CN106947764B - Plant root specific promoter and application thereof - Google Patents

Abstract

The invention discloses a plant root specific promoter and application thereof. The DNA molecule provided by the invention is named as GmGRPLp-1999 and is (1) or (2) or (3) as follows: (1) DNA molecule shown in sequence 1 in the sequence table; (2) a DNA molecule which is hybridized with the DNA sequence defined in 1) under strict conditions and has the function of a promoter; (3) a DNA molecule which has more than 90% of homology with the DNA sequence limited by 1) and has a promoter function. The invention also protects the application of the DNA molecule as a promoter. The invention provides a means for the molecular improvement of the plant at the root, has certain significance for the stress resistance research of the plant, and has wide application space and market prospect in the agricultural field.

Description

Plant root specific promoter and application thereof

Technical Field

The invention relates to a plant root specific promoter and application thereof.

Background

Promoters are parts of genes that control the initiation and extent of expression of a gene. In transgenic breeding, the precise regulation and control of foreign genes in plants is mainly realized by proper promoters. At present, a constitutive promoter is widely used in genetic engineering, but as the constitutive promoter drives the expression of a target gene in plant tissues to be constant and continuous, the constitutive promoter is not limited by time, space and external factors, substances and energy in cells are excessively consumed, a large amount of heterologous proteins or metabolites are generated, the original physiological metabolic balance of plants is damaged, and the plants grow abnormally and even die. Compared with a constitutive promoter, the root-specific promoter drives the exogenous gene to express in the root of a receptor plant, and the excessive consumption of cell substances caused by the continuous and efficient starting of the nonspecific expression of the exogenous gene of the constitutive promoter is overcome. With the increasing safety standards of transgenic plants, including all commercial transgenic crops, it is required that detailed studies must be made on the expression activity of the promoter used, the potential recombination capability, the influence on the surrounding environment and safety.

The soybean is an important crop used for both grain and oil feeding, is mostly in drought, saline-alkali, alpine and other areas in soybean production areas in China, and the applied soybean variety has weak stress tolerance, and serious abiotic and biotic adversity stresses such as drought and waterlogging diseases and insect pests obviously influence the soybean yield. The CaMV35S promoter is currently used in transgenic soybean.

Disclosure of Invention

The invention aims to provide a plant root specific promoter and application thereof.

The invention provides a DNA molecule, named GmGRPLp-1999, obtained from cultivated soybean Williams 82, which is (1) or (2) or (3) as follows:

(1) DNA molecule shown in sequence 1 in the sequence table;

(2) a DNA molecule which is hybridized with the DNA sequence defined in the step (1) under strict conditions and has the function of a promoter;

(3) and (2) a DNA molecule having a promoter function and having 90% or more homology with the DNA sequence defined in (1).

The stringent conditions may be hybridization and membrane washing at 65 ℃ in a solution of 0.1 XSSPE (or 0.1 XSSC), 0.1% SDS.

The recombinant vector, the expression cassette, the transgenic cell line or the recombinant strain containing the DNA molecule belong to the protection scope of the invention.

The recombinant vector can be a recombinant plasmid obtained by inserting the DNA molecule into a multiple cloning site or a recombination site of a plant expression vector. The plant expression vector can be specifically a pC13P1 vector. The recombinant vector can be specifically a recombinant plasmid obtained by inserting the DNA molecule into the multiple cloning site of the pC13P1 vector. The recombinant vector can be specifically a recombinant plasmid obtained by replacing a small fragment between Kpn I and Pst I of the pC13P1 vector by the DNA molecule.

The pC13P1 vector is a circular plasmid shown in a sequence 2 of a sequence table.

The invention also protects the application of the DNA molecule as a promoter. In the application, the promoter is specifically used as a root-specific promoter. In the application, the promoter is specifically used as a plant root specific promoter.

The invention also protects the application of the DNA molecule in promoting the expression of target genes. The promoting expression of the gene of interest may be promoting specific expression of the gene of interest. The specific expression may specifically be root-specific expression. The root-specific expression may specifically be specific expression at the root tip and/or at the post of the root. The promoting of the expression of the target gene can be specifically promoting the expression of the target gene in a plant. The target gene can be GUS gene.

The plant may be a monocot or a dicot. The dicot may be an angiosperm. The angiosperm plant can be a plant of the order Viridales. The plant of the order Sophora may be a plant of the family Leguminosae. The leguminous plant may be a plant of the genus glycine. The plant of Glycine may be soybean, such as cultivated soybean Williams 82.

The plant may be a monocot or a dicot. The dicot may be an angiosperm. The angiosperm plant can be plant of order Capparis. The plant of order Capparis can be a plant of the family Brassicaceae. The cruciferous plant may be an arabidopsis plant. The arabidopsis plant may specifically be arabidopsis thaliana, e.g. type Columbia arabidopsis thaliana.

The invention also provides a method for cultivating the transgenic plant, which is to use the DNA molecule to start the expression of the target gene in the starting plant to obtain the transgenic plant expressing the target gene.

The "expressing the gene of interest" may be "specifically expressing the gene of interest in roots".

The "expressing the target gene" may specifically be "specifically expressing the target gene in the root tip".

The "expressing the target gene" may specifically be "specifically expressing the target gene in the post-root column".

The starting plant may be a monocot or a dicot. The dicot may be an angiosperm. The angiosperm plant can be a plant of the order Viridales. The plant of the order Sophora may be a plant of the family Leguminosae. The leguminous plant may be a plant of the genus glycine. The plant of Glycine may be soybean, such as cultivated soybean Williams 82.

The angiosperm plant can be plant of order Capparis. The plant of order Capparis can be a plant of the family Brassicaceae. The cruciferous plant may be an arabidopsis plant. The arabidopsis plant may specifically be arabidopsis thaliana, e.g. type Columbia arabidopsis thaliana.

The target gene can be GUS gene.

In the method, a specific DNA molecule can be introduced into the starting plant, so that the promoter is used for promoting the expression of a target gene in the starting plant; the specific DNA molecule comprises the DNA molecule and the target gene from upstream to downstream in sequence. The specific DNA molecule can be specifically introduced into the starting plant through a recombinant expression vector containing the specific DNA molecule. The recombinant expression vector may be any of the above recombinant vectors.

The invention also provides a method for preparing the target protein, which is to use the DNA molecule to start the expression of the coding gene of the target protein in a starting plant to obtain the target protein.

The protein of interest is specifically present in plant roots.

The protein of interest is specifically present in the root tip and/or the root center pillar of the plant.

The starting plant may be a monocot or a dicot. The dicot may be an angiosperm. The angiosperm plant can be a plant of the order Viridales. The plant of the order Sophora may be a plant of the family Leguminosae. The leguminous plant may be a plant of the genus glycine. The plant of Glycine may be soybean, such as cultivated soybean Williams 82.

The angiosperm plant can be plant of order Capparis. The plant of order Capparis can be a plant of the family Brassicaceae. The cruciferous plant may be an arabidopsis plant. The arabidopsis plant may specifically be arabidopsis thaliana, e.g. type Columbia arabidopsis thaliana.

