CN109810182A - BnLAX1.c gene, albumen and its application in control cabbage type rape plant type - Google Patents
BnLAX1.c gene, albumen and its application in control cabbage type rape plant type Download PDFInfo
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- CN109810182A CN109810182A CN201910092178.6A CN201910092178A CN109810182A CN 109810182 A CN109810182 A CN 109810182A CN 201910092178 A CN201910092178 A CN 201910092178A CN 109810182 A CN109810182 A CN 109810182A
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Abstract
Include the present invention relates to BnLAX1.c gene, albumen and its application in control cabbage type rape plant type, present invention separation and using one kindBnLAX1.cThe DNA fragmentation of gene, the segment assign the ability that cabbage type rape stem number increases.Wherein, described to containBnLAX1.cThe nucleotide sequence of gene coding region is as shown in sequence table SEQ ID NO:1, and sequence length is 534 bp, and for the amino acid sequence of its coding protein as shown in SEQ ID NO:2, amino acid number is 177.The present invention is separated from cabbage type rapeBnLAX1.cGene, and identify its biological function in terms of cabbage type rape plant type improvement, it is had a very important significance for cultivating high yield cabbage type rape new varieties.
Description
Technical field
The present invention relates to cabbage type rape genetic engineering fields.It is obtained more particularly to separation, clone and by functional verification
A kind of cabbage type rape that can increase stem numberBnLAX1.cApplication of the gene in cabbage type rape plant type genetic improvement.
The method that the present invention uses RT-PCR is separated to control cabbage type rape stem number target geneBnLAX1.c, overexpressionBnLAX1.cGene can add cabbage type rape stem number, it was confirmed that the function and its application approach of the gene.
Background technique
Cabbage type rape (Brassica napusL.) it is the main rape cultivation type in China, there is high yield, resistance
By force, the features such as wide adaptability.In the Components of cabbage type rape cabbage type rape yield, primary effectively branch and its silique number
Contribution it is maximum.The stem of cabbage type rape is most important for the entire growth and development process of rape, and stem not only acts as branch
Support and translocation, and Wild cabbage type can be directly affected by influencing primary effectively branch amount, main inflorescence silique number etc. factors
The yield of rape；There is usually one stem, more stem characters can be improved the yield potential of cabbage type rape for cabbage type rape,
Therefore, it has become one of objective trait of cabbage type rape genetic improvement (Zhang Y, Li Q, Cui Y, et al.
Genetic characterization and fine mapping for multi-inflorescence in Brassica napusL. Theoretical and Applied Genetics, 2018,131 (11): 2311-2319).Pradhan etc.
Find primary branch number, Secondary branches number, single plant silique sum, silique density etc. to yield traits hybrid in mustard type rape
Advantages attributable maximum (Pradhan A K, Sodhi Y S, Mukhopadhyay A, et al. Heterosis
breeding in Indian mustard (Brassica juncea, L. Czern & Coss): Analysis of
component characters contributing to heterosis for yield. Euphytica, 1993, 69
(3): 219-229).It is one of the effective way for improving yield of rape using the good filial generation of hybrid vigour acquired character,
But it is time-consuming too long.The cabbage type rape that obtaining stem number using transgenic technology increased significantly has not been reported yet.
Summary of the invention
In order to overcome drawbacks described above, the present invention provides a kind of BnLAX1.c gene, albumen and its in control cabbage type rape
Application in plant type.The specifically expressed candidate gene in separate living tissue is selected to one according to RNA-seq result, applicant will
The unnamed gene isBnLAX1.c.The present invention separates and includes using one kindBnLAX1.cThe DNA fragmentation of gene, the segment assign
The ability that cabbage type rape stem number increases.Wherein, described to containBnLAX1.cThe nucleotide sequence of gene coding region is such as
Shown in sequence table SEQ ID NO:1, sequence length is 534 bp, the amino acid sequence of its coding protein such as SEQ ID NO:2
Shown, amino acid number is 177.
