CCG, GGG, TCG, GGG, TTG, GGG, CAG, CAG, GTG, CCC, 334, 335, GCC, TTG, GGC, AGC, CCG, GTT, CAC, GCT, GTG, TGG, CCA, CTC, TCC, TGG, GGT, CCA, 382, 383, AAG, TCC, CTT, CCC, GAG, GGC, CAG, CCT, GTG, GAG, CTA, CGG, GGG, TGC, TGG, GCC, 430, 431, AGG, GTC, TGT, GGG, CCT, CAG, TCC, CCT, CTG, AAC, CTC, ACT, GTG, CCC, CAG, ATC, 478, 479, GTC, AAC, ACC, TGC, AGT, GGC, CCA, GAC, ATC, GCA, CGC, TCC, GTC, TCC, TGC, CCA, 526, 527, CTG, CTC, TCC, CGA, GAT, GCC, GTG, GAC, TTC, CTG, CGC, GGC, CAC, CTG, GTC, CCT, 574, 575, AAG, GAG, ATG, TCG, CTG, TGG, GAG, TCA, CTG, GGA, GAG, AGC, TGG, ATG, CGG, CCC, 622, 1, Met, Ser, Leu, Trp, Glu, Ser, Leu, Gly, Glu, Ser, Trp, Met, Arg, Pro, 14, 623, CGT, TCC, GAG, TGG, CCG, CGG, GAG, CCA, CAG, GTG, CCC, CTC, TAC, GTG, CCC, AAG, 670, 15, Arg, Ser, Glu, Trp, Pro, Arg, Glu, Pro, Gln, Val, Pro, Leu, Tyr, Val, Pro, Lys, 30, 671, TTC, CAC, AGC, GGC, TGG, GAG, CCT, CCT, GTG, GAT, GTG, CTG, CAG, GAG, GCC, CCC, 718, 31, Phe, His, Ser, Gly, Trp, Glu, Pro, Pro, Val, Asp, Val, Leu, Gln, Glu, Ala, Pro, 46, 719, TGG, GAG, GTG, GAG, GGG, CTG, GCG, TCT, GCC, CCC, ATC, GAG, GAG, GTG, AGT, CCA, 766, 47, Trp, Glu, Val, Glu, Gly, Leu, Ala, Ser, Ala, Pro, Ile, Glu, Glu, Val, Ser, Pro, 62, 767, GTG, AGC, CGA, CAG, TCC, ATA, AGA, AAC, TCC, CAG, AAG, CAC, AGC, CCC, ACT, TCA, 814, 63, Val, Ser, Arg, Gln, Ser, Ile, Arg, Asn, Ser, Gln, Lys, His, Ser, Pro, Thr, Ser, 78, 815, GAG, CCC, ACC, CCC, CGG, GGG, GAT, GCC, CTA, CCA, CCA, GTC, AGC, TCC, CCA, CAT, 862, 79, Glu, Pro, Thr, Pro, Pro, Gly, Asp, Ala, Leu, Pro, Pro, Val, Ser, Ser, Pro, His, 94, 863, ACT, CAC, AGG, GGC, TAC, CAG, CCA, ACA, CCA, GCC, ATG, GCC, AAG, TAC, GTC, AAG, 910, 95, Thr, His, Arg, Gly, Tyr, Gln, Pro, Thr, Pro, Ala, Met, Ala, Lys, Tyr, Val, Lys, 110, 911, ATC, CTG, TAT, GAC, TTC, ACA, GCC, CGA, AAT, GCC, AAC, GAG, CTA, TCG, GTG, CTC, 958, 111, Ile, Leu, Tyr, Asp, Phe, Thr, Ala, Arg, Asn, Ala, Asn, Glu, Leu, Ser, Val, Leu, 126, 959, AAG, GAT, GAG, GTC, CTA, GAG, GTG, CTG, GAG, GAC, GGC, CGG, CAG, TGG, TGG, AAG, 1006, 127, Lys, Asp, Glu, Val, Leu, Glu, Val, Leu, Glu, Asp, Gly, Arg, Gln, Trp, Trp, Lys, 1421007, CTG, CGC, AGC, CGC, AGC, GGC, CAG, GCG, GGG, TAC, GTG, CCC, TGC, AAC, ATC, CTA, 1054, 143, Leu, Arg, Ser, Arg, Ser, Gly, Gln, Ala, Gly, Tyr, Val, Pro, Cys, Asn, Ile, Leu, 1581055, GGC, GAG, GCG, CGA, CCG, GAG, GAC, GCC, GGC, GCC, CCG, TTC, GAG, CAG, GCC, GGT, 1102, 159, Gly, Glu, Ala, Arg, Pro, Glu, Asp, Ala, Gly, Ala, Pro, Phe, Glu, Gln, Ala, Gly, 174, 1103, CAG, AAA, AGT, ACT, GGG, GCC, CCG, CCA, GCC, GGA, CCC, ACA, AGC, TAC, CCC, CAA, 1150, 175, Gln, Lys, Ser, Thr, Gly, Ala, Pro, Pro, Ala, Arg, Pro, Thr, Ser, Tyr, Pro, Gln, 190, 1151, GCT, TCC, CGG, GGA, ACA, AAG, ACG, AGC, TCA, TGC, AGC, ACA, TGG, AGG, AGG, TCA, 1198, 191, Ala, Ser, Arg, Gly, Thr, Lys, Thr, Ser, Ser, Cys, Ser, Thr, Trp, Thr, Arg, Ser, 206, 1199, AGG, AGG, AGC, TCA, TCC, GGA, AAA, TCA, GCA, ACA, TCA, GGG, CGC, AGC, CAC, AGA, 1246, 207, Thr, Thr, Ser, Ser, Ser, Gly, Lys, Ser, Ala, Thr, Ser, Gly, Arg, Ser, His, Arg, 222, 1247, GGC, ACT, TCC, GCG, TGG, AGC, GCA, GCC, AGC, CCG, TGA, GCC, AGC, CGC, TCA, CCT, 1294, 223, Gly, Thr, Ser, Ala, Trp, Ser, Ala, Ala, Ser, Pro, * *, 233, 1295, AGG, AGT, CGG, GTC, CGG, ACG, AGG, TCC, GCG, CCT, GGC, TGG, AAG, CCA, AGG, CCT, 1342, 1343, TCA, GCC, CGC, GGA, TCG, TGG, AGA, ACC, TGG, GCA, TCC, TGA, CCG, GGC, CGC, AGC, 1390, 1391, TCT, TCT, CCC, TCA, ACA, AGG, AGG, AGC, TGA, AGA, AAG, TGT, GCG, GCG, AGG, AGG, 1438, 1439, GCG, TCC, GCG, TGT, ACA, GCC, AGC, TCA, CCA, TGC, AGA, AGG, CCT, TCC, TGG, AGA, 1486, 1487, AGC, AGC, AAA, GTG, GGT, CGG, AGC, TGG, AAG, AAC, TCA, TGA, ACA, AGT, TTC, ATT, 1534, 1535, CCA, TGA, ATC, AGA, GGA, GGG, GGG, AGG, ACA, GCT, AGG, CCC, AGC, TGC, CTT, GGG, 1582, 1583, CTG, GGG, CCT, GCG, GAG, GGG, AAG, CCC, ACC, CAC, AAT, GCA, TGG, AGT, ATT, ATT, 1630, 1631, TTT, AAA, AAA, AAA, AAA, AAA, AAA, A, 1652
3.PP13191
A: the nucleotide sequence (SEQ ID NO: 7) Length: 1489
1 GCCTGTCCAC ACCTCTGCCC CAGAGCTGCC TCCTGCCTGG CACTGCCGCC ACACTCCCCT
61 CCTGGGATGG GGCTTCTGCT CCCGGGCTCA CTCAAGGAGA CTGCGGCATG TTGACCACAC
121 CAGACTGGGT TTCAGGGAAT GGGCATGCCA GGTGCCAAGG AGCCAAACAG ATGGCTTTCC
181 AGGCAGCAAG GTCCTTGGGG CCTTCTTGGA GGAGCTTGGG TGACAGCCAG GTGAGCACCC
241 AGACCCCAGA CCCTCATGTG CTGTGTGCCT GGCCCCTTCT GTACTGGCCA TTTGTGGCCA
301 GGGCCAAGCC TGTGACTCAA CTCCAGGGGC AAGATGGGGA GTGAGCTGAT GGCTCCGAGA
361 CTGGTCAGGA GCCCAGGCCA GTGAGATGGG GCCTGGAGCC TTGTCTGTGT CACATTAGGT
421 ACCATGGGAG CTGCTGAGAC CTGACATTTT GTCCCCTGCC TACATGGCTT GGCCCATGGA
481 GAAGGAGCAG TGAATGGGAT CGTCGGGGAA GCCCCTCTTC CTGCTCTGCT CCCCTGGAAA
541 CTGTTGCAAA ACTCCCACCG CCTCATGGCA AATGCCCAAA GCATGTTCCG CACCCAGGCG
601 GGGGCCCCTG CTAATGAGAA CCTTGGTGCA GCTGCAGCCA GGAGGGGAGC GGGCCCAGGA
661 GCCAGGCTCA GGTCCAGCTG GTTCCTCTCT GGCGCCTTCT GAACCCGTCT CAGCAGGTCC
721 ACAGCACCTG GGCAGAGGTC AGAGACCAGG GGAGGCCGGG CCTTGCCCTC CCTTCTGCCC
781 AGGGCCCAGT GTTCTTGATA GAAGACCCTT CTGGGGAGCC AGGGAGCTCA GGGGACAGAT
841 AAGGGAAGGA CGCCCCCTGA CTCCAGGCCC CTGAGCCTGG CGGGAAGTGG CTGCGGCCCA
901 GGCAGCCAGT CCTGGTGGTG TTCTCCCTGC ATGCCCTCCG TGGCTGGGCT GCCACCCCAC
961 CCGGCCGGAA TCTGTCTTGA CCTGCAGGAA TACACGGGCG GCGCCAGGCA TTACCTCACA
1021 GCGGGACTAC ACAGTTGCTG GCTTTGCTCC TGGGCAAGGA GGAGCAGGCC AGAGCCTCTT
1081 TTGCTTCCTT TTCTTGCCCA TGCGGCTTCT AGAAGCCAGG CACAGGTTGC CAAGAGGTGA
1141 CACGAAACAG GAGGAAACTC AGTGACCTCT GCCTCTCCCA CATTCCTCCC CGCGGGGGAG
1201 GACCTCGCCG CTCTGAAGAG CACCGTGCAC ATGTGGGTGC ACAAACGTGG GTGTTGGTGT
1261 GGAGGGGGCG CAGATCTCGG TGGATGAACT GCGTCTGGAC TCTTAGATTC ATAAAATATT
1321 CGAGGGTTTG GGAGTCACAG ACCCTCCCCT CTCCTCAGTG CACTTTAGCA TTTGCACGGT
1381 GTCTTCCCCG GACAGCACAG CAATAAATGG TGTGATTGCG TGGAAAAAAA AAAAAAAAAA
1441 AAAAAAAAAA TAAAAAAAAA AAAAAAAAAA AAAAAAAAAA AAAAAAAAA
B: Nucleotide sequence (SEQ ID NO: 8) Length: 126
1 MGSSGKPLFL LCSPGNCCKT PTASWQMPKA CSAPRRGPLL MRTLVQLQPG GERAQEPGSG
61 PAGSSLAPSE PVSAGPQHLG RGQRPGEAGP CPPFCPGPSV LDRRPFWGAR ELRGQIREGR
121 PLTPGP
Nucleotide and amino acid sequence of C. Combination (SEQ ID NO: 9) Clone: PP13191
Start codon: 494 ATG termination codon: 872 TGA Protein Weight: 13131.41
1 G CCT GTC CAC ACC TCT GCC CCA GAG CTG CCT CCT GCC TGG CAC TGC 46
47 CGC CAC ACT CCC CTC CTG GGA TGG GGC TTC TGC TCC CGG GCT CAC TCA 94
95 AGG AGA CTG CGG CAT GTT GAC CAC ACC AGA CTG GGT TTC AGG GAA TGG 142
143 GCA TGC CAG GTG CCA AGG AGC CAA ACA GAT GGC TTT CCA GGC AGC AAG 190
191 GTC CTT GGG GCC TTC TTG GAG GAG CTT GGG TGA CAG CCA GGT GAG CAC 238