The target protein can be GUS. The coding gene of the target protein can be GUS gene.

In the method, a specific DNA molecule can be introduced into the starting plant, so that the expression of the gene coding for the target protein is initiated in the starting plant by the DNA molecule; the specific DNA molecule comprises the DNA molecule and the target gene from upstream to downstream in sequence. The specific DNA molecule can be specifically introduced into the starting plant through a recombinant expression vector containing the specific DNA molecule. The recombinant expression vector may be any of the recombinant vectors described above.

The invention discloses a soybean-derived root-specific promoter GmGRPLp-1999, wherein the promoter GmGRPLp-1999 can effectively promote the specific expression of a target gene in a root tip and a root central cylinder tissue. The invention is applied to the research of transgenic plants, can provide richer promoter elements for the research of the transgenic plants, and can also provide a means for cultivating new varieties of transgenic plants with drought resistance, salt resistance, cold resistance, aluminum resistance, heavy metal resistance and the like. The invention provides a means for the molecular improvement of the plant at the root, has certain significance for the stress resistance research of the plant, and has wide application space and market prospect in the agricultural field.

Drawings

FIG. 1 is a plasmid map of pC13P1 vector.

FIG. 2 shows the soybean transformation procedure.

FIG. 3 shows the GUS staining results of transgenic soybean hairy roots.

FIG. 4 is a plasmid map of pC13(Delta) GUS vector.

FIG. 5 is transgenic Arabidopsis T₁And (5) generation molecular detection results. Wherein, lane M is a DNA molecular weight standard of DL2000 plus.

FIG. 6 is transgenic Arabidopsis T₃And (5) performing GUS staining on the generation plants.

Detailed Description

The following examples are given to facilitate a better understanding of the invention, but do not limit the invention. The experimental procedures in the following examples are conventional unless otherwise specified. The test materials used in the following examples were purchased from a conventional biochemical reagent store unless otherwise specified. The quantitative tests in the following examples, all set up three replicates and the results averaged.

Cultivation of soybean Williams 82: reference documents: functional analysis of wild soybean salt stress-related micrornas [ D ]. chinese academy of agricultural sciences, 2011; the public is available from the institute of crop science, academy of agricultural sciences, china.

pC13P1 vector: a circular plasmid shown in a sequence 2 of a sequence table. The pC13P1 vector does not contain any promoter, but contains a GUS reporter gene. The plasmid map of the pC13P1 vector is shown in FIG. 1.

pC13(Delt) GUS vector: a circular plasmid shown in a sequence 3 of a sequence table. The pC13(Delt) GUS vector does not contain the GUS gene, but contains kanamycin resistance and hygromycin resistance. The plasmid map of pC13(Delt) GUS vector is shown in FIG. 4.

Agrobacterium tumefaciens GV 3101: reference documents: effect of different agrobacterium tumefaciens strain types on trichoderma genetic transformation efficiency [ J ] northern horticulture, 2006 (3): 14-15.; the public is available from the institute of crop science, academy of agricultural sciences, china.

Columbia type Arabidopsis thaliana: reference documents: crowning. Columbia type arabidopsis callus culture study [ J ]. south river agriculture, 2009 (1): 42-44; the public is available from the institute of crop science, academy of agricultural sciences, china.

The YEP liquid culture medium consists of a solvent and a solvent; the solutes and their concentrations in YEP liquid medium were: NaCl 5g/L, yeast extract 5g/L, tryptone 10 g/L; the solvent is water. YEP broth pH was 7.0.

The germination culture medium consists of a solvent and a solvent; the solutes and their concentrations in the germination medium are: b5 salt 3.1g/L, sucrose 20g/L, agar 7g/L, 1 ml/L; the solvent is water. The pH of the germination medium was 5.8.

The co-culture medium consists of a solvent and a solvent; the solutes and their concentrations in the co-cultivation medium are: MS salt 0.31g/L, MES 3.9g/L, sucrose 30g/L, agar 7g/L, MS organic 1ml/L, DTT 150mg/L, AS 0.02 g/L; the solvent is water. The pH of the co-cultivation medium was 5.4.

The induction culture medium consists of a solvent and a solvent; the solutes and their concentrations in the induction medium are: 2.165g/L of MS salt, 30g/L of cane sugar, 7g/L of agar, 1ml/L of MS organic matter, 250mg/L of cefuroxime axetil and 250mg/L of carbenicillin sodium; the solvent is water. The pH of the induction medium was 5.8.

B5 salt: phyto Tech, catalog No.: G768.

MS organic: phyto Tech, catalog No.: and M533.

MS salt: phyto Tech, catalog No.: and M524.

DTT: Sigma-Aldrich, Cat number: D5545.

AS: Sigma-Aldrich, Cat number: D134406.

and (3) cephamycin: Sigma-Aldrich, Cat number: C7039.

carbenicillin sodium: incoco, good No.: 1758-9317.

GUS histochemical staining procedures in the following examples are as follows: the tissue sample to be stained is placed in the detection solution and incubated at 37 ℃ for 12-16 h. The staining of the samples was observed and photographed. Blue under white background is GUS expression site.

Composition of the detection solution: 50mmol/L phosphate buffer (4.095g Na)₂HPO₄And 2.537g NaH₂PO₄Constant volume to 1L waterIn solution), 10mmol/L EDTA (pH 7.0), 0.1% (mass percent) Triton X-100, 2mmol/L potassium ferricyanide, 2mmol/L potassium ferrocyanide, 5% (mass percent) X-Gluc, and the balance of water.

Example 1 discovery of promoters

A promoter is found from soybean root genome DNA by taking cultivated soybean Williams 82 as a research material through a large amount of sequence analysis, expression analysis and functional verification, and is named as GmGRPLp-1999, and is shown as a sequence 1 in a sequence table.

Example 2 construction of recombinant expression vector containing GmGRPLp-1999 promoter

1. Extracting the genomic DNA of the root system of the cultivated soybean Williams 82.

2. And (3) taking the genomic DNA obtained in the step (1) as a template, carrying out PCR amplification by adopting a primer pair consisting of a primer F-1999 and a primer R-1999, and recovering a PCR amplification product of about 2000 bp.

F-1999：5′-GGTACCAATAGTAGGCTTCTCTAG-3′；

R-1999：5′-CTGCAGCTTAAACTTTTTAGGG-3′。

F-1999 and R-1999, the Kpn I and Pst I cleavage sites are underlined, respectively.

3. And (3) carrying out double digestion on the PCR amplification product in the step 2 by using restriction enzymes Kpn I and Pst I, and recovering the digestion product.

4. The pC13P1 vector was double-digested with restriction enzymes Kpn I and Pst I, and the vector backbone of about 8700bp was recovered.

5. And (4) connecting the enzyme digestion product obtained in the step (3) with the vector framework obtained in the step (4) to obtain a recombinant expression vector pCAM-GRPLp-1999. Based on the sequencing results, the recombinant expression vector pCAM-GRPLp-1999 was structurally described as follows: the small fragment between Kpn I and Pst I cleavage sites of pC13P1 vector was substituted for the double-stranded DNA molecule shown as 1-1999 nucleotides from the 5' end of sequence 1 in the sequence listing.