In order to achieve the above-mentioned object of the invention, the technical scheme adopted by the invention is as follows: a kind of cabbage type rape regulation of plant form
Albumen, which is characterized in that the albumen is for following (a) or (b):
(a) protein that the amino acid sequence shown in SEQ ID NO:2 forms；
(b) by the amino acid sequence of SEQ ID NO:2 by one or several amino acid residues substitution and/or missing and/or
Addition and the protein as derived from SEQ ID NO:2 relevant to cabbage type rape regulation of plant form.
It is a further object to provide the genes of coding foregoing proteins.
The gene is DNA molecular any in following (a1)-(a3)；
(a1) DNA molecular shown in SEQ ID NO:1；
(a2) hybridize under strict conditions with (a1) DNA sequence dna limited and encode cabbage type rape regulation of plant form GAP-associated protein GAP
DNA molecular；
(a3) at least have 70% with (a1) DNA sequence dna limited, at least have 75%, at least with 80%, at least with 85%, extremely
Less with 90%, at least with 95%, at least with 96%, at least with 97%, at least with 98% or at least have it is 99% homologous
Property and encode cabbage type rape regulation of plant form GAP-associated protein GAP DNA molecular.
It is a further object to provide the expression cassette containing forementioned gene, recombinant vector, recombinant microorganism or turn
Gene cell system.
A further object of the present invention is to provide the application of (b1) or (b2) or (b3) or (b4):
(b1) foregoing proteins, or, the gene, or, containing the expression cassette of the gene, recombinant vector, recombinant microorganism or turning
Gene cell system, the application in regulation cabbage type rape plant type；
(b2) albumen, or, the gene, or, containing the expression cassette of the gene, recombinant vector, recombinant microorganism or turning
Gene cell system is cultivating the application in cabbage type rape new varieties；
(b3) albumen, or, the gene, or, containing the expression cassette of the gene, recombinant vector, recombinant microorganism or turning
Gene cell system, the application in regulation cabbage type rape stem number；
(b4) albumen, or, the gene, or, containing the expression cassette of the gene, recombinant vector, recombinant microorganism or turning
Gene cell system, the application in regulation plant stem number.
The present invention also provides a kind of methods of genetically modified plants for cultivating increase plant stem number, to improve purpose plant
Described in albumen content or activity, obtain genetically modified plants；The plant stem number of the genetically modified plants is higher than the mesh
Plant.
A method of the genetically modified plants for increasing plant stem number are cultivated, cultivate to obtain transgenosis using preceding method
Plant；The Expressed in Transgenic Plant cabbage type rape regulation of plant form albumen, or contain forementioned gene.
Carry the present inventionBnLAX1.cThe expression vector of gene can be by using Ti-plasmids, plant viral vector, directly
The standard biologics technical method such as DNA conversion, microinjection, electroporated importing plant cell (Weissbach, 1998, Method
for Plant Molecular Biology VIII, Academy Press, New York, pp.411-463;
Geiserson and Corey, 1998, Plant Molecular Biology (2nd Edition)).
It includes of the invention for can be usedBnLAX1.cThe expression vector conversion host of gene is (including cabbage type rape
Various plants), cultivate the plant variety of plant type improvement.Plant host can also be rice, tobacco, soybean, tomato, wheat etc..
Gene of the present invention is higher in plant tip separate living tissue expression, therefore can be by gene of the invention and any point
The promoter of raw organizing specific expression is connected into suitable expression vector after combining, and converts plant host, increases mitogenetic group of plant
The quantity knitted, and then change the plant type of plant.