239 CCA GAC CCC AGA CCC TCA TGT GCT GTG TGC CTG GCC CCT TCT GTA CTG 286
287 GCC ATT TGT GGC CAG GGC CAA GCC TGT GAC TCA ACT CCA GGG GCA AGA 334
335 TGG GGA GTG AGC TGA TGG CTC CGA GAC TGG TCA GGA GCC CAG GCC AGT 382
383 GAG ATG GGG CCT GGA GCC TTG TCT GTG TCA CAT TAG GTA CCA TGG GAG 430
431 CTG CTG AGA CCT GAC ATT TTG TCC CCT GCC TAC ATG GCT TGG CCC ATG 478
479 GAG AAG GAG CAG TGA ATG GGA TCG TCG GGG AAG CCC CTC TTC CTG CTC 526
1 Met Gly Ser Ser Gly Lys Pro Leu Phe Leu Leu 11
527 TGC TCC CCT GGA AAC TGT TGC AAA ACT CCC ACC GCC TCA TGG CAA ATG 574
12 Cys Ser Pro Gly Asn Cys Cys Lys Thr Pro Thr Ala Ser Trp Gln Met 27
575 CCC AAA GCA TGT TCC GCA CCC AGG CGG GGG CCC CTG CTA ATG AGA ACC 622
28 Pro Lys Ala Cys Ser Ala Pro Arg Arg Gly Pro Leu Leu Met Arg Thr 43
623 TTG GTG CAG CTG CAG CCA GGA GGG GAG CGG GCC CAG GAG CCA GGC TCA 670
44 Leu Val Gln Leu Gln Pro Gly Gly Glu Arg Ala Gln Glu Pro Gly Ser 59
671 GGT CCA GCT GGT TCC TCT CTG GCG CCT TCT GAA CCC GTC TCA GCA GGT 718
60 Gly Pro Ala Gly Ser Ser Leu Ala Pro Ser Glu Pro Val Ser Ala Gly 75
719 CCA CAG CAC CTG GGC AGA GGT CAG AGA CCA GGG GAG GCC GGG CCT TGC 766
76 Pro Gln His Leu Gly Arg Gly Gln Arg Pro Gly Glu Ala Gly Pro Cys 91
767 CCT CCC TTC TGC CCA GGG CCC AGT GTT CTT GAT AGA AGA CCC TTC TGG 814
92 Pro Pro Phe Cys Pro Gly Pro Ser Val Leu Asp Arg Arg Pro Phe Trp 107
815 GGA GCC AGG GAG CTC AGG GGA CAG ATA AGG GAA GGA CGC CCC CTG ACT 862
108 Gly Ala Arg Glu Leu Arg Gly Gln Ile Arg Glu Gly Arg Pro Leu Thr 123
863 CCA GGC CCC TGA GCC TGG CGG GAA GTG GCT GCG GCC CAG GCA GCC AGT 910
124 Pro Gly Pro *** 127
911 CCT GGT GGT GTT CTC CCT GCA TGC CCT CCG TGG CTG GGC TGC CAC CCC 958
959 ACC GGG CCC GAA TCT GTC TTG ACC TGC AGG AAT ACA CGG GCG GCG CCA 1006
1007 GGC ATT ACC TCA CAG CGG GAC TAC ACA GTT GCT GGC TTT GCT CCT GGG 1054
1055 CAA GGA GGA GCA GGC CAG AGC CTC TTT TGC TTC CTT TTC TTG CCC ATG 1102
1103 CCG CTT CTA GAA GCC ACG CAC AGG TTG CCA AGA GGT GAC ACG AAA CAG 1150
1151 GAG GAA ACT CAG TGA CCT CTG CCT CTC CCA CAT TCC TCC CCG CGG GGG 1198
1199 AGG ACC TCG CCG CTC TGA AGA GCA CCG TGC ACA TGT GGG TGC ACA AAC 1246
1247 GTG GGT GTT GGT GTG GAC GGG GGC CAG ATC TCC GTG GAT GAA CTG GGT 1294
1295 CTG GAC TCT TAG ATT CAT AAA ATA TTC GAG GGT TTG GGA GTC ACA GAC 1342
1343 CCT CCC CTC TCC TCA GTG CAC TTT AGC ATT TGC ACG GTG TCT TCC CCG 1390
1391 GAC AGC ACA GCA ATA AAT GGT GTG ATT GCG TGG AAA AAA AAA AAA AAA 1438
1439 AAA AAA AAA AAA TAA AAA AAA AAA AAA AAA AAA AAA AAA AAA AAA AAA 1486
1487 AAA 1489
4.PP13439
A: the nucleotide sequence (SEQ ID NO: 10) Length: 2935
1 GCCAGCCCGC AGAGCACGGT CCTGGGGGTG TGATCACAGC TGCTGACCGC GGCTCCCGCC
61 GTGTGATGTG ACCACTCGGA GGTGGGGCGA GGGGGCCGGG GGCCAGCAGC AGGGAGTGTG
121 TAAAAGGTCT TCTTCCTCAC AATACGATTG AAGGTGTTTG GGAGGTAGGG CTCAGATCCA
181 GGAGTGCCGT CCTCCAAGCC TTCCATAGGA TTTTGTGACG CCCTTGAGAA CCGCCCTTGT
241 AGACTCAGTC CTGAACTACC GCCACGCCTG TATTTGCTCC TCCTCATCTC GGATTATTGC
301 TTTCTTTACT TCCTCTGTGA AGCACTAAAT ACTAGTAGCT TTTTAAGAAC AGACCACAAC
361 AGTTTCTTCG TTTGACCATT ACTAAGTCTG CCTCCAAAAT GGGGAGTTCC GTTAACTTTA
421 TCAGCCAGCC CTGCCAATTT TAGTTAAGAT AAGCCTTCGT GGGCCTCCAA AGGCCTCTGA
481 GAGACTAGGT ACCCAGAGAC CTCTGAGGTA TGTTAAATGG TTTAGTTCGT GGTGGTGCAA
541 TGAAAACTTA AAGAAGGAAA GGAATCTGCA ATTTGAACTG TTAACAGACC CCACATTTAC
601 ACACTGCCTA TTTGAATAAG AATCTCTTTC AGTATTAAAT GTTTTGTGAC ACACAAATGA
661 ATTTGTACAC AGTCTTTTAG CTTGAATTTG TAGACAGCCT TTAAATGTTG TTTGGGGATC
721 TTTGGACTAA TTGTATAAGC GCTCAGTCTG GTCTCAAACT CATGATCTCA GGTTATCCAT
781 GCGCCTCGGC CTCCGAAAAT GCTGCGATTA TAGGCGTGAG CCACCACTCC CAGCCCCTTC
841 TTGTGATGAT GTGAGATGAT AAAATGCCTG TGTGAAGATG AAGTGAGGCG AATAACCTAG
901 GCAGTGTGAC ATAAGTGGAA AACGCCAGAA ATAAACAATT CATGTTATAA ATTGTGCACC
961 ATTCTGAGTA GTGTGATGAA ATCTTGCACT CCTCCATTCC ATCCTGCCAG CATTGCCAGT
1021 CACTCGGTGG CCATCTCTGT GATCAGATTA ACAGTGGTTT TATCAGTATC ATAGGGCTTA
1081 TGTTCAAGTA ACTGTATTTT ACTTAATAAT GGCTACAAAG TGCAAAAGTA GCGATATAAT
1141 ATTTTTGGAC CATGGTTGAC TGCAGGTAAC TGAAATTTTA GTTACATGAA ACCACAGATA
1201 AGGGGGGACT GCACTACACA TAAAATAGGC ATAAGTAACC TCAAGACAAT AGACATTATT
1261 AAAATATAGA AAAGTAATAA AGAAGTGATA AATGGAGAAC TTTTGCTTTA ATATAATTTA
1321 TTGTAAGTTT ATATAATTTA ATTTTTAGTA ACTTCAATTT TTAATATTTT AATTTTTAAT
1381 GATTTATGTG TTTAACAACT GACTCACAAA ATTGCTGAAA ATTTCCCAAT CTTGATGAAA
1441 ATTTCCCAGT CTGCTCTCAG GAGCCTATAT GAGCCAGCCC AGAATACCAC TGGAGTTATA
1501 ATTTTAGAAG ATTACCTTGG TAGCAATCTG TAGAAATGTT TTGTAGGGGA GCTACAGTAG
1561 GGATCAAAAG TGGAGACAGA TTACGAAATG TCTTCAGACC CTGCCAGCAT GAGGGTTCAA
1621 ATGATGTCTA GTCAGTGGAG ATCACGCAAA TTTCACTGGT TTGTGTTCCC ACGTGGATCC
1681 GTAGGACCTT CAATGCAAGT CTCCTTCAGG TATTCCTGGT GGTATCTTGG CATTGTCTCA
1741 CATGTGTTTT GAGAGCCCCA CTTCCTTGGG GAGTCTTCCG CTTTGGCTTC CTTGTTGATG
1801 ACCGTGGGCC AAAGGGCTTT GTGTGTGAGG CTCCACCTCA GCCATAGCTG GTGTCCTCCT
1861 GAAGAGGATT TATGCCAAAA GGAGTCACAT GCCAGGACTT GTTTTTTTCA AGTTTGCAAA
1921 TCTGGGGGGA GGGTGGGGCT CTAGAAAGAT GCTGGAACCT GGTATGTGCC CTGCAGAAAT
1981 GTCCTTGACT CCTTTTGTCA CCAGGAGCCT AAATTTACTG TGTTTCTCTG TGACTCACAG
2041 AGGCAGGGGG AAAGACAATC CTCTCCACTT CTTCCTGCAG CTACTTGCTC CATCAGCCTC
2101 AGCCAGTCTG GGGCTGGGCA GGGAAACAGG CCCTCTTCCT TCTCCTGGTT TGTCCCCTGG
2161 TTTGCTCAGC GAGACCTTCG TGTCTGAGGG CCCCCTTCTC ACCTTGCAGA TCAGCTGGTG
2221 TGGTACTGAA TGCCTGATGG ATGGCCAGGG GCCCCTGATC CATCTGCATT CCCAGAATAT
2281 AAGAATCTCA TTTGGGGCCA CCCAGCCCCC ACTACTCTTA GTATATGAAA TAAGGTGGAA
2341 TAGATAAGAG GATGGAACTG GCCCCTGGGT GCATGATGGG GCCAATAACG GAGGTACAGG
2401 GTGCATATGT GGCTGGGCTG TGAGACAGGG AAAGAGGATG GAATTGAGAT GCAAGGTAGA
2461 AAAGCATGAC TATGTTCAGT TTGGGACTTG ATGAAATAGA TACTGGTAGG ACAGTTCACT
2521 AGGCACTGGG CCTTCAGGAG AAAGAGAAGA AAGGAATGGT GGCATTGGGA GTAGTAAAGC
2581 TGTGGGTATG AATGAGTATC TATGAGACTC ACCTCCCTGT GAGGACATAG AATGAGGAGA
2641 CACAGTCTAC AGGTAGACTA GGTCGAGTTC AGAAGCCTGA AGAACACCAG TGTTTAAGGG
2701 ATGGTGTTGG GAAAGGGAGC CAGGCCAGCC AGAGAGGAAT GTTCCGGAAC TCTGAAAGGA
2761 AGGAAATGGC AGGAACAAAG GAGCTGGGGC ACAACAGTGG TGACTCACAC TGGGAACACT
2821 TCGGCCCATT GTGTTTATGC TTATAGTTTC CCAAGTTTAT CTTTTCAGGG TTATGATTAC
2881 GTTAACCTCA CCTCCCTCCC TCCCTCTAAA ACAAAACAAA AAAAAAAAAA AAAAA
B: Nucleotide sequence (SEQ ID NO: 11) Length: 152
1 MPGLVFFKFA NLGGGWGSRK MLEPGMCPAE MSLTPFVTRS LNLLCFSVTH RGRGKDNPLH
61 FFLQLLAPSA SASLGLGRET GPLPSPGLSP GLLSETFVSE GPLLTLQISW CGTECLMDGQ
121 GPLIHLHSQN IRISFGATQP PLLLVYEIRW NR
Nucleotide and amino acid sequence of C. Combination (SEQ ID NO: 12) Clone: PP13439
Start codon: 1889 ATG termination codon: 2345 TAA Protein Weight: 16504.36
...