Example 3 acquisition and expression characterization of transgenic soybean hairy root GmGRPLp-1999

1. The recombinant expression vector pCAM-GRPLp-1999 obtained in example 2 was introduced into Agrobacterium rhizogenes K599 to obtain recombinant Agrobacterium.

2. Inoculating the recombinant Agrobacterium obtained in step 1 into YEP liquid culture medium (containing 50mg/L kanamycin), and shake culturing at 28 deg.C and 200rmp to obtain OD_600nm0.6-0.8 of recombinant agrobacterium liquid.

3. Selecting seeds of cultivated soybean Williams 82 without plant diseases and insect pests, placing the seeds in a culture dish, placing the culture dish in a dryer, adding 100ml of sodium hypochlorite into a beaker of the dryer, adding 3.5ml of 12N hydrochloric acid into the beaker, closing the dryer, and placing for 16h for disinfection (figure 2A).

4. The seeds sterilized in step 3 were inoculated into a germination medium (FIG. 2B), and cultured at 28 ℃ for 5 days. After 5 days (FIG. 2C), the soybean cotyledons were removed.

5. Cutting the soybean cotyledon with hypocotyl obtained in step 4 with a knife, cutting off the redundant hypocotyl, keeping the hypocotyl with 0.5cm length of cotyledon, separating cotyledon node from the middle with a blade to obtain two single leaves (figure 2D), and scratching the joint of cotyledon and hypocotyl with a blade for 7-8 times. And (3) soaking the treated cotyledon node in the recombinant agrobacterium liquid obtained in the step (2) for 30min (fig. 2E).

6. The cotyledonary node soaked in step 5 was transferred to the co-culture medium and cultured in the dark at 22 ℃ for 5 days (FIG. 2F).

7. The cotyledonary node cultured in step 6 was washed 3 times in 1/2MS liquid medium, transferred to induction medium, and cultured at 28 ℃ for 10-12 days, and a large amount of hairy roots were observed to be generated from the scratched part (FIG. 2G). Hairy roots were cut from the cotyledon base and subjected to GUS histochemical staining.

8. Agrobacterium rhizogenes K599 was used instead of recombinant Agrobacterium as a Control (CK) following steps 2-7.

The results are shown in FIG. 3. As can be seen from the figure, in hairy roots transformed into GmGRPLp-1999, the expression activity of GUS gene was detected only in the root tip and the middle column (blue color), and was not detected in other tissues. No GUS expression was observed in the control hairy roots transformed with Agrobacterium rhizogenes K599. The results of three repeated experiments are consistent.

Example 4 acquisition and expression characterization of transgenic Arabidopsis thaliana GmGRPLp-1999

1. The recombinant expression vector pCAM-GRPLp-1999 obtained in example 2 was introduced into Agrobacterium tumefaciens GV3101 to obtain recombinant Agrobacterium tumefaciens GRPLp-1999.

2. The pC13(Delt) GUS vector was introduced into Agrobacterium tumefaciens GV3101 to obtain recombinant Agrobacterium pC13(Delt) GUS.

3. Inoculating the recombinant Agrobacterium GRPLp-1999 obtained in step 1 into YEP liquid medium (containing 50mg/L kanamycin), shaking-culturing at 28 deg.C and 220rmp to obtain OD_600nm0.4-0.6 of recombinant agrobacterium liquid GRPLp-1999.

4. Inoculating the recombinant Agrobacterium pC13(Delt) GUS obtained in the step 2 into YEP liquid culture medium (containing 50mg/L kanamycin), and performing shake culture at 28 ℃ and 200rmp to obtain OD_600nm0.4-0.6. the recombinant agrobacterium liquid is recombinant agrobacterium pC13(Delt) GUS.

5. Mixing 1 volume part of the recombinant agrobacterium liquid GRPLp-1999 obtained in the step 3 and 1 volume part of the recombinant agrobacterium liquid pC13(Delt) GUS obtained in the step 4, and adding silwet L-77 to obtain a mixed liquid, wherein the volume percentage of the silwet L-77 in the mixed liquid is 0.5%.

6. The Columbia arabidopsis thaliana at the flowering stage is taken, and the infection is carried out once every week until T1 seeds are harvested.

The steps of each infection are as follows: and (4) soaking the mixture in the mixed bacterial liquid obtained in the step (5) for 1min, shading the mixture overnight after soaking, and recovering normal culture (16 h/8 h in darkness) the next day.

T₁The plant grown by the seed generation is T₁And (5) plant generation. From T₁The seeds produced by the selfing of the generation plants are T₂And (5) seed generation. From T₂The plant grown by the seed generation is T₂And (5) plant generation. From T₂The seeds produced by the selfing of the generation plants are T₃And (5) seed generation. From T₃The plant grown by the seed generation is T₃And (5) plant generation.

7. The T obtained in the step 6₁Inoculating the seeds to 1/2MS culture medium containing 50mg/L hygromycin, and screening T capable of normally growing₁And (5) plant generation. Extraction of T₁The genome DNA of the generation plant is taken as a template, and a primer pair consisting of a primer F-1999 and a primer R-1999 is adoptedPCR detection was performed. The recombinant expression vector pCAM-GRPLp-1999 was used as a positive control.

If an amplification product of about 1999bp is obtained, the plant to be detected is a transgenic positive plant, if the amplification product is not obtained, the plant to be detected is a non-transgenic plant (part of the detection results are shown in figure 5. in figure 5, CK + is a positive control, and lanes 1-7 are randomly selected T₁A progeny plant).

Identifying T as positive for transgene₁The plant generation is continued to be cultured at T₃And generating to obtain transgenic homozygous lines.

8. The pC13P1 vector is adopted to replace a recombinant expression vector pCAM-GRPLp-1999, and the operation is carried out according to the steps 1 to 7, so as to obtain a transgenic empty vector strain.

9. Respectively taking Columbia type arabidopsis thaliana (wild type) plants and T of transgenic homozygous lines in the same period (3 days and 10 days of emergence)₃T of generation plant and empty vector line₃GUS staining is carried out on the generation plants.

The results are shown in FIG. 6. The results showed that the expression activity of GUS gene was detectable in the root tip of transgenic plants (blue color) and not in other tissues. There was no GUS expression in both Columbia type Arabidopsis (CK) and empty vector Arabidopsis plants. The results of three repeated experiments are consistent.

<110> institute of crop science of Chinese academy of agricultural sciences

<120> plant root specific promoter and application thereof

<160>3

<210>1

<211>1999

<212>DNA

<213> Soybean (Glycine max (Linn.) Merr.)