Include using the recycling of DNA QIAquick Gel Extraction KitBnLAX1.cThe DNA fragmentation of gene coding region, the side connected using digestion
This segment is connected into pMDC83 skeleton carrier by method, is constructed the Overexpression vector of the gene, is named as pMDC83-BnLAX1.c。
Using electric robin by pMDC83-BnLAX1.cIn vector introduction Agrobacterium tumefaciems, Agrobacterium tumefaciens strain is
GV3101.The genetic transforming method of mediation is infected by pMDC83- by AgrobacteriumBnLAX1.cConvert cabbage type rape receptor material
Expect J9712, has successfully been obtainedBnLAX1.cThe transgenic plant that gene expression amount is significantly improved relative to wild type, observation hair
It is existing, compared with WT lines, it is overexpressedBnLAX1.cTransgenic brassica napus stem number increase, explanationBnLAX1.c
Plant plant type can be regulated and controled.
In conclusion the present invention using cabbage type rape as research material, passes through analysis cabbage type rape variety " first 9712 "
Each histoorgan RNA-seq result, it was found that a specifically expressed candidate gene in separate living tissue, the gene may be
It plays an important role in the formation atomization of apical meristem, is by the unnamed geneBnLAX1.c。
Cabbage type rape is edible oil important source material, and improving its yield and improveing its quality is always the mesh that people make great efforts
Mark.In the present invention, it is overexpressedBnLAX1.cGene increases the stem number of cabbage type rape, explanationBnLAX1.cGene participates in
The regulation of plant form of cabbage type rape.Therefore, it is separated from cabbage type rapeBnLAX1.cGene, and identify it in Wild cabbage type oil
The biological function of dish plant type improvement aspect, has a very important significance for cultivating high yield cabbage type rape new varieties.
Detailed description of the invention
Sequence table SEQ ID NO:1 be the present invention separation clone includeBnLAX1.cThe nucleotides sequence of gene coding region
Column, sequence length are 534 bp, and 1-534 are its code areas, encode 177 amino acid；
Sequence table SEQ ID NO:2 isBnLAX1.cThe amino acid sequence of DNA encoding the protein；
Fig. 1 BnLAX1.cExpression of the gene in each histoorgan of cabbage type rape；
Fig. 2BnLAX1.cOverexpression vector constructs schematic diagram；
Fig. 3BnLAX1.cIn overexpressing plantsBnLAX1.cThe expression of gene, CK1 and CK2 are transgene negative seedling；
BnLAX1-2, BnLAX1-4, BnLAX1-10, BnLAX1-11, BnLAX1-12, BnLAX1-18 areBnLAX1.cTransgenic positive
Cabbage type rape；
Fig. 4BnLAX1.cThe plant type of overexpression transgenic brassica napus shows, and in figure: WT is Wild type control plants；BnLAX1.c- OE is to be overexpressedBnLAX1.cTransgenic brassica napus.
Specific embodiment
Following embodiment defines the present invention, and describes the present invention in clone and includeBnLAX1.cGene completely encodes
The DNA fragmentation of section, and verifyingBnLAX1.cThe method of gene function.According to being described below and these embodiments, this field
Technical staff can determine essential characteristic of the invention, and without departing from the spirit and scope of the invention, can be to this
Various changes and modifications are made in invention, so that it is suitable for different purposes and conditions.
It is endogenous that embodiment 1:RNA-seq analyzes cabbage type rapeBnLAX1.cExpression of the gene in each histoorgan
The histoorgan sample for taking cabbage type rape strain " first 9712 " each period, is immediately placed in quick-frozen in liquid nitrogen, and shifts
It is saved to 70 DEG C of refrigerators, until RNA is extracted.Total serum IgE extracting is mentioned using the RNAiso Plus kit of Dalian TaKaRa company
It takes, send to Science and Technology Ltd., Shanghai Major Biological Medical Technology Co., Ltd. and carry out RNA-Seq analysis, according to high-flux sequence interpretation of result gene
FPKM value, withBnubi(BnaA10g06670D) it is calculated as reference geneBnLAX1.cRelative expression in each tissue
Amount, as shown in Figure 1.