1???G?CCA?GCC?CGC?AGA?GCA?CGG?TCC?TGG?GGG?TGT?GAT?CAC?AGC?TGC?TGA?????46???47?CCG?CGG?CTC?CCG?CCG?TGT?GAT?GTG?ACC?ACT?CGG?AGG?TGG?GGC?GAG?GGG?????94???95?GCC?GGG?GGC?CAG?CAG?CAG?GGA?GTG?TGT?AAA?AGG?TCT?TCT?TCC?TCA?CAA????142??143?TAG?GAT?TGA?AGG?TGT?TTG?GGA?CGT?AGG?GCT?CAG?ATC?CAG?GAG?TGC?CGT????190??191?CCT?CCA?AGC?CTT?CCA?TAG?GAT?TTT?GTG?ACG?CCC?TTG?AGA?ACC?GCC?CTT????238?239?GTA?GAC?TCA?GTC?CTG?AAC?TAC?CGC?CAC?GCC?TGT?ATT?TGC?TCC?TCC?TCA?????286?287?TCT?CGG?ATT?ATT?GCT?TTC?TTT?ACT?TCC?TCT?GTG?AAG?CAC?TAA?ATA?CTA?????334?335?GTA?GCT?TTT?TAA?GAA?CAG?ACC?ACA?ACA?GTT?TCT?TCG?TTT?GAC?CAT?TAC?????382?383?TAA?GTC?TGC?CTC?CAA?AAT?GGG?GAG?TTC?CGT?TAA?CTT?TAT?CAG?CCA?GCC?????430?431?CTG?CCA?ATT?TTA?GTT?AAG?ATA?AGC?CTT?CGT?GGG?CCT?CCA?AAG?GCC?TCT?????478?479?GAG?AGA?CTA?GGT?ACC?CAG?AGA?CCT?CTG?AGG?TAT?GTT?AAA?TGG?TTT?AGT?????526?527?TCG?TGG?TGG?TGC?AAT?GAA?AAC?TTA?AAG?AAG?GAA?AGG?AAT?CTG?CAA?TTT?????574?575?GAA?CTG?TTA?ACA?GAC?CCC?ACA?TTT?ACA?CAC?TGC?CTA?TTT?GAA?TAA?GAA?????622?623?TCT?CTT?TCA?GTA?TTA?AAT?GTT?TTG?TGA?CAC?ACA?AAT?GAA?TTT?GTA?CAC?????670?671?AGT?CTT?TTA?GCT?TGA?ATT?TGT?AGA?CAG?CCT?TTA?AAT?GTT?GTT?TGG?GGA?????718?719?TCT?TTG?GAC?TAA?TTG?TAT?AAG?CGC?TCA?GTC?TGG?TCT?CAA?ACT?CAT?GAT?????766?767?CTC?AGG?TTA?TCC?ATG?CGC?CTC?GGC?CTC?CGA?AAA?TGC?TGC?GAT?TAT?AGG?????814?815?CGT?GAG?CCA?CCA?CTC?CCA?GCC?CCT?TCT?TGT?GAT?GAT?GTG?AGA?TGA?TAA?????862?863?AAT?GCC?TGT?GTG?AAG?ATG?AAG?TGA?GGC?GAA?TAA?CCT?AGG?CAG?TGT?GAC?????910?911?ATA?AGT?GGA?AAA?CGC?CAG?AAA?TAA?ACA?ATT?CAT?GTT?ATA?AAT?TGT?GCA?????958?959?CCA?TTC?TGA?GTA?GTG?TGA?TGA?AAT?CTT?GCA?CTC?CTC?CAT?TCC?ATC?CTG????10061007?CCA?GCA?TTG?CCA?GTC?ACT?CGG?TGG?CCA?TCT?CTG?TGA?TCA?GAT?TAA?CAG????10541055?TGG?TTT?TAT?CAG?TAT?CAT?AGG?GCT?TAT?GTT?CAA?GTA?ACT?GTA?TTT?TAC????11021103?TTA?ATA?ATG?GCT?ACA?AAG?TGC?AAA?AGT?AGC?GAT?ATA?ATA?TTT?TTG?GAC????11501151?CAT?GGT?TGA?CTG?CAG?GTA?ACT?GAA?ATT?TTA?GTT?ACA?TGA?AAC?CAC?AGA????11981199?TAA?GGG?GGG?ACT?GCA?CTA?CAC?ATA?AAA?TAG?GCA?TAA?GTA?ACC?TCA?AGA????12461247?CAA?TAG?ACA?TTA?TTA?AAA?TAT?AGA?AAA?GTA?ATA?AAG?AAG?TGA?TAA?ATG????12941295?GAG?AAC?TTT?TGC?TTT?AAT?ATA?ATT?TAT?TGT?AAG?TTT?ATA?TAA?TTT?AAT????13421343?TTT?TAG?TAA?CTT?CAA?TTT?TTA?ATA?TTT?TAA?TTT?TTA?ATG?ATT?TAT?GTG????13901391?TTT?AAC?AAC?TGA?CTC?ACA?AAA?TTG?CTG?AAA?ATT?TCC?CAA?TCT?TGA?TGA????14381439?AAA?TTT?CCC?AGT?CTG?CTC?TCA?GGA?GCC?TAT?ATG?AGC?CAG?CCC?AGA?ATA????14861487?CCA?CTG?GAG?TTA?TAA?TTT?TAG?AAG?ATT?ACC?TTG?GTA?GCA?ATC?TGT?AGA????15341535?AAT?GTT?TTG?TAG?GGG?AGC?TAC?AGT?AGG?GAT?CAA?AAG?TGG?AGA?CAG?ATT????15821583?ACG?AAA?TGT?CTT?CAG?ACC?CTG?CCA?GCA?TGA?GGG?TTC?AAA?TGA?TGT?CTA????16301631?GTC?AGT?GGA?GAT?CAC?GCA?AAT?TTC?ACT?GGT?TTG?TGT?TCC?CAC?GTG?GAT????16781679?CCG?TAG?GAC?CTT?CAA?TGC?AAG?TCT?CCT?TCA?GGT?ATT?CCT?GGT?GGT?ATC????17261727?TTG?GCA?TTG?TCT?CAC?ATG?TGT?TTT?GAG?AGC?CCC?ACT?TCC?TTG?GGG?AGT????17741775?CTT?CCG?CTT?TGG?CTT?CCT?TGT?TGA?TGA?CCG?TGG?GCC?AAA?GGG?CTT?TGT????18221823?GTG?TGA?GGC?TCC?ACC?TCA?GCC?ATA?GCT?GGT?GTC?CTC?CTG?AAG?AGG?ATT????18701871?TAT?GCC?AAA?AGG?AGT?CAC?ATG?CCA?GGA?CTT?GTT?TTT?TTC?AAG?TTT?GCA????1918???1?????????????????????????Met?Pro?Gly?Leu?Val?Phe?Phe?Lys?Phe?Ala??????101919?AAT?CTG?GGG?GGA?GGG?TGG?GGC?TCT?AGA?AAG?ATG?CTG?GAA?CCT?GGT?ATG????1966??11?Asn?Leu?Gly?Gly?Gly?Trp?Gly?Ser?Arg?Lys?Met?Leu?Glu?Pro?Gly?Met??????261967?TGC?CCT?GCA?GAA?ATG?TCC?TTG?ACT?CCT?TTT?GTC?ACC?AGG?AGC?CTA?AAT????2014??27?Cys?Pro?Ala?Glu?Met?Ser?Leu?Thr?Pro?Phe?Val?Thr?Arg?Ser?Leu?Asn??????422015?TTA?CTG?TGT?TTC?TCT?GTG?ACT?CAC?AGA?GGC?AGG?GGG?AAA?GAC?AAT?CCT????2062??43?Leu?Leu?Cys?Phe?Ser?Val?Thr?His?Arg?Gly?Arg?Gly?Lys?Asp?Asn?Pro??????582063?CTC?CAC?TTC?TTC?CTG?CAG?CTA?CTT?GCT?CCA?TCA?GCC?TCA?GCC?AGT?CTG????2110??59?Leu?His?Phe?Phe?Leu?Gln?Leu?LeuAla?Pro?Ser?Ala?Ser?Ala?Ser?Leu???????742111?GGG?CTG?GGC?AGG?GAA?ACA?GGC?CCT?CTT?CCT?TCT?CCT?GGT?TTG?TCC?CCT????2158??75?Gly?Leu?Gly?Arg?Glu?Thr?Gly?Pro?Leu?Pro?Ser?Pro?Gly?Leu?Ser?Pro??????902159?GGT?TTG?CTC?AGC?GAG?ACC?TTC?GTG?TCT?GAG?GGC?CCC?CTT?CTC?ACC?TTG????2206??91?Gly?Leu?Leu?Ser?Glu?Thr?Phe?Val?Ser?Glu?Gly?Pro?Leu?Leu?Thr?Leu?????1062207?CAG?ATC?AGC?TGG?TGT?GGT?ACT?GAA?TGC?CTG?ATG?GAT?GGC?CAG?GGG?CCC????2254?107?Gln?Ile?Ser?Trp?Cys?Gly?Thr?Glu?Cys?Leu?Met?Asp?Gly?Gln?Gly?Pro?????1222255?CTG?ATC?CAT?CTG?CAT?TCC?CAG?AAT?ATA?AGA?ATC?TCA?TTT?GGG?GCC?ACC????2302???123??Leu?Ile?His?Leu?His?Ser?Gln?Asn?Ile?Arg?Ile?Ser?Phe?Gly?Ala?Thr?????138??2303??CAG?CCC?CCA?CTA?CTC?TTA?GTA?TAT?GAA?ATA?AGG?TGG?AAT?AGA?TAA?GAG????2350???139??Gln?Pro?Pro?Leu?Leu?Leu?Val?Tyr?Glu?Ile?Arg?Trp?Asn?Arg?***?????????153??2351??GAT?GGA?ACT?GGC?CCC?TGG?GTG?CAT?GAT?GGG?GCC?AAT?AAC?GGA?GGT?ACA????2398??2399??GGG?TGC?ATA?TGT?GGC?TGG?GCT?GTG?AGA?CAG?GGA?AAG?AGG?ATG?GAA?TTG????2446??2447??AGA?TGC?AAG?GTA?GAA?AAG?CAT?GAC?TAT?GTT?CAG?TTT?GGG?ACT?TGA?TGA????2494??2495??AAT?AGA?TAC?TGG?TAG?GAC?AGT?TCA?CTA?GGC?ACT?GGG?CCT?TCA?GGA?GAA????2542??2543??AGA?GAA?GAA?AGG?AAT?GGT?GGC?ATT?GGG?AGT?AGT?AAA?GCT?GTG?GGT?ATG????2590??2591??AAT?GAG?TAT?CTA?TGA?GAC?TCA?CCT?CCC?TGT?GAG?GAC?ATA?GAA?TGA?GGA????2638??2639??GAC?ACA?GTC?TAC?AGG?TAG?ACT?AGG?TCG?AGT?TCA?GAA?GCC?TGA?AGA?ACA????2686??2687??CCA?GTG?TTT?AAG?GGA?TGG?TGT?TGG?GAA?AGG?GAG?CCA?GGC?CAG?CCA?GAG????2734??2735??AGG?AAT?GTT?CCG?GAA?CTC?TGA?AAG?GAA?GGA?AAT?GGC?AGG?AAC?AAA?GGA????2782??2783??GCT?GGG?GCA?CAA?CAG?TGG?TGA?CTC?ACA?CTG?GGA?ACA?CTT?CGG?CCC?ATT????2830??2831??GTG?TTT?ATG?CTT?ATA?GTT?TCC?CAA?GTT?TAT?CTT?TTC?AGG?GTT?ATG?ATT????2878??2879??ACG?TTA?ACC?TCA?CCT?CCC?TCC?CTC?CCT?CTA?AAA?CAA?AAC?AAA?AAA?AAA????2926??2927??AAA?AAA?AAA????????????????????????????????????????????????????????2935
5.?PP13479