<400>1

aatagtaggc ttctctagtt tctttttcaa cctaaagaat taaaattgat gaaaaataag 60

aaaatgatat tttattattt gtatttataa ataaaaaata ggtgaaaaat aaaataatta 120

tggatatttg atttaagaga aataaaaaaa taagaaaaaa aatacaagat aatccattgt 180

cttttatggc aatttattgt taaaattata attttgaaaa tttgactgat cgacctgtta 240

aactaggaac ccacataatg accggttcag ttattctaaa aaagcaaact atttcagaat 300

tggtgagcac ttgtcaaatc ggtctaaaat caaccaaaat cggtataaaa tcagatagaa 360

attggttaat ttgaactggt attctgattt tttttctgtt tctttaaatt taatataaaa 420

tattcaaaga taattaaaaa cacatattta atcaagatta aaaataggta taaattttct 480

tattgatgat ttattttata aaacacataa tttactcact tatgttaatt tactattttt 540

ttatcttaat tttatttata attttatatt atattaattt tttttctttt ctattttaga 600

tgtgattttt ttatttttaa tgaatttttt attttattaa aatttaactt ttattttgat 660

attttttaac ttttatttgt actttcaatg ttataaacct ggaaaaatga tactttattt 720

tagtttatga tgctaagaag ttatgatatt attattatta ttattaataa taaaatcttg 780

aactaatgaa ccttaaatca gtacattgaa ccgttcggtt caatttcaaa aatattggtt 840

aaaactatga aaaatcgatc atgaattaac tttgacaata aactatttta gtttttccac 900

ggtaagacaa ttgattatga atgaaaacca tcaattattt tcaataaaaa aaagttcttt 960

tctattttcc caccattttt atgcaatcaa acgaagcttt atggcgcata tgtagacaat 1020

tttttttgaa caaaagagta atgtaggtat tgaaattacg ttccacggtt ccacctgtca 1080

aagtttctga ttacaatagt tcttgagatt gtatatcttg cctatcttga aagcaactgc 1140

aaaaggatgt caatcaggaa aaaaaaaaag agatgtcaat tagagccaaa aagattggac 1200

taaaattata atatttccaa tattattaag gaacgggagg tgggaaacgt acatgtatgg 1260

aaagaaagga acgtgaggtg ggaaacgtac atgtatggaa ggaaggaaga ggaaaaaaga 1320

gagtagaaaa agataataaa taatgtgata taaaaaaaat aaataaaaaa agatgaaaag 1380

aatttgaaaa gaataaatat atgttaataa taacggtaga aaaaatacat aaaaaaataa 1440

cgtaaagaaa acaatcttgt cttctttttg taaaggatat tatatatgac tattaacgaa 1500

agcctttttt agagaggtta aggctatgct tctataaaag ttaagttctc ttaaacgaaa 1560

aagcaacaaa tttttactct tacacttgct actttgaaaa gtgtaaaatt aaaattgcct 1620

tggaaagcac gcaacaaaat ttctaattga ttttcaagca taccctaaat ttaactaact 1680

tacaacttag atgtgagacc ctacatcacc tcacaaaata caaaacctag ttcaagggtt 1740

tctctgccag ctaaaactca ttctaataca tggttattgc ttcattaaca tagtttctct 1800

caaatccgtt aaactatatt ttaataatta ttacctcgat gaccattttc ctaaacaatt 1860

taccccactt gcatttcaaa gccctatata aaccaccatt cactgccaat gactcccatc 1920

cccacaaaag ctcttaagct cccgtagtta aatctgcgag ccattagtta aagaaaaaaa 1980

aaaccctaaa aagtttaag 1999

<210>2

<211>8766

<212>DNA

<213> Artificial sequence

<220>

<223>

<400>2

cgagctcggt acccggggat cctgcagtct agaagcttcc atggtagatc tgagggtaaa 60

tttctagttt ttctccttca ttttcttggt taggaccctt ttctcttttt atttttttga 120

gctttgatct ttctttaaac tgatctattt tttaattgat tggttatggt gtaaatatta 180

catagcttta actgataatc tgattacttt atttcgtgtg tctatgatga tgatgatagt 240

tacagaaccg acgactcgtc cgtcctgtag aaaccccaac ccgtgaaatc aaaaaactcg 300

acggcctgtg ggcattcagt ctggatcgcg aaaactgtgg aattgatcag cgttggtggg 360

aaagcgcgtt acaagaaagc cgggcaattg ctgtgccagg cagttttaac gatcagttcg 420

ccgatgcaga tattcgtaat tatgcgggca acgtctggta tcagcgcgaa gtctttatac 480

cgaaaggttg ggcaggccag cgtatcgtgc tgcgtttcga tgcggtcact cattacggca 540

aagtgtgggt caataatcag gaagtgatgg agcatcaggg cggctatacg ccatttgaag 600

ccgatgtcac gccgtatgtt attgccggga aaagtgtacg tatcaccgtt tgtgtgaaca 660

acgaactgaa ctggcagact atcccgccgg gaatggtgat taccgacgaa aacggcaaga 720

aaaagcagtc ttacttccat gatttcttta actatgccgg aatccatcgc agcgtaatgc 780

tctacaccac gccgaacacc tgggtggacg atatcaccgt ggtgacgcat gtcgcgcaag 840

actgtaacca cgcgtctgtt gactggcagg tggtggccaa tggtgatgtc agcgttgaac 900

tgcgtgatgc ggatcaacag gtggttgcaa ctggacaagg cactagcggg actttgcaag 960

tggtgaatcc gcacctctgg caaccgggtg aaggttatct ctatgaactc gaagtcacag 1020

ccaaaagcca gacagagtct gatatctacc cgcttcgcgt cggcatccgg tcagtggcag 1080

tgaagggcca acagttcctg attaaccaca aaccgttcta ctttactggc tttggtcgtc 1140

atgaagatgc ggacttacgt ggcaaaggat tcgataacgt gctgatggtg cacgaccacg 1200

cattaatgga ctggattggg gccaactcct accgtacctc gcattaccct tacgctgaag 1260

agatgctcga ctgggcagat gaacatggca tcgtggtgat tgatgaaact gctgctgtcg 1320

gctttcagct gtctttaggc attggtttcg aagcgggcaa caagccgaaa gaactgtaca 1380

gcgaagaggc agtcaacggggaaactcagc aagcgcactt acaggcgatt aaagagctga 1440

tagcgcgtga caaaaaccac ccaagcgtgg tgatgtggag tattgccaac gaaccggata 1500

cccgtccgca aggtgcacgg gaatatttcg cgccactggc ggaagcaacg cgtaaactcg 1560

acccgacgcg tccgatcacc tgcgtcaatg taatgttctg cgacgctcac accgatacca 1620

tcagcgatct ctttgatgtg ctgtgcctga accgttatta cggatggtat gtccaaagcg 1680

gcgatttgga aacggcagag aaggtactgg aaaaagaact tctggcctgg caggagaaac 1740

tgcatcagcc gattatcatc accgaatacg gcgtggatac gttagccggg ctgcactcaa 1800

tgtacaccga catgtggagt gaagagtatc agtgtgcatg gctggatatg tatcaccgcg 1860

tctttgatcg cgtcagcgcc gtcgtcggtg aacaggtatg gaatttcgcc gattttgcga 1920

cctcgcaagg catattgcgc gttggcggta acaagaaagg gatcttcact cgcgaccgca 1980

aaccgaagtc ggcggctttt ctgctgcaaa aacgctggac tggcatgaac ttcggtgaaa 2040

aaccgcagca gggaggcaaa caagctagcc accaccacca ccaccacgtg tgaattacag 2100

gtgaccagct cgaatttccc cgatcgttca aacatttggc aataaagttt cttaagattg 2160

aatcctgttg ccggtcttgc gatgattatc atataatttc tgttgaatta cgttaagcat 2220