Embodiment 2: cabbage type rapeBnLAX1.cThe molecular cloning of gene
Take cabbage type rape variety " Darmor-bzh " three leaf one heart stage seedling, liquid nitrogen flash freezer, place saved in -70 DEG C of refrigerators with
It is standby to extract total serum IgE.Total serum IgE extracting is extracted using the RNAiso Plus kit of Dalian TaKaRa company.Cabbage type rape cDNA
Synthesis said by the HiScript 1st Strand cDNA Synthesis Kit of Nanjing Vazyme Biotechnology Co., Ltd.
Bright book operation carries out the first chain synthesis.
The first chain of cDNA with the synthesis of above-mentioned kit is amplification template, with the F:5'- of design
ACACAGACACCACGTAACAATAC-3'(SEQ ID NO:3) and R:5'-GGGGACACTAACAAACTAAGAT-3'(SEQ
ID NO:4) it is primer, cDNA amplification, amplification condition are carried out using RT-PCR are as follows: 94 DEG C of 3 min, 94 DEG C of 15 s, 59 DEG C
15 s, 72 DEG C of 30 s, totally 35 recycle；72℃ 10 min.Electrophoretic analysis is carried out after PCR, uses health for century biology
The DNA QIAquick Gel Extraction Kit of Science and Technology Ltd. recycles purpose amplified fragments.Amplified fragments are connected into Beijing Quan Shijin biotechnology
The pEASY-Blunt carrier T of Co., Ltd, converts competent escherichia coli cell, and picking white colony carries out bacterium colony PCR mirror
Determine positive colony, positive colony is sent to the sequencing of Yangzhou Qing Ke Biotechnology Co., Ltd, is ordered through the errorless plasmid of sequence verification
It is entitledBnLAX1.c-T。
Embodiment 3:BnLAX1.cThe building of gene overexpression carrier
In order to preferably analyzeBnLAX1.cThe function of gene, applicant's overexpression in cabbage type rape by it, passes through sight
The phenotype of transgenic plant is examined to study the function of the gene.
Overexpression vector construction method is as follows: with above-mentioned errorless through sequence verificationBnLAX1.cGene clone carrier matter
GrainBnLAX1.c- T is template, uses primer LAX1F(5'-gACTAGTcATGGACCAGTCCACTCTTCA-3') (SEQ ID
NO:5), the additional connector of sequence specific primersSpeI site) and LAX1 R(5'-atGGCGCGCCatAGACAAACCACGTCTAT
GCA-3') (SEQ ID NO:6), the additional connector of sequence specific primersAscI site) utilize RT-PCR progress cDNA amplification, amplification
Condition are as follows: 94 DEG C of 3 min, 94 DEG C of 15 s, 58 DEG C of 15 s, 72 DEG C of 30 s, totally 35 recycle；72℃ 10 min.PCR knot
Electrophoretic analysis is carried out after beam, and health is used to recycle purpose amplified fragments for the DNA QIAquick Gel Extraction Kit of century Biotechnology Co., Ltd.
Amplified fragments are connected in the pEASY-Blunt carrier T of Beijing Quanshijin Biotechnology Co., Ltd, Escherichia coli are converted
Competent cell, picking white colony carry out bacterium colony PCR to identify positive colony, positive colony are sent to Yangzhou and holds up biology section, section
The sequencing of skill Co., Ltd.
IncludeBnLAX1.cThe cloning vector plasmids of genetic fragment pass throughSpeI+AscAfter I double digestion, is recycled and tried using DNA
Agent box recycles target DNA fragment, this segment is connected with the pMDC83 skeleton carrier of corresponding digestion and is built intoBnLAX1.cGene
Overexpression vector, be named as pMDC83-BnLAX1.c(Fig. 2).
Embodiment 4:pMDC83-BnLAX1.cThe cabbage type rape genetic transformation of Overexpression vector
Using electroporated method by pMDC83-BnLAX1.cPlasmid imports in Agrobacterium tumefaciems GV3101 bacterial strain competent cell.