A: nucleotide sequence, (SEQ, ID, NO:13) length: 2323, 1, GGCCGGATTC, CCAGTGGTGG, CGAGGAGGTG, GTATTTTTTT, AAGTGTCAGT, GTGGCATTGT, 61, GGTTGCCTCA, ATAAAACAGA, GTCTTTTTTT, GTCTTTTTTT, TTTGAGACAG, GGTCTCGCTC, 121, TCACCCAGGC, TGGAGTGCAG, TGACACAGTT, GCAGCTCACT, GCAGCCTCGA, CCTCCTGGGC, 181, TCAAGCAATC, CTCCCGCCTC, AGCCTCCCGA, GTAGCTGAGT, CTACAGATCT, CCACAATGGA, 241, AGTTCGAAGC, AAGCAAAAGC, CACGCAAACC, ACAGGCCGAT, CTGTCTGAGC, CCTAGGATTT, 301, GGCCCGGTTC, TGCTTCAGCC, ACCAGCACCG, TCTGCTCCTC, CTCAGAATCC, TTCCTCCCCC, 361, GTGGCCCGCC, CGCCGTGTCC, CTCCTCCTCC, ACGGGCCGCC, CACCGTGTCC, TTCCCTCCCC, 421, CGTGGCCCAC, CCACCATGTC, CTTCCCTCCC, CTGTGGCCCA, CCCGCCATGT, CCCTGCCTCC, 481, CACCCGACAT, GCCCCTTGAA, GCTGCCTGGG, CCCTGCTGTT, GTCCCCACTG, CCTGTGTGAC, 541, TCTGCGCCCC, CTTCCCTACC, CTGCCCCACC, CTGGTTCAGG, GAGCGTCCAG, GCCCATTCTC, 601, ATCCTCAGGG, CCTTCCCTGG, CCCTTGCCAC, TCTGTGCCGT, GTCATGACCT, GAAGCTGCAG, 661, GTGGGCGCCT, CCCCCTTCCG, TCATGGCTGT, CCCCCTTCTG, TGAGGTGTCC, CAGCCGCCTG, 721, ATTGCCGGAG, TCCCAGGGTG, CTCGGTGCTG, TCGTGGAGCC, TGGGACATTC, ACTGTCTGGG, 781, ATTGATTCCA, GGGTTGGAGC, CACACCTGGT, CTGGGGCATT, CGCTGTCCTG, GGTCAGAGCC, 841, CCTCCTGGTC, TGGGACATTC, GCTGTCTGGG, GTTGGAGCCA, CACCTGGTCT, GGGGCATTTG, 901, CTGTCCGGGG, TCGGAGCCTC, ACCTGGTGAA, GATACAGAAC, ATGCTGCTGC, CCTAACCCCG, 961, TGTGGTGTGC, CCCCTGTCCC, CGGGTGTCGT, TCCCATAGCC, AGCCCTTGTC, TCATCTCGTC1021, TCATCCTCTA, GATGCTGTGG, GCCCTGAGGG, AAAGGATCAC, AGAGGCGCTG, AGCCGGGATG1081, GCTACGTGTA, CAAGTACGAC, CTCTCCCTCC, CTGTGGAGCG, GCTCTACGAC, ATCGTGACTG1141, ACCTGCGCGC, CCGCCTCGGC, CCGCACGCCA, AGCACGTGGT, GGGCTATGGC, CACCTTGGAG1201, ATGGTAACCT, GCACCTCAAT, GTGACGGCGG, AGGCCTTCAG, CCCCTCGCTC, CTGGCTGCCC1261, TGGAGCCCCA, CGTGTACGAG, TGGACGGCCG, GGCAGCAGGG, CAGCGTCAGC, GCGGAGCACG1321, GAGTGGGCTT, CAGGAAGAGG, GACGTCCTGG, GCTACAGCAA, GCCACCGGGG, GCCCTGCAGC1381, TCATGCAGCA, GCTCAAGGCC, CTGCTGGACC, CCAAGGGCAT, CCTCAACCCC, TACAAGACGC1441, TGCCCAGCCA, GGCCTGACGG, CCACTCCTGC, TGCTGCCAAG, GCCCACTGGG, GGTCGGCGGG1501, TGGCTCTCGG, GCGGGGGTGT, TGCGGTGGCT, CTGAGGGATG, AGCCGGCAGT, GGGCAGGGGA1561, CCAGGCACCT, GGTTGAAGGG, ACTGGGAGCC, CGCACTGGGG, AACTGCCGGA, CGCAGGCCCT1621, CGGGCAGGAG, CATCTGGCAG, AGTGGGGGGC, GTGGCAGGCA, CCCTCCTTTG, CAGGGCGAGG1681, TGGGGCCTCT, GCAGCCATCC, TGGACAGGCC, GGGGTGGCGG, CAGCTTTGCC, CACGTGGAAG1741, CGGGGTGGGT, CTCACTTGCG, TGGTGGCCCC, TGGCCCCATC, TTGCCTGCTG, CGGCCTGGGG1801, AGCAGGCGCT, GGGTGGTGGT, TCTGCCTGCT, TGCTGCTCGT, TCCCCGGGCA, TGCGTGGGCA1861, GCGGGGGGCA, TGCGTGGGCA, GCAGGGGGCG, TGGGCAGCGG, GGGCATGGGC, AGGACCACGT1921, GGGCCGTGAT, CGTGGGTTGC, CGAGAGGACC, TGAGCGCTGC, GGCTCTGCTG, AATGGAGCCG1981, GGTCCCTCAG, GCCGTGGACG, CCCTCGGGAG, GGGGGTGACT, GTGGCTTGTG, TCTGGACAGG2041, AATGTGTCAT, TTCCCACATC, TTCTAGAGGG, CTGCCAGCTG, GGAAGACAGT, TATCAGGGCA2101, AGCTGTGCTC, TGAGTTTCGG, GTTCTGCTCC, TACAAAGAAC, GTGCGGTGCT, GCGGGCGAGG2161, GCCCCGGCAC, GGACAAGGGC, CACTGCAGAG, TGTGTTTCTG, CTCGTCAGCT, GCCCTGGGCA, 2221, GCGGATGGGC, TGGGCGATGC, AGCTGGATGC, ACATCTCATT, CTGTCATGAA, TGTCCAGTAA, 2281, AAATCTGAAT, TGGTTGCAAA, AAAAAAAAAA, AAAAAAAAAA, AAA
B: nucleotide sequence, (SEQ, ID, NO:14), length: 203, 1, MSFPPLWPTR, HVPASHPTCP, LKLPGPCCCP, HCLCDSAPPS, LPCPTLVQGA, SRPILILRAF, 61, PGPCHSVPCH, DLKLQVGASP, FRHGCPPSVR, CPSRLIAGVP, GCSVLSWSLG, HSLSGIDSRV, 121, GATPGLGHSL, SWVRAPPGLG, HSLSGVGATP, GLGHLLSGVG, ASPGEDTEHA, AALTPCGVPP, 181, VPGCRSHSQP, LSHLVSSSRC, CGP, C. nucleotides and amino acid composite sequence, (SEQ, ID, NO:15), clone number: PP13479, start code: 436, ATG, stop coding: 1045, TGA, protein molecular weight: 17771.81, 1, GGC, CGG, ATT, CCC, AGT, GGT, GGC, GAG, GAG, GTG, GTA, TTT, TTT, TAA, GTG, TCA, 48, 49, GTG, TGG, CAT, TGT, GGT, TGC, CTC, AAT, AAA, ACA, GAG, TCT, TTT, TTT, GTC, TTT, 96, 97, TTT, TTT, TGA, GAC, AGG, GTC, TCG, CTC, TCA, CCC, AGG, CTG, GAG, TGC, AGT, GAC, 144, 145, ACA, GTT, GCA, GCT, CAC, TGC, AGC, CTC, GAC, CTC, CTG, GGC, TCA, AGC, AAT, CCT, 192, 193, CCC, GCC, TCA, GCC, TCC, CGA, GTA, GCT, GAG, TCT, ACA, GAT, CTC, CAC, AAT, GGA, 240, 241, AGT, TCG, AAG, CAA, GCA, AAA, GCC, ACG, CAA, ACC, ACA, GGC, CGA, TCT, GTC, TGA, 288, 289, GCC, CTA, GGA, TTT, GGC, CCG, GTT, CTG, CTT, CAG, CCA, CCA, GCA, CCG, TCT, GCT, 336, 337, CCT, CCT, CAG, AAT, CCT, TCC, TCC, CCC, GTG, GCC, CGC, CCG, CCG, TGT, CCC, TCC, 384, 385, TCC, TCC, ACG, GGC, CGC, CCA, CCG, TGT, CCT, TCC, CTC, CCC, CGT, GGC, CCA, CCC, 432, 433, ACC, ATG, TCC, TTC, CCT, CCC, CTG, TGG, CCC, ACC, CGC, CAT, GTC, CCT, GCC, TCC, 480, 1, Met, Ser, Phe, Pro, Pro, Leu, Trp, Pro, Thr, Arg, His, Val, Pro, Ala, Ser, 15, 481, CAC, CCG, ACA, TGC, CCC, TTG, AAG, CTG, CCT, GGG, CCC, TGC, TGT, TGT, CCC, CAC, 528, 16, His, Pro, Thr, Cys, Pro, Leu, Lys, Leu, Pro, Gly, Pro, Cys, Cys, Cys, Pro, His, 31, 529, TGC, CTG, TGT, GAC, TCT, GCG, CCC, CCT, TCC, CTA, CCC, TGC, CCC, ACC, CTG, GTT, 576, 32, Cys, Leu, Cys, Asp, Ser, Ala, Pro, Pro, Ser, Leu, Pro, Cys, Pro, Thr, Leu, Val, 47, 577, CAG, GGA, GCG, TCC, AGG, CCC, ATT, CTC, ATC, CTC, AGG, GCC, TTC, CCT, GGC, CCT, 624, 48, Gln, Gly, Ala, Ser, Arg, Pro, Ile, Leu, Ile, Leu, Arg, Ala, Phe, Pro, Gly, Pro, 63, 625, TGC, CAC, TCT, GTG, CCG, TGT, CAT, GAC, CTG, AAG, CTG, CAG, GTG, GGC, GCC, TCC, 672, 64, Cys, His, Ser, Val, Pro, Cys, His, Asp, Leu, Lys, Leu, Gln, Val, Gly, Ala, Ser, 79, 673, CCC, TTC, CGT, CAT, GGC, TGT, CCC, CCT, TCT, GTG, AGG, TGT, CCC, AGC, CGC, CTG, 720, 80, Pro, Phe, Arg, His, Gly, Cys, Pro, Pro, Ser, Val, Arg, Cys, Pro, Ser, Arg, Leu, 95, 721, ATT, GCC, GGA, GTC, CCA, GGG, TGC, TCG, GTG, CTG, TCG, TGG, AGC, CTG, GGA, CAT, 768, 96, Ile, Ala, Gly, Val, Pro, Gly, Cys, Ser, Val, Leu, Ser, Trp, Ser, Leu, Gly, His, 111, 769, TCA, CTG, TCT, GGG, ATT, GAT, TCC, AGG, GTT, GGA, GCC, ACA, CCT, GGT, CTG, GGG, 816, 112, Ser, Leu, Ser, Gly, Ile, Asp, Ser, Arg, Val, Gly, Ala, Thr, Pro, Gly, Leu, Gly, 127, 817, CAT, TCG, CTG, TCC, TGG, GTC, AGA, GCC, CCT, CCT, GGT, CTG, GGA, CAT, TCG, CTG, 864, 128, His, Ser, Leu, Ser, Trp, Val, Arg, Ala, Pro, Pro, Gly, Leu, Gly, His, Ser, Leu, 143, 865, TCT, GGG, GTT, GGA, GCC, ACA, CCT, GGT, CTG, GGG, CAT, TTG, CTG, TCC, GGG, GTC, 912, 144, Ser, Gly, Val, Gly, Ala, Thr, Pro, Gly, Leu, Gly, His, Leu, Leu, Ser, Gly, Val, 159, 913, GGA, GCC, TCA, CCT, GGT, GAA, GAT, ACA, GAA, CAT, GCT, GCT, GCC, CTA, ACC, CCG, 960, 160, Gly, Ala, Ser, Pro, Gly, Glu, Asp, Thr, Glu, His, Ala, Ala, Ala, Leu, Thr, Pro, 175, 961, TGT, GGT, GTG, CCC, CCT, GTC, CCC, GGG, TGT, CGT, TCC, CAT, AGC, CAG, CCC, TTG, 1008, 176, Cys, Gly, Val, Pro, Pro, Val, Pro, Gly, Cys, Arg, Ser, His, Ser, Gln, Pro, Leu, 1911009, TCT, CAT, CTC, GTC, TCA, TCC, TCT, AGA, TGC, TGT, GGG, CCC, TGA, GGG, AAA, GGA, 1056, 192, Ser, His, Leu, Val, Ser, Ser, Ser, Arg, Cys, Cys, Gly, Pro, * *, 204, 1057, TCA, CAG, AGG, CGC, TGA, GCC, GGG, ATG, GCT, ACG, TGT, ACA, AGT, ACG, ACC, TCT, 1104, 1105, CCC, TCC, CTG, TGG, AGC, GGC, TCT, ACG, ACA, TCG, TGA, CTG, ACC, TGC, GCG, CCC, 1152, 1153, GCC, TCG, GCC, CGC, ACG, CCA, AGC, ACG, TGG, TGG, GCT, ATG, GCC, ACC, TTG, GAG, 1200, 1201, ATG, GTA, ACC, TGC, ACC, TCA, ATG, TGA, CGG, CGG, AGG, CCT, TCA, GCC, CCT, CGC, 