gtaataatta acatgtaatg catgacgtta tttatgagat gggtttttat gattagagtc 2280

ccgcaattat acatttaata cgcgatagaa aacaaaatat agcgcgcaaa ctaggataaa 2340

ttatcgcgcg cggtgtcatc tatgttacta gatcgggaat taaactatca gtgtttgaca 2400

ggatatattg gcgggtaaac ctaagagaaa agagcgttta ttagaataac ggatatttaa 2460

aagggcgtga aaaggtttat ccgttcgtcc atttgtatgt gcatgccaac cacagggttc 2520

ccctcgggat caaagtactt tgatccaacc cctccgctgc tatagtgcag tcggcttctg 2580

acgttcagtg cagccgtctt ctgaaaacga catgtcgcac aagtcctaag ttacgcgaca 2640

ggctgccgcc ctgccctttt cctggcgttt tcttgtcgcg tgttttagtc gcataaagta 2700

gaatacttgc gactagaacc ggagacatta cgccatgaac aagagcgccg ccgctggcct 2760

gctgggctat gcccgcgtca gcaccgacga ccaggacttg accaaccaac gggccgaact 2820

gcacgcggcc ggctgcacca agctgttttc cgagaagatc accggcacca ggcgcgaccg 2880

cccggagctg gccaggatgc ttgaccacct acgccctggc gacgttgtga cagtgaccag 2940

gctagaccgc ctggcccgca gcacccgcga cctactggac attgccgagc gcatccagga 3000

ggccggcgcg ggcctgcgta gcctggcaga gccgtgggcc gacaccacca cgccggccgg 3060

ccgcatggtg ttgaccgtgt tcgccggcat tgccgagttc gagcgttccc taatcatcga 3120

ccgcacccgg agcgggcgcg aggccgccaa ggcccgaggc gtgaagtttg gcccccgccc 3180

taccctcacc ccggcacaga tcgcgcacgc ccgcgagctg atcgaccagg aaggccgcac 3240

cgtgaaagag gcggctgcac tgcttggcgt gcatcgctcg accctgtacc gcgcacttga 3300

gcgcagcgag gaagtgacgc ccaccgaggc caggcggcgc ggtgccttcc gtgaggacgc 3360

attgaccgag gccgacgccc tggcggccgc cgagaatgaa cgccaagagg aacaagcatg 3420

aaaccgcacc aggacggcca ggacgaaccg tttttcatta ccgaagagat cgaggcggag 3480

atgatcgcgg ccgggtacgt gttcgagccg cccgcgcacg tctcaaccgt gcggctgcat 3540

gaaatcctgg ccggtttgtc tgatgccaag ctggcggcct ggccggccag cttggccgct 3600

gaagaaaccg agcgccgccg tctaaaaagg tgatgtgtat ttgagtaaaa cagcttgcgt 3660

catgcggtcg ctgcgtatat gatgcgatga gtaaataaac aaatacgcaa ggggaacgca 3720

tgaaggttat cgctgtactt aaccagaaag gcgggtcagg caagacgacc atcgcaaccc 3780

atctagcccg cgccctgcaa ctcgccgggg ccgatgttct gttagtcgat tccgatcccc 3840

agggcagtgc ccgcgattgg gcggccgtgc gggaagatca accgctaacc gttgtcggca 3900

tcgaccgccc gacgattgac cgcgacgtga aggccatcgg ccggcgcgac ttcgtagtga 3960

tcgacggagc gccccaggcg gcggacttgg ctgtgtccgc gatcaaggca gccgacttcg 4020

tgctgattcc ggtgcagcca agcccttacg acatatgggc caccgccgac ctggtggagc 4080

tggttaagca gcgcattgag gtcacggatg gaaggctaca agcggccttt gtcgtgtcgc 4140

gggcgatcaa aggcacgcgc atcggcggtg aggttgccga ggcgctggcc gggtacgagc 4200

tgcccattct tgagtcccgt atcacgcagc gcgtgagcta cccaggcact gccgccgccg 4260

gcacaaccgt tcttgaatca gaacccgagg gcgacgctgc ccgcgaggtc caggcgctgg 4320

ccgctgaaat taaatcaaaa ctcatttgag ttaatgaggt aaagagaaaa tgagcaaaag 4380

cacaaacacg ctaagtgccg gccgtccgag cgcacgcagc agcaaggctg caacgttggc 4440

cagcctggca gacacgccag ccatgaagcg ggtcaacttt cagttgccgg cggaggatca 4500

caccaagctg aagatgtacg cggtacgcca aggcaagacc attaccgagc tgctatctga 4560

atacatcgcg cagctaccag agtaaatgag caaatgaata aatgagtaga tgaattttag 4620

cggctaaagg aggcggcatg gaaaatcaag aacaaccagg caccgacgcc gtggaatgcc 4680

ccatgtgtgg aggaacgggc ggttggccag gcgtaagcgg ctgggttgtc tgccggccct 4740

gcaatggcac tggaaccccc aagcccgagg aatcggcgtg acggtcgcaa accatccggc 4800

ccggtacaaa tcggcgcggc gctgggtgat gacctggtgg agaagttgaa ggccgcgcag 4860

gccgcccagc ggcaacgcat cgaggcagaa gcacgccccg gtgaatcgtg gcaagcggcc 4920

gctgatcgaa tccgcaaaga atcccggcaa ccgccggcag ccggtgcgcc gtcgattagg 4980

aagccgccca agggcgacga gcaaccagat tttttcgttc cgatgctcta tgacgtgggc 5040

acccgcgata gtcgcagcat catggacgtg gccgttttcc gtctgtcgaa gcgtgaccga 5100

cgagctggcg aggtgatccg ctacgagctt ccagacgggc acgtagaggt ttccgcaggg 5160

ccggccggca tggccagtgt gtgggattac gacctggtac tgatggcggt ttcccatcta 5220

accgaatcca tgaaccgata ccgggaaggg aagggagaca agcccggccg cgtgttccgt 5280

ccacacgttg cggacgtact caagttctgc cggcgagccg atggcggaaa gcagaaagac 5340

gacctggtag aaacctgcat tcggttaaac accacgcacg ttgccatgca gcgtacgaag 5400

aaggccaaga acggccgcct ggtgacggta tccgagggtg aagccttgat tagccgctac 5460

aagatcgtaa agagcgaaac cgggcggccg gagtacatcg agatcgagct agctgattgg 5520

atgtaccgcg agatcacaga aggcaagaac ccggacgtgc tgacggttca ccccgattac 5580

tttttgatcg atcccggcat cggccgtttt ctctaccgcc tggcacgccg cgccgcaggc 5640

aaggcagaag ccagatggtt gttcaagacg atctacgaac gcagtggcag cgccggagag 5700

ttcaagaagt tctgtttcac cgtgcgcaag ctgatcgggt caaatgacct gccggagtac 5760

gatttgaagg aggaggcggg gcaggctggc ccgatcctag tcatgcgcta ccgcaacctg 5820

atcgagggcg aagcatccgc cggttcctaa tgtacggagc agatgctagg gcaaattgcc 5880

ctagcagggg aaaaaggtcg aaaaggtctc tttcctgtgg atagcacgta cattgggaac 5940

ccaaagccgt acattgggaa ccggaacccg tacattggga acccaaagcc gtacattggg 6000

aaccggtcac acatgtaagt gactgatata aaagagaaaa aaggcgattt ttccgcctaa 6060

aactctttaa aacttattaa aactcttaaa acccgcctgg cctgtgcata actgtctggc 6120

cagcgcacag ccgaagagct gcaaaaagcg cctacccttc ggtcgctgcg ctccctacgc 6180

cccgccgctt cgcgtcggcc tatcgcggcc gctggccgct caaaaatggc tggcctacgg 6240

ccaggcaatc taccagggcg cggacaagcc gcgccgtcgc cactcgaccg ccggcgccca 6300

catcaaggca ccctgcctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca 6360

gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca 6420

gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc atgacccagt cacgtagcga 6480