Picking single colonie is inoculated in overnight incubation in 25 mL YEB culture mediums (containing 50 mg/L rifampins), and 5 mL bacterium solutions is taken to be transferred to
In 100 mL YEB culture mediums (containing 50 mg/L rifampins), culture to OD600Bacterium solution is placed 10 by=0.7-0.8 on ice
Min, 5,000 4 DEG C of rpm are centrifuged 10 min and collect thallus, and the cleaning of 100 mL aseptic double-distilled waters is added twice.4 mL 10% are added
Glycerol suspension thalline is transferred in 50 mL centrifuge tubes.5500 4 DEG C of rpm are centrifuged 10 min and collect thallus, and 500 μ L are added
Thallus is resuspended in 10% glycerol, is transferred in 1.5 mL centrifuge tubes.
50 μ L competent cells are taken, 5 μ L pMDC83- are addedBnLAX1.cRecombinant plasmid is transferred to after being mixed with pipette tips
In 0.1 cm electrotransformation cup.Electrotransformation parameter: 500 μ L LB culture is added after electric shock immediately by 200 Ω, 1.7 KV, 2.5 F
Liquid.After 37 DEG C of 220 rpm cultivates 1 h, 100 μ L bacterium solutions is taken to be coated with the LA culture medium of Kanamycin resistance containing kanamycin
Screen transformant, 28 DEG C of 16 h of culture.
The genetic transforming method for converting cabbage type rape is to use for reference Hua Zhong Agriculture University's crop genetic improvement state key reality
Test room method for transformation improve after method utilize agriculture bar using the hypocotyl of cabbage type rape aseptic seedlings as explant
Bacterium mediated method realizes genetic transformation of the exogenous sequences in cabbage type rape.
Used medium formula is as follows:
Inoculation medium (M0): MURASHIGE & SKOOG MEDIUM(Duchefa Biochemie company)+30.0 g/L
Sucrose Sucrose+8 g/L agar Agar(pH 5.8-pH 6.0).
Co-culture medium (M1): M01.0 mg/L 2,4 dichlorophenoxyacetic acid of+18.0 g/L PEARLITOL 50C annitol+
Kinetin+100 μM of acetosyringone AS(pH 5.8 of 2,4-D+0.3 mg/L kinetin).
Callus differential medium (M2): M1+ 300.0 mg/L Ticarcillin/Clavulanate Acid Timentin+25 mg/L hygromycin
Hygromycin B。
Raw seedling culture medium (M3): MURASHIGE & SKOOG MEDIUM(Duchefa Biochemie company)+10.0
G/L glucose Glucose+0.25 g/L xylose Xylose+0.6 g/L morpholino b acid MES+2.0 mg/L zeatin
Zeatin+0.1 mg/L heteroauxin IAA+300.0 mg/L Ticarcillin/Clavulanate Acid Timentin+25 mg/L hygromycin
Hygromycin B。
Strengthening seedling and rooting culture medium (M4): M0+ 300.0 mg/L Ticarcillin/Clavulanate Acid Timentin.
MURASHIGE & SKOOG MEDIUM is referred to as MS culture medium.
Specific steps are as follows:
(1) it sterilizes:
A. 1 min of cabbage type rape seed is impregnated with 75% alcohol first, notices that the time cannot be too long；
B. then with 2% 20 min of hypochlorite disinfectant；
C. last to use aseptic water washing seed 4-5 times, it is cleaned up as far as possible.
(2) it sows:
A. sterilized seed is multicast to M with aseptic nipper0On culture medium, 30, every ware；
B. the culture tank of inoculation is placed into incubator, dark culture 6-7 d at 24 DEG C.
(3) bacterium is shaken:
After sowing 5-6 d, by Agrobacterium inoculation in sterile triangular flask or centrifuge tube containing LB liquid medium, it is placed in 28
It is cultivated in DEG C 180-220 rpm shaking table.