1248, 1249, TCC, TGG, CTG, CCC, TGG, AGC, CCC, ACG, TGT, ACG, AGT, GGA, CGG, CCG, GGC, AGC, 1296, 1297, AGG, GCA, GCG, TCA, GCG, CGG, AGC, ACG, GAG, TGG, GCT, TCA, GGA, AGA, GGG, ACG, 1344, 1345, TCC, TGG, GCT, ACA, GCA, AGC, CAC, CGG, GGG, CCC, TGC, AGC, TCA, TGC, AGC, AGC, 1392, 1393, TCA, AGG, CCC, TGC, TGG, ACC, CCA, AGG, GCA, TCC, TCA, ACC, CCT, ACA, AGA, CGC, 1440, 1441, TGC, CCA, GCC, AGG, CCT, GAC, GGC, CAC, TCC, TGC, TGC, TGC, CAA, GGC, CCA, CTG, 1488, 1489, GGG, GTC, GGC, GGG, TGG, CTC, TCG, GGC, GGG, GGT, GTT, GCG, GTG, GCT, CTG, AGG, 1536, 1537, GAT, GAG, CCG, GCA, GTG, GGC, AGG, GGA, CCA, GGC, ACC, TGG, TTG, AAG, GGA, CTG, 1584, 1585, GGA, GCC, CGC, ACT, GGG, GAA, CTG, CCG, GAC, GCA, GGC, CCT, CGG, GCA, GGA, GCA, 1632, 1633, TCT, GGC, AGA, GTG, GGG, GGC, GTG, GCA, GGC, ACC, CTC, CTT, TGC, AGG, GCG, AGG, 1680, 1681, TGG, GGC, CTC, TGC, AGC, CAT, CCT, GGA, CAG, GCC, GGG, GTG, GCG, GCA, GCT, TTG, 1728, 1729, CCC, ACG, TGG, AAG, CGG, GGT, GGG, TCT, CAC, TTG, CGT, GGT, GGC, CCC, TGG, CCC, 1776, 1777, CAT, CTT, GCC, TGC, TGC, GGC, CTG, GGG, AGC, AGG, CGC, TGG, GTG, GTG, GTT, CTG, 1824, 1825, CCT, GCT, TGC, TGC, TCG, TTC, CCC, GGG, CAT, GCG, TGG, GCA, GCG, GGG, GGC, ATG, 1872, 1873, CGT, GGG, CAG, CAG, GGG, GCG, TGG, GCA, GCG, GGG, GCA, TGG, GCA, GGA, CCA, CGT, 1920, 1921, GGG, CCG, TGA, TCG, TGG, GTT, GCC, GAG, AGG, ACC, TGA, GCG, CTG, CGG, CTC, TGC, 1968, 1969, TGA, ATG, GAG, CCG, GGT, CCC, TCA, GGC, CGT, GGA, CGC, CCT, CGG, GAG, GGG, GGT, 2016, 2017, GAC, TGT, GGC, TTG, TGT, CTG, GAC, AGG, AAT, GTG, TCA, TTT, CCC, ACA, TCT, TCT, 2064, 2065, AGA, GGG, CTG, CCA, GCT, GGG, AAG, ACA, GTT, ATC, AGG, GCA, AGC, TGT, GCT, CTG, 2112, 2113, AGT, TTC, GGG, TTC, TGC, TCC, TAC, AAA, GAA, CGT, GCG, GTG, CTG, CGG, GCG, AGG, 2160, 2161, GCC, CCG, GCA, CGG, ACA, AGG, GCC, ACT, GCA, GAG, TGT, GTT, TCT, GCT, CGT, CAG, 2208, 2209, CTG, CCC, TGG, GCA, GCG, GAT, GGG, CTG, GGC, GAT, GCA, GCT, GGA, TGC, ACA, TCT, 2256, 2257, CAT, TCT, GTC, ATG, AAT, GTC, CAG, TAA, AAA, TCT, GAA, TTG, GTT, GCA, AAA, AAA, 2304, 2305, AAA, AAA, AAA, AAA, AAA, AAA, A, 2323
6.?PP13842
A: nucleotide sequence, (SEQ, ID, NO:16) length: 1259, 1, GCTGCAGCCC, CTGCCCCCGC, CCCTCCTCGC, TGGGTGCTCA, GAAGGCTGAC, AGCTGCGCCA, 61, GGCTGAGGCG, GCAGTCGATG, CTGGAGTTGT, CCGGGCCCGT, GTAGGCCAGG, CCCAGGGGCT, 121, CTAGGAAGGC, CCGGCAGGCC, CCAGCGCTGC, CTTTGCGGAT, TCTGTTTTTG, AGCCGTGGAC, 181, TTGGGTTGTA, AATTTATTTG, TGGGGAGTGC, GCTCCAGGAA, GAGCCACCAT, CCCTGCCCCC, 241, GTTTTCCCAC, CGGGGAGTCT, GTACAGAGAT, TTTTCTACGT, TTTTATTTTT, TGCCTCAGAG, 301, GGATGGGATT, GGGGAGGAGG, GGATGGGCAG, CGGAGGGTTG, GGGGCATGGT, CTGCAGGCTC, 361, ATCTGTGTCC, GCTTTCACTC, CACTAATGCT, GTCTCAGTGT, TTTCTCTCTC, TCTCTTTCGA, 421, GCTTGCACTC, CGGTACCCGA, CCCGGCGCCC, TGGCCCATCC, CATGCCGGGG, GGCCAGTGGA, 481, AAGAAGACAG, GCCGTCCAGC, CCGTGCCCGC, CTGCGGCGGG, GGCACCCAGC, AAGCCCGCCC, 541, ACCGCCCGCT, GCCTCACCTG, CTTCGCCACA, GACTCTTGTT, CCCAGCCCCT, TGGGGCCTCC, 601, GTGTTTGGGG, TGGGGGAGCT, GCTTAGAGAC, TGTGCCCGTC, CTCGGCCCCC, CACCCTGAAG, 661, TGCCAGCACC, ACCAGCACCA, GATCCTCCGC, CGCCACACCG, CACTGAGGAC, ACGCCGGCCG, 721, GGCCGCCTCG, TCTCAAGTTG, TATAAAGTTG, TCTCCGTGTC, CCCTCCTCCC, TCTGCCCCCA, 781, GTGTTTCTTC, TGATTTTTTT, TTCCCCTTTC, CCTCCCTCCC, CCTCCGCATT, CTTCCCTTGG, 841, TTCAGCACAG, GTAAAACGGT, TCCCCTCCCT, CCCTGCCTTC, ATGGATCACC, AGCTCACGTC, 901, ATGTTGCCTT, CTCTTTTCTT, TGTGTGTGTG, TTTATTTAAG, TTATTTTTCT, TCCTCCTCTC, 961, CCTTTTCTTT, TTGGCCCTCC, CTCCCTCCCT, CTTCTGCCAT, GTAACTGGAG, GATGTGCTAT1021, GAGTTTGCAA, ACAGCTGGAC, TGTCAGGCTG, CTTTTTTTCC, AGATGTTCCT, CCTCTGCCTC1081, CCCTTCCCCT, CCTCTCCCCT, CCTTTTCCTT, CCTTCCTTCC, TTTCCTTGGA, GCACTGAGCA1141, CCATTTGGAA, GCTTGAGAGA, AACCAAAATT, AAAGAGAGAA, AGAGAGAAAA, AAAAAAAAAA1201, AAAAAAAAAA, AAAAAAAAAA, AAAAAAAAAA, AAAAAAAAAA, AAAAAAAAAA, AACCTCGGG, B: nucleotide sequence, (SEQ, ID, NO:17), length: 197, 1, MVCRLICVRF, HSTNAVSVFS, LSLFRACTPV, PDPAPWPIPC, RGASGKKTGR, PARARLRRGH, 61, PASPPTARCL, TCFATDSCSQ, PLGASVFGVG, ELLRDCARPR, PPTLKCQHHQ, HQILRRHTAL121, RTRRPGRLVS, SCIKLSPCPL, LPLPPVFLLI, FFSPFPPSPS, AFFPWFSTGK, TVPLPPCLHG181, SPAHVMLPSL, FFVCVFI
C. Nucleotide and amino acid composite sequence (SEQ ID NO:18) clone number: PP13842
start code: 346ATG, stop coding: 937, TAA, protein molecular weight: 21622.56, 1, GCT, GCA, GCC, CCT, GCC, CCC, GCC, CCT, CCT, CGC, TGG, GTG, CTC, AGA, AGG, CTG, 48, 49, ACA, GCT, GCG, CCA, GGC, TGA, GGC, GGC, AGT, CGA, TGC, TGG, AGT, TGT, CCG, GGC, 96, 97, CCG, TGT, AGG, CCA, GGC, CCA, GGG, GCT, CTA, GGA, AGG, CCC, GGC, AGG, CCC, CAG, 144, 145, CGC, TGC, CTT, TGC, GGA, TTC, TGT, TTT, TGA, GCC, GTG, GAC, TTG, GGT, TGT, AAA, 192, 193, TTT, ATT, TGT, GGG, GAG, TGC, GCT, CCA, GGA, AGA, GCC, ACC, ATC, CCT, GCC, CCC, 240, 241, GTT, TTC, CCA, CCG, GGG, AGT, CTG, TAC, AGA, GAT, TTT, TCT, ACG, TTT, TTA, TTT, 288, 289, TTT, GCC, TCA, GAG, GGA, TGG, GAT, TGG, GGA, GGA, GGG, GAT, GGG, CAG, CGG, AGG, 336, 337, GTT, GGG, GGC, ATG, GTC, TGC, AGG, CTC, ATC, TGT, GTC, CGC, TTT, CAC, TCC, ACT, 384, 1, Met, Val, Cys, Arg, Leu, Ile, Cys, Val, Arg, Phe, His, Ser, Thr, 13, 385, AAT, GCT, GTC, TCA, GTG, TTT, TCT, CTC, TCT, CTC, TTT, CGA, GCT, TGC, ACT, CCG, 432, 14, Asn, Ala, Val, Ser, Val, Phe, Ser, Leu, Ser, Leu, Phe, Arg, Ala, Cys, Thr, Pro, 29, 433, GTA, CCC, GAC, CCG, GCG, CCC, TGG, CCC, ATC, CCA, TGC, CGG, GGG, GCC, AGT, GGA, 480, 30, Val, Pro, Asp, Pro, Ala, Pro, Trp, Pro, Ile, Pro, Cys, Arg, Gly, Ala, Ser, Gly, 45, 481, AAG, AAG, ACA, GGC, CGT, CCA, GCC, CGT, GCC, CGC, CTG, CGG, CGG, GGG, CAC, CCA, 528, 46, Lys, Lys, Thr, Gly, Arg, Pro, Ala, Arg, Ala, Arg, Leu, Arg, Arg, Gly, His, Pro, 61, 529, GCA, AGC, CCG, CCC, ACC, GCC, CGC, TGC, CTC, ACC, TGC, TTC, GCC, ACA, GAC, TCT, 576, 62, Ala, Ser, Pro, Pro, Thr, Ala, Arg, Cys, Leu, Thr, Cys, Phe, Ala, Thr, Asp, Ser, 77, 577, TGT, TCC, CAG, CCC, CTT, GGG, GCC, TCC, GTG, TTT, GGG, GTG, GGG, GAG, CTG, CTT, 624, 78, Cys, Ser, Gln, Pro, Leu, Gly, Ala, Ser, Val, Phe, Gly, Val, Gly, Glu, Leu, Leu, 93, 625, AGA, GAC, TGT, GCC, CGT, CCT, CGG, CCC, CCC, ACC, CTG, AAG, TGC, CAG, CAC, CAC, 672, 94, Arg, Asp, Cys, Ala, Arg, Pro, Arg, Pro, Pro, Thr, Leu, Lys, Cys, Gln, His, His, 109, 673, CAG, CAC, CAG, ATC, CTC, CGC, CGC, CAC, ACC, GCA, CTG, AGG, ACA, CGC, CGG, CCG, 720, 110, Gln, His, Gln, Ile, Leu, Arg, Arg, His, Thr, Ala, Leu, Arg, Thr, Arg, Arg, Pro, 