tagcggagtg tatactggct taactatgcg gcatcagagc agattgtact gagagtgcac 6540

catatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat caggcgctct 6600

tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca 6660

gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac 6720

atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt 6780

ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg 6840

cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc 6900

tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc 6960

gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc 7020

aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac 7080

tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt 7140

aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct 7200

aactacggct acactagaag gacagtattt ggtatctgcg ctctgctgaa gccagttacc 7260

ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt 7320

ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg 7380

atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc 7440

atgcattcta ggtactaaaa caattcatcc agtaaaatat aatattttat tttctcccaa 7500

tcaggcttga tccccagtaa gtcaaaaaat agctcgacat actgttcttc cccgatatcc 7560

tccctgatcg accggacgca gaaggcaatg tcataccact tgtccgccct gccgcttctc 7620

ccaagatcaa taaagccact tactttgcca tctttcacaa agatgttgct gtctcccagg 7680

tcgccgtggg aaaagacaag ttcctcttcg ggcttttccg tctttaaaaa atcatacagc 7740

tcgcgcggat ctttaaatgg agtgtcttct tcccagtttt cgcaatccac atcggccaga 7800

tcgttattca gtaagtaatc caattcggct aagcggctgt ctaagctatt cgtataggga 7860

caatccgata tgtcgatgga gtgaaagagc ctgatgcact ccgcatacag ctcgataatc 7920

ttttcagggc tttgttcatc ttcatactct tccgagcaaa ggacgccatc ggcctcactc 7980

atgagcagat tgctccagcc atcatgccgt tcaaagtgca ggacctttgg aacaggcagc 8040

tttccttcca gccatagcat catgtccttt tcccgttcca catcataggt ggtcccttta 8100

taccggctgt ccgtcatttt taaatatagg ttttcatttt ctcccaccag cttatatacc 8160

ttagcaggag acattccttc cgtatctttt acgcagcggt atttttcgat cagttttttc 8220

aattccggtg atattctcat tttagccatt tattatttcc ttcctctttt ctacagtatt 8280

taaagatacc ccaagaagct aattataaca agacgaactc caattcactg ttccttgcat 8340

tctaaaacct taaataccag aaaacagctt tttcaaagtt gttttcaaag ttggcgtata 8400

acatagtatc gacggagccg attttgaaac cgcggtgatc acaggcagca acgctctgtc 8460

atcgttacaa tcaacatgct accctccgcg agatcatccg tgtttcaaac ccggcagctt 8520

agttgccgtt cttccgaata gcatcggtaa catgagcaaa gtctgccgcc ttacaacggc 8580

tctcccgctg acgccgtccc ggactgatgg gctgcctgta tcgagtggtg attttgtgcc 8640

gagctgccgg tcggggagct gttggctggc tggtggcagg atatattgtg gtgtaaacaa 8700

attgacgctt agacaactta ataacacatt gcggacgttt ttaatgtact gaattaacgc 8760

cgaatt 8766

<210>3

<211>9041

<212>DNA

<213> Artificial sequence

<220>

<223>

<400>3

agctgtgaat tacaggtgac cagctcgaat ttccccgatc gttcaaacat ttggcaataa 60

agtttcttaa gattgaatcc tgttgccggt cttgcgatga ttatcatata atttctgttg 120

aattacgtta agcatgtaat aattaacatg taatgcatga cgttatttat gagatgggtt 180

tttatgatta gagtcccgca attatacatt taatacgcga tagaaaacaa aatatagcgc 240

gcaaactagg ataaattatc gcgcgcggtg tcatctatgt tactagatcg ggaattaaac 300

tatcagtgtt tgacaggata tattggcggg taaacctaag agaaaagagc gtttattaga 360

ataacggata tttaaaaggg cgtgaaaagg tttatccgtt cgtccatttg tatgtgcatg 420

ccaaccacag ggttcccctc gggatcaaag tactttgatc caacccctcc gctgctatag 480

tgcagtcggc ttctgacgtt cagtgcagcc gtcttctgaa aacgacatgt cgcacaagtc 540

ctaagttacg cgacaggctg ccgccctgcc cttttcctgg cgttttcttg tcgcgtgttt 600

tagtcgcata aagtagaata cttgcgacta gaaccggaga cattacgcca tgaacaagag 660

cgccgccgct ggcctgctgg gctatgcccg cgtcagcacc gacgaccagg acttgaccaa 720

ccaacgggcc gaactgcacg cggccggctg caccaagctg ttttccgaga agatcaccgg 780

caccaggcgc gaccgcccgg agctggccag gatgcttgac cacctacgcc ctggcgacgt 840

tgtgacagtg accaggctag accgcctggc ccgcagcacc cgcgacctac tggacattgc 900

cgagcgcatc caggaggccg gcgcgggcct gcgtagcctg gcagagccgt gggccgacac 960

caccacgccg gccggccgca tggtgttgac cgtgttcgcc ggcattgccg agttcgagcg 1020

ttccctaatc atcgaccgca cccggagcgg gcgcgaggcc gccaaggccc gaggcgtgaa 1080

gtttggcccc cgccctaccc tcaccccggc acagatcgcg cacgcccgcg agctgatcga 1140

ccaggaaggc cgcaccgtga aagaggcggc tgcactgctt ggcgtgcatc gctcgaccct 1200

gtaccgcgca cttgagcgca gcgaggaagt gacgcccacc gaggccaggc ggcgcggtgc 1260

cttccgtgag gacgcattga ccgaggccga cgccctggcg gccgccgaga atgaacgcca 1320

agaggaacaa gcatgaaacc gcaccaggac ggccaggacg aaccgttttt cattaccgaa 1380

gagatcgagg cggagatgat cgcggccggg tacgtgttcg agccgcccgc gcacgtctca 1440

accgtgcggc tgcatgaaat cctggccggt ttgtctgatg ccaagctggc ggcctggccg 1500

gccagcttgg ccgctgaaga aaccgagcgc cgccgtctaa aaaggtgatg tgtatttgag 1560

taaaacagct tgcgtcatgc ggtcgctgcg tatatgatgc gatgagtaaa taaacaaata 1620

cgcaagggga acgcatgaag gttatcgctg tacttaacca gaaaggcggg tcaggcaaga 1680

cgaccatcgc aacccatcta gcccgcgccc tgcaactcgc cggggccgat gttctgttag 1740

tcgattccga tccccagggc agtgcccgcg attgggcggc cgtgcgggaa gatcaaccgc 1800

taaccgttgt cggcatcgac cgcccgacga ttgaccgcga cgtgaaggcc atcggccggc 1860