(4) it prepares explant and infects:
A. the seedling for after planting growing 6-7 d is cut with aseptic nipper and scalpel, it is 0.8- that its hypocotyl, which is cut into length,
The explant section of 1.0 cm.Its hypocotyl is placed in M when cutting seedling1It is cut in fluid nutrient medium, cuts better effect in this way.It cuts
When it is fast, quasi-, do not draw；
B. the OD of Agrobacterium is measured600Value (OD in LB culture medium600=0.3 or so preferably), will preparatory cultured bacterium solution with
6000 rpm are centrifuged 10 min, abandon supernatant, with the MS Liquid Culture containing 100 μM acetosyringone AS isometric with bacterium solution
Bacterium is resuspended base, repeats later primary.2 mL bacterium solutions finally are taken, the MS liquid of 100 μM of acetosyringone AS is contained with 20 mL
The dilution of body culture medium；
C. the explant cut is placed in the bacterium solution re-suspension liquid for having adjusted concentration, disseminates 10 min, pays attention to time of infection
Should not be too long, it otherwise will lead to explant death.It is appropriate that 150-200 explant is disseminated in the bacterium solution of every 20 mL；
(5) explant infected is transferred to M1On culture medium, it is advisable with 20-25 explant of every ware, is placed in 24 DEG C of half-lights
Lower culture 36-48 h；
(6) by explant from M1Go to M2On culture medium, and it is transferred to culture (24 DEG C of dark 8 h of 16 h/ of light) in illumination box
3 weeks；
(7) explant is transferred to M3On culture medium, every 2-3 weeks subculture is primary, until there is green bud；
(8) explant is finally transferred to M4It takes root in culture medium, rootage duration needs 2-4 weeks.
Embodiment 5: the identification of transgenic brassica napus positive plant
Cabbage type rape genomic DNA is extracted using express method, its step are as follows:
(1) take two panels young leaflet tablet (about 0.2 g), shred and be fitted into the centrifuge tube of 2 mL, be added 250 μ L DNA buffer and
Two steel balls (6.7 mm of diameter), proof press 50 Hz, 180 s smash leaf sample；
(2) sample smashed is placed in 95 DEG C of 10 min of incubation；
(3) sample taking-up is cooled to room temperature, 12000 rpm are centrifuged 5 min；
(4) 50 μ L supernatants are drawn to be transferred in 1.5 new mL centrifuge tubes, it is spare after 5 times of dilution.
DNA buffer formula:
Tris-HCl (pH=7.5) 500 mM
NaCl 300mM
300 mM of sucrose Sucrose
Take 1 μ L DNA as template, with primer 35S(5'-TCCCACTATCCTTCGCAAG-3') (SEQ ID NO:7) and R
(5'-atGGCGCGCCatAGACAAACCACGTCTATGCA-3') (SEQ ID NO:8), R(5'-
GACTAGTcATGGACCAGTCCACTCTTCA-3') (SEQ ID NO:9) and GFP(5'-TCCCACTATCCTTCGCAAG-3')
(SEQ ID NO:10) carries out PCR amplification, amplification condition are as follows: 94 DEG C of 5 min；94 DEG C of 30 s, 58 DEG C of 30 s, 72 DEG C 30
S, totally 30 recycle；72℃ 10 min.Using transgenic brassica napus DNA as template, specific target fragment can be amplified,
Prove purpose carrier 35S::BnLAX1.cIt has been integrated into cabbage type rape genome.
Embodiment 6: it is overexpressedBnLAX1.cGene render transgenic cabbage type rape stem number increases
The present invention is using the method for fluorescence detection real-time quantitative in partial transgenic cabbage type rape plantBnLAX1.cGene
Expression detected, embodiment 3 is shown in the extraction and reverse transcription of RNA.Fluorescence is carried out on 7500 quantitative PCR apparatus of ABI company
Quantitative PCR, with the qG1 F(5'-CATCTTCTTCTTTACACAGCCG-3' of design) (SEQ ID NO:11) and qG1 R(5'-
ACCAGATCTGAGCTTTCAAGAA-3') (SEQ ID NO:12) is that primer carries out quantitative fluorescent PCR, reaction condition are as follows: 95 DEG C
1 min, 95 DEG C of 15 s, 60 DEG C of 20 s, 72 DEG C of 31 s acquire fluorescence signal, totally 40 circulations；60 DEG C 95 DEG C of to, every 1
DEG C acquisition first order fluorescence signal, continue 1 s.After reaction, software (7500 Software carried with ABI7500
V2.0.1 it) is analyzed and is drawn.The results show that having successfully been obtainedBnLAX1.cThe expression quantity of gene is relative to wild typeBnLAX1.cThe transgenic plant (Fig. 3) that gene expression amount significantly improves.