125, 721, GGC, CGC, CTC, GTC, TCA, AGT, TGT, ATA, AAG, TTG, TCT, CCG, TGT, CCC, CTC, CTC, 768, 126, Gly, Arg, Leu, Val, Ser, Ser, Cys, Ile, Lys, Leu, Ser, Pro, Cys, Pro, Leu, Leu, 141, 769, CCT, CTG, CCC, CCA, GTG, TTT, CTT, CTG, ATT, TTT, TTT, TCC, CCT, TTC, CCT, CCC, 816, 142, Pro, Leu, Pro, Pro, Val, Phe, Leu, Leu, Ile, Phe, Phe, Ser, Pro, Phe, Pro, Pro, 157, 817, TCC, CCC, TCC, GCA, TTC, TTC, CCT, TGG, TTC, AGC, ACA, GGT, AAA, ACG, GTT, CCC, 864, 158, Ser, Pro, Ser, Ala, Phe, Phe, Pro, Trp, Phe, Ser, Thr, Gly, Lys, Thr, Val, Pro, 173, 865, CTC, CCT, CCC, TGC, CTT, CAT, GGA, TCA, CCA, GCT, CAC, GTC, ATG, TTG, CCT, TCT, 912, 174, Leu, Pro, Pro, Cys, Leu, His, Gly, Ser, Pro, Ala, His, Val, Met, Leu, Pro, Ser, 189, 913, CTT, TTC, TTT, GTG, TGT, GTG, TTT, ATT, TAA, GTT, ATT, TTT, CTT, CCT, CCT, CTC, 960, 190, Leu, Phe, Phe, Val, Cys, Val, Phe, Ile, * *, 198, 961, CCT, TTT, CTT, TTT, GGC, CCT, CCC, TCC, CTC, CCT, CTT, CTG, CCA, TGT, AAC, TGG, 10081009, AGG, ATG, TGC, TAT, GAG, TTT, GCA, AAC, AGC, TGG, ACT, GTC, AGG, CTG, CTT, TTT, 10561057, TTC, CAG, ATG, TTC, CTC, CTC, TGC, CTC, CCC, TTC, CCC, TCC, TCT, CCC, CTC, CTT, 11041105, TTC, CTT, CCT, TCC, TTC, CTT, TCC, TTG, GAG, CAC, TGA, GCA, CCA, TTT, GGA, AGC, 11521153, TTG, AGA, GAA, ACC, AAA, ATT, AAA, GAG, AGA, AAG, AGA, GAA, AAA, AAA, AAA, AAA, 12001201, AAA, AAA, AAA, AAA, AAA, AAA, AAA, AAA, AAA, AAA, AAA, AAA, AAA, AAA, AAA, AAA, 12481249, AAA, ACC, TCG, GG, 1259
7.PP14673
A: the nucleotide sequence (SEQ ID NO: 19) Length: 2333
1 GCGGCCGTAG CGGCCGGGGC TGCGGTAGCC ACTTTAGATT TGGGCAAGGA CTTTAGATTC
61 GGGCTCTGTT CTGTTTCCGC CGTCCTGCTT CCTGCCGAGG CTGGCCCAGG CAGCCGCGCT
121 TCGAAGGACG CCGCCGGGAG CTGCGGAGCA TGCGTGGAGT GGCAGTGCTA ACGGCTGGTG
181 TCTCGCACTG TTGGCCTGTG AAGGTACGTG AAGCTGAAAG CCTGGAATGG CTGGAAAGGG
241 GTCATCAGGC AGGCGGCCCC TGCTGCTGGG GCTGCTGGTG GCCGTAGCCA CTGTCCACCT
301 GGTCATCTGT CCCTACACCA AAGTGGAGGA GAGCTTCAAC CTGCAGGCCA CACATGACCT
361 GCTCTACCAC TGGCAAGACC TGGAGCAGTA CGACCATCTT GAGTTCCCCG GAGTCGTCCC
421 CAGGACGTTC CTCGGGCCAG TGGTGATCGC AGTGTTCTCC AGCCCCGCGG TTTACGTGCT
481 TTCGCTGTTA GAAATGTCCA AGTTTTACTC TCAGCTAATA GTTAGAGGAG TGCTTGGACT
541 CGGCGTGATT TTTGGACTCT GGACGTTACA AAAGGAAGTG AGACGGCACT TCGGGGCCAT
601 GGTGGCCACC ATGTTCTGCT GGGTGACGGC CATGCAGTTC CACCTGATGT TCTACTGCAC
661 GCGGACACTG CCCAATGTGC TGGCCCTGCC TGTAGTCCTG CTGGCCCTCG CGGCCTGGCT
721 GCGGCACGAG TGGGCCCGCT TCATCTGGCT GTCAGCCTTC GCCATCATCG TGTTCAGGGT
781 GGAGCTGTGC CTGTTCCTGG GCCTCCTGCT GCTGCTGGCC TTGGGCAACC GAAAGGTTTC
841 TGTAGTCAGA GCCCTTCGCC ACGCCGTCCC GGCAGGGATC CTCTGTTTAG GACTGACGGT
901 TGCTGTGGAC TCTTATTTTT GGCGGCAGCT CACTTGGCCG GAAGGAAAGG TGCTTTGGTA
961 CAACACTGTC CTGAACAAAA GCTCCAACTG GGGGACCTCC CCGCTGCTGT GGTACTTCTA
1021 CTCAGCCCTG CCCCGCGGCC TGGGCTGCAG CCTGCTCTTC ATCCCCCTGG GCTTGGTAGA
1081 CAGAAGGACG CACGCGCCGA CGGTGCTGGC ACTGGGCTTC ATGGCACTCT ACTCCCTCCT
1141 GCCACACAAG GAGCTACGCT TCATCATCTA TGCCTTCCCC ATGCTCAACA TCACGGCTGC
1201 CAGAGGCTGC TCCTACCTGC TGAATAACTA TAAAAAGTCT TGGCTGTACA AAGCGGGGTC
1261 TCTGCTTGTG ATCGGACACC TCGTGGTGAA TGCCGCCTAC TCAGCCACGG CCCTGTATGT
1321 GTCCCATTTC AACTACCCAG GTGGCGTCGC AATGCAGAGG CTGCACCAGC TGGTGCCCCC
1381 CCAGACAGAC GTCCTTCTGC ACATTGACGT GGCAGCCGCC CAGACAGGTG TGTCTCGGTT
1441 TCTCCAAGTC AACAGCGCCT GGAGGTACGA CAAGAGGGAG GATGTGCAGC CGGGGACAGG
1501 CATGCTGGCA TACACACACA TCCTCATGGA GGCGGCCCCT GGGCTCCTGG CCCTCTACAG
1561 GGACACACAC CGGGTCCTGG CCAGCGTCGT GGGGACCACA GGTGTGAGTC TGAACCTGAC
1621 CCAACTGCCC CCCTTCAACG TCCACCTGCA GACAAAGCTG GTGCTTCTGG AGAGGCTCCC
1681 CCGGCCGTCC TGAGGGGGAC CAGGCAGCCC TCAGCAGCCA CAGGCCTTCC AGGAGCTGTT
1741 ATCACTACCA GTTTCTGGCA CAATTCCAGC ACAATTATGA CAATTCAGAG AAGCAAGTCA
1801 AAGGACTGGG CACCTGCCTC TGACAGACAC CAGACCAGGT CCAGGGCCTC CTCCACAGCC
1861 TCAGCTGGGG CTCTCAGCAC CAAAGAACGA GGGGCCCAGG TCTTGTTGGC ACCCCGGGAG
1921 CCACTGCCCA GGGTGATTGG TGGCCAGCTC AGGGCTTCCT GCGGGTGACT GTCGCCCAGA
1981 CCAGGTGCCA TTCATGACTA ATCAGGAGCA GCGGGCTCAC CCAGGCACCT GTCTGCCAGG
2041 AGGCCACCGT GTGTCCTGCC CACCCAGGGG GAGCTGTATT TTGGCAGCAC CCCACGCTTG
2101 CTGCCCGAGG GCCTCTTGGG GCACCTAAGA CAGCACCCCC TCTCAGGGGA GACCATGGTG
2161 GCCCCGGCCG CACCCCCCCA CCCTGGTGCC ACCACTGCAA CTTTTGTATT CACAGGCATC
2221 CCATCTCCAT CACAGATAAA ATCTTAGGAG ATAAACACAT TCAAAAAGGA ATGAGATAAA
2281 AAGAATAAGG CAATAAATGT TGATTGGAAC CTCTCAAAAA AAAAAAAAAA AAA
B: Nucleotide sequence (SEQ ID NO: 20) Length: 488
1 MAGKGSSGRR PLLLGLLVAV ATVHLVICPY TKVEESFNLQ ATHDLLYHWQ DLEQYDHLEF
61 PGVVPRTFLG PVVIAVFSSP AVYVLSLLEM SKFYSQLIVR GVLGLGVIFG LWTLQKEVRR
121 HFGAMVATMF CWVTAMQFHL MFYCTRTLPN VLALPVVLLA LAAWLRHEWA RFIWLSAFAI
181 IVFRVELCLF LGLLLLLALG NRKVSVVRAL RHAVPAGILC LGLTVAVDSY FWRQLTWPEG
241 KVLWYNTVLN KSSNWGTSPL LWYFYSALPR GLGCSLLFIP LGLVDRRTHA PTVLALGFMA
301 LYSLLPHKEL RFIIYAFPML NITAARGCSY LLNNYKKSWL YKAGSLLVIG HLVVNAAYSA
361 TALYVSHFNY PGGVAMQRLH QLVPPQTDVL LHIDVAAAQT GVSRFLQVNS AWRYDKREDV
421 QPGTGMLAYT HILMEAAPGL LALYRDTHRV LASVVGTTGV SLNLTQLPPF NVHLQTKLVL
481 LERLPRPS
Nucleotide and amino acid sequence of C. Combination (SEQ ID NO: 21) Clone: PP14673
Start codon: 227 ATG termination codon: 1691 TGA Protein Weight: 54651.70
1 G CGG CCG TAG CGG CCG GGG CTG CGG TAG CCA CTT TAG ATT TGG GCA 46
47 AGG ACT TTA GAT TCG GGC TCT GTT CTG TTT CCG CCG TCC TGC TTC CTG 94
95 CCG AGG CTG GCC CAG GCA GCC GCG CTT CGA AGG ACG CCG CCG GGA GCT 142
143 GCG GAG CAT GCG TGG AGT GGC AGT GCT AAC GGC TGG TGT CTC GCA CTG 190
191 TTG GCC TGT GAA GGT ACG TGA AGC TGA AAG CCT GGA ATG GCT GGA AAG 238
1 Met Ala Gly Lys 4
239 GGG TCA TCA GGC AGG CGG CCC CTG CTG CTG GGG CTG CTG GTG GCC GTA 286
5 Gly Ser Ser Gly Arg Arg Pro Leu Leu Leu Gly Leu Leu Val Ala Val 20
287 GCC ACT GTC CAC CTG GTC ATC TGT CCC TAC ACC AAA GTG GAG GAG AGC 334
21 Ala Thr Val His Leu Val Ile Cys Pro Tyr Thr Lys Val Glu Glu Ser 36
335 TTC AAC CTG CAG GCC ACA CAT GAC CTG CTC TAC CAC TGG CAA GAC CTG 382
37 Phe Asn Leu Gln Ala Thr His Asp Leu Leu Tyr His Trp Gln Asp Leu 52
383 GAG CAG TAC GAC CAT CTT GAG TTC CCC GGA GTC GTC CCC AGG ACG TTC 430
53 Glu Gln Tyr Asp His Leu Glu Phe Pro Gly Val Val Pro Arg Thr Phe 68
431 CTC GGG CCA GTG GTG ATC GCA GTG TTC TCC AGC CCC GCG GTT TAC GTG 478
69 Leu Gly Pro Val Val Ile Ala Val Phe Ser Ser Pro Ala Val Tyr Val 84
479 CTT TCG CTG TTA GAA ATG TCC AAG TTT TAC TCT CAG CTA ATA GTT AGA 526