gcgacttcgt agtgatcgac ggagcgcccc aggcggcgga cttggctgtg tccgcgatca 1920

aggcagccga cttcgtgctg attccggtgc agccaagccc ttacgacata tgggccaccg 1980

ccgacctggt ggagctggtt aagcagcgca ttgaggtcac ggatggaagg ctacaagcgg 2040

cctttgtcgt gtcgcgggcg atcaaaggca cgcgcatcgg cggtgaggtt gccgaggcgc 2100

tggccgggta cgagctgccc attcttgagt cccgtatcac gcagcgcgtg agctacccag 2160

gcactgccgc cgccggcaca accgttcttg aatcagaacc cgagggcgac gctgcccgcg 2220

aggtccaggc gctggccgct gaaattaaat caaaactcat ttgagttaat gaggtaaaga 2280

gaaaatgagc aaaagcacaa acacgctaag tgccggccgt ccgagcgcac gcagcagcaa 2340

ggctgcaacg ttggccagcc tggcagacac gccagccatg aagcgggtca actttcagtt 2400

gccggcggag gatcacacca agctgaagat gtacgcggta cgccaaggca agaccattac 2460

cgagctgcta tctgaataca tcgcgcagct accagagtaa atgagcaaat gaataaatga 2520

gtagatgaat tttagcggct aaaggaggcg gcatggaaaa tcaagaacaa ccaggcaccg 2580

acgccgtgga atgccccatg tgtggaggaa cgggcggttg gccaggcgta agcggctggg 2640

ttgtctgccg gccctgcaat ggcactggaa cccccaagcc cgaggaatcg gcgtgacggt 2700

cgcaaaccat ccggcccggt acaaatcggc gcggcgctgg gtgatgacct ggtggagaag 2760

ttgaaggccg cgcaggccgc ccagcggcaa cgcatcgagg cagaagcacg ccccggtgaa 2820

tcgtggcaag cggccgctga tcgaatccgc aaagaatccc ggcaaccgcc ggcagccggt 2880

gcgccgtcga ttaggaagcc gcccaagggc gacgagcaac cagatttttt cgttccgatg 2940

ctctatgacg tgggcacccg cgatagtcgc agcatcatgg acgtggccgt tttccgtctg 3000

tcgaagcgtg accgacgagc tggcgaggtg atccgctacg agcttccaga cgggcacgta 3060

gaggtttccg cagggccggc cggcatggcc agtgtgtggg attacgacct ggtactgatg 3120

gcggtttccc atctaaccga atccatgaac cgataccggg aagggaaggg agacaagccc 3180

ggccgcgtgt tccgtccaca cgttgcggac gtactcaagt tctgccggcg agccgatggc 3240

ggaaagcaga aagacgacct ggtagaaacc tgcattcggt taaacaccac gcacgttgcc 3300

atgcagcgta cgaagaaggc caagaacggc cgcctggtga cggtatccga gggtgaagcc 3360

ttgattagcc gctacaagat cgtaaagagc gaaaccgggc ggccggagta catcgagatc 3420

gagctagctg attggatgta ccgcgagatc acagaaggca agaacccgga cgtgctgacg 3480

gttcaccccg attacttttt gatcgatccc ggcatcggcc gttttctcta ccgcctggca 3540

cgccgcgccg caggcaaggc agaagccaga tggttgttca agacgatcta cgaacgcagt 3600

ggcagcgccg gagagttcaa gaagttctgt ttcaccgtgc gcaagctgat cgggtcaaat 3660

gacctgccgg agtacgattt gaaggaggag gcggggcagg ctggcccgat cctagtcatg 3720

cgctaccgca acctgatcga gggcgaagca tccgccggtt cctaatgtac ggagcagatg 3780

ctagggcaaa ttgccctagc aggggaaaaa ggtcgaaaag gtctctttcc tgtggatagc 3840

acgtacattg ggaacccaaa gccgtacatt gggaaccgga acccgtacat tgggaaccca 3900

aagccgtaca ttgggaaccg gtcacacatg taagtgactg atataaaaga gaaaaaaggc 3960

gatttttccg cctaaaactc tttaaaactt attaaaactc ttaaaacccg cctggcctgt 4020

gcataactgt ctggccagcg cacagccgaa gagctgcaaa aagcgcctac ccttcggtcg 4080

ctgcgctccc tacgccccgc cgcttcgcgt cggcctatcg cggccgctgg ccgctcaaaa 4140

atggctggcc tacggccagg caatctacca gggcgcggac aagccgcgcc gtcgccactc 4200

gaccgccggc gcccacatca aggcaccctg cctcgcgcgt ttcggtgatg acggtgaaaa 4260

cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag 4320

cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg cagccatgac 4380

ccagtcacgt agcgatagcg gagtgtatac tggcttaact atgcggcatc agagcagatt 4440

gtactgagag tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac 4500

cgcatcaggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg 4560

cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat 4620

aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc 4680

gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc 4740

tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga 4800

agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt 4860

ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg 4920

taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc 4980

gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg 5040

gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc 5100

ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg 5160

ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc 5220

gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct 5280

caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt 5340

taagggattt tggtcatgca ttctaggtac taaaacaatt catccagtaa aatataatat 5400

tttattttct cccaatcagg cttgatcccc agtaagtcaa aaaatagctc gacatactgt 5460

tcttccccga tatcctccct gatcgaccgg acgcagaagg caatgtcata ccacttgtcc 5520

gccctgccgc ttctcccaag atcaataaag ccacttactt tgccatcttt cacaaagatg 5580

ttgctgtctc ccaggtcgcc gtgggaaaag acaagttcct cttcgggctt ttccgtcttt 5640

aaaaaatcat acagctcgcg cggatcttta aatggagtgt cttcttccca gttttcgcaa 5700

tccacatcgg ccagatcgtt attcagtaag taatccaatt cggctaagcg gctgtctaag 5760

ctattcgtat agggacaatc cgatatgtcg atggagtgaa agagcctgat gcactccgca 5820

tacagctcga taatcttttc agggctttgt tcatcttcat actcttccga gcaaaggacg 5880

ccatcggcct cactcatgag cagattgctc cagccatcat gccgttcaaa gtgcaggacc 5940

tttggaacag gcagctttcc ttccagccat agcatcatgt ccttttcccg ttccacatca 6000

taggtggtcc ctttataccg gctgtccgtc atttttaaat ataggttttc attttctccc 6060

accagcttat ataccttagc aggagacatt ccttccgtat cttttacgca gcggtatttt 6120

tcgatcagtt ttttcaattc cggtgatatt ctcattttag ccatttatta tttccttcct 6180

cttttctaca gtatttaaag ataccccaag aagctaatta taacaagacg aactccaatt 6240

cactgttcct tgcattctaa aaccttaaat accagaaaac agctttttca aagttgtttt 6300

caaagttggc gtataacata gtatcgacgg agccgatttt gaaaccgcgg tgatcacagg 6360

cagcaacgct ctgtcatcgt tacaatcaac atgctaccct ccgcgagatc atccgtgttt 6420

caaacccggc agcttagttg ccgttcttcc gaatagcatc ggtaacatga gcaaagtctg 6480

ccgccttaca acggctctcc cgctgacgcc gtcccggact gatgggctgc ctgtatcgag 6540

tggtgatttt gtgccgagct gccggtcggg gagctgttgg ctggctggtg gcaggatata 6600

ttgtggtgta aacaaattga cgcttagaca acttaataac acattgcgga cgtttttaat 6660

gtactgaatt aacgccgaat taattcgggg gatctggatt ttagtactgg attttggttt 6720

taggaattag aaattttatt gatagaagta ttttacaaat acaaatacat actaagggtt 6780

tcttatatgc tcaacacatg agcgaaaccc tataggaacc ctaattccct tatctgggaa 6840

ctactcacac attattatgg agaaactcga gcttgtcgat cgacagatcc ggtcggcatc 6900

tactctattt ctttgccctc ggacgagtgc tggggcgtcg gtttccacta tcggcgagta 6960

cttctacaca gccatcggtc cagacggccg cgcttctgcg ggcgatttgt gtacgcccga 7020

cagtcccggc tccggatcgg acgattgcgt cgcatcgacc ctgcgcccaa gctgcatcat 7080

cgaaattgcc gtcaaccaag ctctgataga gttggtcaag accaatgcgg agcatatacg 7140

cccggagtcg tggcgatcct gcaagctccg gatgcctccg ctcgaagtag cgcgtctgct 7200

gctccataca agccaaccac ggcctccaga agaagatgtt ggcgacctcg tattgggaat 7260

ccccgaacat cgcctcgctc cagtcaatga ccgctgttat gcggccattg tccgtcagga 7320

cattgttgga gccgaaatcc gcgtgcacga ggtgccggac ttcggggcag tcctcggccc 7380

aaagcatcag ctcatcgaga gcctgcgcga cggacgcact gacggtgtcg tccatcacag 7440

tttgccagtg atacacatgg ggatcagcaa tcgcgcatat gaaatcacgc catgtagtgt 7500

attgaccgat tccttgcggt ccgaatgggc cgaacccgct cgtctggcta agatcggccg 7560

cagcgatcgc atccatagcc tccgcgaccg gttgtagaac agcgggcagt tcggtttcag 7620

gcaggtcttg caacgtgaca ccctgtgcac ggcgggagat gcaataggtc aggctctcgc 7680

taaactcccc aatgtcaagc acttccggaa tcgggagcgc ggccgatgca aagtgccgat 7740

aaacataacg atctttgtag aaaccatcgg cgcagctatt tacccgcagg acatatccac 7800

gccctcctac atcgaagctg aaagcacgag attcttcgcc ctccgagagc tgcatcaggt 7860

cggagacgct gtcgaacttt tcgatcagaa acttctcgac agacgtcgcg gtgagttcag 7920

gctttttcat atctcattgc cccccgggat ctgcgaaagc tcgagagaga tagatttgta 7980

gagagagact ggtgatttca gcgtgtcctc tccaaatgaa atgaacttcc ttatatagag 8040

gaaggtcttg cgaaggatag tgggattgtg cgtcatccct tacgtcagtg gagatatcac 8100

atcaatccac ttgctttgaa gacgtggttg gaacgtcttc tttttccacg atgctcctcg 8160

tgggtggggg tccatctttg ggaccactgt cggcagaggc atcttgaacg atagcctttc 8220

ctttatcgca atgatggcat ttgtaggtgc caccttcctt ttctactgtc cttttgatga 8280

agtgacagat agctgggcaa tggaatccga ggaggtttcc cgatattacc ctttgttgaa 8340

aagtctcaat agccctttgg tcttctgaga ctgtatcttt gatattcttg gagtagacga 8400

gagtgtcgtg ctccaccatg ttatcacatc aatccacttg ctttgaagac gtggttggaa 8460

cgtcttcttt ttccacgatg ctcctcgtgg gtgggggtcc atctttggga ccactgtcgg 8520

cagaggcatc ttgaacgata gcctttcctt tatcgcaatg atggcatttg taggtgccac 8580

cttccttttc tactgtcctt ttgatgaagt gacagatagc tgggcaatgg aatccgagga 8640

ggtttcccga tattaccctt tgttgaaaag tctcaatagc cctttggtct tctgagactg 8700

tatctttgat attcttggag tagacgagag tgtcgtgctc caccatgttg gcaagctgct 8760

ctagccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca 8820

cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct 8880

cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat 8940

tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg aattcgagct 9000

cggtacccgg ggatcctcta gagtcgacct gcaggcatgc a 9041

Claims (11)

1. A DNA molecule is a DNA molecule shown as a sequence 1 in a sequence table.

2. A recombinant vector, expression cassette or recombinant bacterium comprising the DNA molecule of claim 1.

3. The recombinant vector of claim 2, wherein: the recombinant vector is obtained by inserting the DNA molecule of claim 1 into a multiple cloning site or a recombination site of a plant expression vector.

4. A transgenic cell line of non-plant propagation material containing the DNA molecule of claim 1.

5. Use of the DNA molecule of claim 1 as a promoter.

6. Use of the DNA molecule of claim 1 for promoting expression of a gene of interest.

7. The use of claim 6, wherein: the target gene expression is the specific expression of the target gene.

8. The use of claim 7, wherein: the specific expression is root-specific expression.

9. A method for producing a transgenic plant, wherein the DNA molecule of claim 1 is used to promote the expression of a desired gene in a starting plant, thereby producing a transgenic plant expressing the desired gene.

10. The method of claim 9, wherein: the "expressing the gene of interest" is "specifically expressing the gene of interest in roots".

11. A method for producing a target protein by promoting expression of a gene encoding the target protein in a starting plant using the DNA molecule of claim 1.

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