85 Leu Ser Leu Leu Glu Met Ser Lys Phe Tyr Ser Gln Leu Ile Val Arg 100
527 GGA GTG CTT GGA CTC GGC GTG ATT TTT GGA CTC TGG ACG TTA CAA AAG 574
101 Gly Val Leu Gly Leu Gly Val Ile Phe Gly Leu Trp Thr Leu Gln Lys 116
575 GAA GTG AGA CGG CAC TTC GGG GCC ATG GTG GCC ACC ATG TTC TGC TGG 622
117 Glu Val Arg Arg His Phe Gly Ala Met Val Ala Thr Met Phe Cys Trp 132
623 GTG ACG GCC ATG CAG TTC CAC CTG ATG TTC TAC TGC ACG CGG ACA CTG 670
133 Val Thr Ala Met Gln Phe His Leu Met Phe Tyr Cys Thr Arg Thr Leu 148
671 CCC AAT GTG CTG GCC CTG CCT GTA GTC CTG CTG GCC CTC GCG GCC TGG 718
149 Pro Asn Val Leu Ala Leu Pro Val Val Leu Leu Ala Leu Ala Ala Trp 164
719 CTG CGG CAC GAG TGG GCC CGC TTC ATC TGG CTG TCA GCC TTC GCC ATC 766
165 Leu Arg His Glu Trp Ala Arg Phe Ile Trp Leu Ser Ala Phe Ala Ile 180
767 ATC GTG TTC AGG GTG GAG CTG TGC CTG TTC CTG GGC CTC CTG CTG CTG 814
181 Ile Val Phe Arg Val Glu Leu Cys Leu Phe Leu Gly Leu Leu Leu Leu 196
815 CTG GCC TTG GGC AAC CGA AAG GTT TCT GTA GTC AGA GCC CTT CGC CAC 862
197 Leu Ala Leu Gly Asn Arg Lys Val Ser Val Val Arg Ala Leu Arg His 212
863 GCC GTC CCG GCA GGG ATC CTC TGT TTA GGA CTG ACG GTT GCT GTG GAC 910
213 Ala Val Pro Ala Gly Ile Leu Cys Leu Gly Leu Thr Val Ala Val Asp 228
911 TCT TAT TTT TGG CGG CAG CTC ACT TGG CCG GAA GGA AAG GTG CTT TGG 958
229 Ser Tyr Phe Trp Arg Gln Leu Thr Trp Pro Glu Gly Lys Val Leu Trp 244
959 TAC AAC ACT GTC CTG AAC AAA AGC TCC AAC TGG GGG ACC TCC CCG CTG 1006
245 Tyr Asn Thr Val Leu Asn Lys Ser Ser Asn Trp Gly Thr Ser Pro Leu 260
1007 CTG TGG TAC TTC TAC TCA GCC CTG CCC CGC GGC CTG GGC TGC AGC CTG 1054
261 Leu Trp Tyr Phe Tyr Ser Ala Leu Pro Arg Gly Leu Gly Cys Ser Leu 276
1055 CTC TTC ATC CCC CTG GGC TTG GTA GAC AGA AGG ACG CAC GCG CCG ACG 1102
277 Leu Phe Ile Pro Leu Gly Leu Val Asp Arg Arg Thr His Ala Pro Thr 292
1103 GTG CTG GCA CTG GGC TTC ATG GCA CTC TAC TCC CTC CTG CCA CAC AAG 1150
293 Val Leu Ala Leu Gly Phe Met Ala Leu Tyr Ser Leu Leu Pro His Lys 308
1151 GAG CTA CGC TTC ATC ATC TAT GCC TTC CCC ATG CTC AAC ATC ACG GCT 1198
309 Glu Leu Arg Phe Ile Ile Tyr Ala Phe Pro Met Leu Asn Ile Thr Ala 324
1199 GCC AGA GGC TGC TCC TAC CTG CTG AAT AAC TAT AAA AAG TCT TGG CTG 1246
325 Ala Arg Gly Cys Ser Tyr Leu Leu Asn Asn Tyr Lys Lys Ser Trp Leu 340
1247 TAC AAA GCG GGG TCT CTG CTT GTG ATC GGA CAC CTC GTG GTG AAT GCC 1294
341 Tyr Lys Ala Gly Ser Leu Leu Val Ile Gly His Leu Val Val Asn Ala 356
1295 GCC TAC TCA GCC ACG GCC CTG TAT GTG TCC CAT TTC AAC TAC CCA GGT 1342
357 Ala Tyr Ser Ala Thr Ala Leu Tyr Val Ser His Phe Asn Tyr Pro Gly 372
1343 GGC GTC GCA ATG CAG AGG CTG CAC CAG CTG GTG CCC CCC CAG ACA GAC 1390
373 Gly Val Ala Met Gln Arg Leu His Gln Leu Val Pro Pro Gln Thr Asp 388
1391 GTC CTT CTG CAC ATT GAC GTG GCA GCC GCC CAG ACA GGT GTG TCT CGG 1438
389 Val Leu Leu His Ile Asp Val Ala Ala Ala Gln Thr Gly Val Ser Arg 404
1439 TTT CTC CAA GTC AAC AGC GCC TGG AGG TAC GAC AAG AGG GAG GAT GTG 1486
405 Phe Leu Gln Val Asn Ser Ala Trp Arg Tyr Asp Lys Arg Glu Asp Val 420
1487 CAG CCG GGG ACA GGC ATG CTG GCA TAC ACA CAC ATC CTC ATG GAG GCG 1534
421 Gln Pro Gly Thr Gly Met Leu Ala Tyr Thr His Ile Leu Met Glu Ala 436
1535 GCC CCT GGG CTC CTG GCC CTC TAC AGG GAC ACA CAC CGG GTC CTG GCC 1582
437 Ala Pro Gly Leu Leu Ala Leu Tyr Arg Asp Thr His Arg Val Leu Ala 452
1583 AGC GTC GTG GGG ACC ACA GGT GTG AGT CTG AAC CTG ACC CAA CTG CCC 1630
453 Ser Val Val Gly Thr Thr Gly Val Ser Leu Asn Leu Thr Gln Leu Pro 468
1631 CCC TTC AAC GTC CAC CTG CAG ACA AAG CTG GTG CTT CTG GAG AGG CTC 1678
469 Pro Phe Asn Val His Leu Gln Thr Lys Leu Val Leu Leu Glu Arg Leu 484
1679 CCC CGG CCG TCC TGA GGG GGA CCA GGC AGC CCT CAG CAG CCA CAG GCC 1726
485 Pro Arg Pro Ser *** 489
1727 TTC CAG GAG CTG TTA TCA CTA CCA GTT TCT GGC ACA ATT CCA GCA CAA 1774
1775 TTA TGA CAA TTC AGA GAA GCA AGT CAA AGG ACT GGG CAC CTG CCT CTG 1822
1823 ACA GAC ACC AGA CCA GGT CCA GGG CCT CCT CCA CAG CCT CAG CTG GGG 1870
1871 CTC TCA GCA CCA AAG AAC GAG GGG CCC AGG TCT TGT TGG CAC CCC GGG 1918
1919 AGC CAC TGC CCA GGG TGA TTG GTG GCC AGC TCA GGG CTT CCT GCG GGT 1966
1967 GAC TGT CGC CCA GAC CAG GTG CCA TTC ATG ACT AAT CAG GAG CAG CGG 2014
2015 GCT CAC CCA GGC ACC TGT CTG CCA GGA GGC CAC CGT GTG TCC TGC CCA 2062
2063 CCC AGG GGG AGC TGT ATT TTG GCA GCA CCC CAC GCT TGC TGC CCG AGG 2110
2111 GCC TCT TGG GGC ACC TAA GAC AGC ACC CCC TCT CAG GGG AGA CCA TGG 2158
2159 TGG CCC CGG CCG CAC CCC CCC ACC CTG GTG CCA CCA CTG CAA CTT TTG 2206
2207 TAT TCA CAG GCA TCC CAT CTC CAT CAC AGA TAA AAT CTT AGG AGA TAA 2254
2255 ACA CAT TCA AAA AGG AAT GAG ATA AAA AGA ATA AGG CAA TAA ATG TTG 2302
2303 ATT GGA ACC TCT CAA AAA AAA AAA AAA AAA A 2333
8.PP14776
A: the nucleotide sequence (SEQ ID NO: 22) Length: 1405
1 GTTTGGGTAG ATTTATAGGT GTTTAATAGC CATAGTCAAA GCACATTTAT TAATGATAAA
61 TGGAAATGGA GACCCCTAAG GTATGCAGTA AAGACAGGTC CTCGGTTTGG GATTCGCCAC
121 CGTTTCCTCC CACGCATTTT AATGAAAGAT TTAAATGAAA ACACAGATTT GCTTTCCACA
181 TTTTAAAAAT TATTCAAAAC TAGAGTCAAA GGGGAGAATC TGATACACTG GATAACAGAA
241 TTGAAATTCA ATAAGGCATT CAGAAGCAAA AGCAGTTATG CTGTAACAAG TCAGCTGCAC
301 TGCCATCCCA CTGAAACTAT GATCGATACC TGGCATTTAT GAGAACTTAT GATACACCAG
361 GAACTTCGAT GTGCATCATC CCAGATGATC CTCACAGAAA TCCTAGGAGG TTCTACGATG
421 AAATCAGGAC CCCGCTCCTC TCTGACATCC GCATCGATTA TCCCCCCAGC TCAGTGGTGC
481 AGGCCACCAA GACCCTGTTC CCCAACTACT TCAACGGCTC GGAGATCATC ATTGCGGGGA
541 AGCTGGTGGA CAGGAAGCTG GATCACCTGC ACGTGGAGGT CACCGCCAGC AACAGTAAGA
601 AATTCATCAT CCTGAAGACA GATGTGCCTG TGCGGCCTCA GAAGGCAGGG AAAGATGTCA
661 CAGGAAGCCC CAGGCCTGGA GGCGATGGAG AGGGGGACCC CAACCACATC GAGCGTCTCT
721 GGAGCTACCT CACCACAAAG GAGCTGCTGA GCTCCTGGCT GCAAAGTGAC GATGAACCGG
781 AGAAGGAGCG GCTGCGGCAG CGGGCCCAGG CCCTGGCTGT GAGCTACCGC TTCCTCACTC
841 CCTTCACCTC CATGAAGCTG AGGGGGCCGG TCCCACGCAT GGACGGCCTG GAGGAGGCCC
901 ACGGCATGTC GGCTGCCATG GGACCCGAAC CGGTGGTGCA GAGCGTGCGA GGAGCTGGCA
961 CGCAGCCAGG ACCTTTGCTC AAGAAGCCAT ACCAGCCAAG AATTAAAATC TCTAAAACAT
1021 CAGGTAAAGC AAAGGATGCG GTTGTGTGTG GATTGAGAGT AAGAGATGTT TAAAAAGTGT
1081 AAAACCAGGC CAGGTGTACT GGCTCACGCC TGTAATCCCA GCACTTTGGG AGGCCGAGGT
1141 GGGTGTATTG CTTGAGGTCA GGAGTTCAAG ACCAGCCTGG CCAACATGGT GCAACCTCGT
1201 CTCTACTAAG AATACAAAAA TTAGCCAGGC ATGGTGGCAG GGGCCTGTAG TTCCAGCTAC
1261 TGGGGAGGCT GAGGCAGGAG AATCACTTGA ACCTGGGAAG TTGAGGTTAC AGTGAGCCGA
1321 GATTGTGCCA CCACACTCTA GCCTGGACTA CAAGAGTGAA ACTCTGTCTC AAAAAAAAAA
1381 AAAAAAAAAA AAAAAAAAAA AAAAA
B: Nucleotide sequence (SEQ ID NO: 23) Length: 244
1 MRTYDTPGTS MCIIPDDPHR NPRRFYDEIR TPLLSDIRID YPPSSVVQAT KTLFPNYFNG
61 SEIIIAGKLV DRKLDHLHVE VTASNSKKFI ILKTDVPVRP QKAGKDVTGS PRPGGDGEGD
121 PNHIERLWSY LTTKELLSSW LQSDDEPEKE RLRQRAQALA VSYRFLTPFT SMKLRGPVPR
181 MDGLEEAHGM SAAMGPEPVV QSVRGAGTQP GPLLKKPYQP RIKISKTSGK AKDAVVCGLR
241 VRDV
Nucleotide and amino acid sequence of C. Combination (SEQ ID NO: 24) Clone: PP14776
Start codon: 339 ATG termination codon: 1071 TAA Protein Weight: 27185.72
1 GT TTG GGT AGA TTT ATA GGT GTT TAA TAG CCA TAG TCA AAG CAC ATT 47
48 TAT TAA TGA TAA ATG GAA ATG GAG ACC CCT AAG GTA TGC AGT AAA GAC 95
96 AGG TCC TCG GTT TGG GAT TCG CCA CCG TTT CCT CCC ACG CAT TTT AAT 143
144 GAA AGA TTT AAA TGA AAA CAC AGA TTT GCT TTC CAC ATT TTA AAA ATT 191
192 ATT CAA AAC TAG AGT CAA AGG GGA GAA TCT GAT ACA CTG GAT AAC AGA 239
240 ATT GAA ATT CAA TAA GGC ATT CAG AAG CAA AAG CAG TTA TGC TGT AAC 287
288 AAG TCA GCT GCA CTG CCA TCC CAC TGA AAC TAT GAT CGA TAC CTG GCA 335
336 TTT ATG AGA ACT TAT GAT ACA CCA GGA ACT TCG ATG TGC ATC ATC CCA 383
1 Met Arg Thr Tyr Asp Thr Pro Gly Thr Ser Met Cys Ile Ile Pro 15
384 GAT GAT CCT CAC AGA AAT CCT AGG AGG TTC TAC GAT GAA ATC AGG ACC 431
16 Asp Asp Pro His Arg Asn Pro Arg Arg Phe Tyr Asp Glu Ile Arg Thr 31
432 CCG CTC CTC TCT GAC ATC CGC ATC GAT TAT CCC CCC AGC TCA GTG GTG 479
32 Pro Leu Leu Ser Asp Ile Arg Ile Asp Tyr Pro Pro Ser Ser Val Val 47
480 CAG GCC ACC AAG ACC CTG TTC CCC AAC TAC TTC AAC GGC TCG GAG ATC 527
48 Gln Ala Thr Lys Thr Leu Phe Pro Asn Tyr Phe Asn Gly Ser Glu Ile 63
528 ATC ATT GCG GGG AAG CTG GTG GAC AGG AAG CTG GAT CAC CTG CAC GTG 575
64 Ile Ile Ala Gly Lys Leu Val Asp Arg Lys Leu Asp His Leu His Val 79
576 GAG GTC ACC GCC AGC AAC AGT AAG AAA TTC ATC ATC CTG AAG ACA GAT 623
80 Glu Val Thr Ala Ser Asn Ser Lys Lys Phe Ile Ile Leu Lys Thr Asp 95
624 GTG CCT GTG CGG CCT CAG AAG GCA GGG AAA GAT GTC ACA GGA AGC CCC 671
96 Val Pro Val Arg Pro Gln Lys Ala Gly Lys Asp Val Thr Gly Ser Pro 111
672 AGG CCT GGA GGC GAT GGA GAG GGG GAC CCC AAC CAC ATC GAG CGT CTC 719
112 Arg Pro Gly Gly Asp Gly Glu Gly Asp Pro Asn His Ile Glu Arg Leu 127
720 TGG AGC TAC CTC ACC ACA AAG GAG CTG CTG AGC TCC TGG CTG CAA AGT 767
128 Trp Ser Tyr Leu Thr Thr Lys Glu Leu Leu Ser Ser Trp Leu Gln Ser 143
768 GAC GAT GAA CCG GAG AAG GAG CGG CTG CGG CAG CGG GCC CAG GCC CTG 815
144 Asp Asp Glu Pro Glu Lys Glu Arg Leu Arg Gln Arg Ala Gln Ala Leu 159
816 GCT GTG AGC TAC CGC TTC CTC ACT CCC TTC ACC TCC ATG AAG CTG AGG 863
160 Ala Val Ser Tyr Arg Phe Leu Thr Pro Phe Thr Ser Met Lys Leu Arg 175
864 GGG CCG GTC CCA CGC ATG GAC GGC CTG GAG GAG GCC CAC GGC ATG TCG 911
176 Gly Pro Val Pro Arg Met Asp Gly Leu Glu Glu Ala His Gly Met Ser 191
912 GCT GCC ATG GGA CCC GAA CCG GTG GTG CAG AGC GTG CGA GGA GCT GGC 959
192 Ala Ala Met Gly Pro Glu Pro Val Val Gln Ser Val Arg Gly Ala Gly 207
960 ACG CAG CCA GGA CCT TTG CTC AAG AAG CCA TAC CAG CCA AGA ATT AAA 1007
208 Thr Gln Pro Gly Pro Leu Leu Lys Lys Pro Tyr Gln Pro Arg Ile Lys 223
1008 ATC TCT AAA ACA TCA GGT AAA GCA AAG GAT GCG GTT GTG TGT GGA TTG 1055
224 Ile Ser Lys Thr Ser Gly Lys Ala Lys Asp Ala Val Val Cys Gly Leu 239
1056 AGA GTA AGA GAT GTT TAA AAA GTG TAA AAC CAG GCC AGG TGT ACT GGC 1103
240 Arg Val Arg Asp Val *** 245
1104 TCA CGC CTG TAA TCC CAG CAC TTT GGG AGG CCG AGG TGG GTG TAT TGC 1151
1152 TTG AGG TCA GGA GTT CAA GAC CAG CCT GGC CAA CAT GGT GCA ACC TCG 1199
1200 TCT CTA CTA AGA ATA CAA AAA TTA GCC AGG CAT GGT GGC AGG GGC CTG 1247
1248 TAG TTC CAG CTA CTG GGG AGG CTG AGG CAG GAG AAT CAC TTG AAC CTG 1295
1296 GGA AGT TGA GGT TAC AGT GAG CCG AGA TTG TGC CAC CAC ACT CTA GCC 1343
1344 TGG ACT ACA AGA GTG AAA CTC TGT CTC AAA AAA AAA AAA AAA AAA AAA 1391
1392 AAA AAA AAA AAA AA 1405
...
Claims (10)
1. isolating people's albumen with promotion 3T3 cell transformation function is characterized in that it comprises the polypeptide of the aminoacid sequence with the group of being selected from down: SEQ ID NO:2,5,8,11,14,17,20,23;
Or its conservative property variation polypeptide or its active fragments or its reactive derivative.
2. polypeptide as claimed in claim 1 is characterized in that, this polypeptide is the polypeptide with aminoacid sequence of the group of being selected from down: SEQ ID NO:2,5,8,11,14,17,20,23.
3. isolating polynucleotide is characterized in that, it comprises a nucleotide sequence, and this nucleotide sequence is shown at least 85% homogeny with a kind of nucleotides sequence that is selected from down group:
(a) coding is as the polynucleotide of polypeptide as described in claim 1 and 2;
(b) with polynucleotide (a) complementary polynucleotide.
4. polynucleotide as claimed in claim 3 is characterized in that, the polypeptide of this polynucleotide encoding has the aminoacid sequence of the group of being selected from down: SEQ ID NO:2,5,8,11,14,17,20,23.
5. polynucleotide as claimed in claim 3 is characterized in that, the sequence of these polynucleotide is selected from down group:
SEQ ID NO:3,6,9,12,15,18,21,24 coding region sequence or full length sequence.
6. a carrier is characterized in that, it contains the described polynucleotide of claim 3.
7. a genetically engineered host cell is characterized in that, it is a kind of host cell that is selected from down group:
(a) host cell that transforms or transduce with the described carrier of claim 6;
(b) host cell that transforms or transduce with the described polynucleotide of claim 3.
8. preparation method with polypeptide of the people's protein-active that promotes 3T3 cell transformation function is characterized in that this method comprises:
(a) being fit to express under the proteic condition of people with promotion 3T3 cell transformation function, cultivate the described host cell of claim 7;
(b) from culture, isolate polypeptide with the people's protein-active that promotes 3T3 cell transformation function.
9. an energy and claim 1 are described has the people's protein-specific bonded antibody that promotes 3T3 cell transformation function.
10. nucleic acid molecule, it contains a successive 10-800 Nucleotide in the described polynucleotide of claim 3.
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