To overexpressionBnLAX1.cTransgenic plant carry out phenotypic analysis, find compared with Wild type control plants, mistake
ExpressionBnLAX1.cTransgenic plant stem number increase (Fig. 4).
The isolated cabbage type rape gene relevant to plant type of the present invention is capable of specific aim acquisition regulation plant plant type
Candidate gene has certain theoretical direction effect with plant plant type related gene to research plant branching mechanism and separation.This
Isolated plant type related gene is invented from plant itself, is had fewer environmental impacts.Rape is carried out using isolated gene
Plant type improves molecular breeding, for cultivating the cabbage type rape new varieties of plant type improvement, improving cabbage type rape yield and harvest
Index has very important significance.
Basic principles and main features and advantages of the present invention of the invention have been shown and described above.The skill of the industry
Art personnel it should be appreciated that the present invention is not limited to the above embodiments, the above embodiments and description only describe
The principle of the present invention, without departing from the spirit and scope of the present invention, various changes and improvements may be made to the invention, these
Changes and improvements all fall within the protetion scope of the claimed invention.The claimed scope of the invention by appended claims and
Its equivalent thereof.
Sequence table
<110>Yangzhou University
<120>BnLAX1.c gene, albumen and its application in control cabbage type rape plant type
<130> xhx2019013001
<141> 2019-01-30
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atggaccagt ccactcttca tagcctaaac ccatattcat cttccaccac ttcctcatca 60
tcttcttctt tacacagccg caagggcaga ataaagggca acaaaaatca gtcaatgtcc 120
acgttatcga cggatccaca gagcgtggct gcccgtgaga gacgccaccg aatcagcgac 180
cgtttgaaga ttctacagag catggttcct ggtggtgcga agttggacac cgtctctatg 240
ctcgacgaag ccattagcta cgtcaagttc ttgaaagctc agatctggtt tcaccacaat 300
atgcttcttt tcttcaacga ctacgaaact acgtcgcctt gtacttattc cccagtcggc 360
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acgtattcac aagggacgcc actatatatg gttgctgacc cgaataatcc gatgtggtat 480
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35 40 45
Val Ala Ala Arg Glu Arg Arg His Arg Ile Ser Asp Arg Leu Lys Ile
50 55 60
Leu Gln Ser Met Val Pro Gly Gly Ala Lys Leu Asp Thr Val Ser Met
65 70 75 80
Leu Asp Glu Ala Ile Ser Tyr Val Lys Phe Leu Lys Ala Gln Ile Trp
85 90 95
Phe His His Asn Met Leu Leu Phe Phe Asn Asp Tyr Glu Thr Thr Ser
100 105 110
Pro Cys Thr Tyr Ser Pro Val Gly Val Ser Glu Phe Glu Ser Arg Leu
115 120 125
Phe Gly Cys Asp Glu Asp Tyr Thr Pro Val Pro Lys Thr Tyr Ser Gln
130 135 140
Gly Thr Pro Leu Tyr Met Val Ala Asp Pro Asn Asn Pro Met Trp Tyr
145 150 155 160
Ser Ser Val Asp Asp Glu Gln Gln Glu Thr Met His Arg Arg Gly Leu
165 170 175
Ser
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Claims (7)
1. a kind of cabbage type rape regulation of plant form albumen, which is characterized in that the albumen is for following (a) or (b):
(a) protein that the amino acid sequence shown in SEQ ID NO:2 forms；
(b) by the amino acid sequence of SEQ ID NO:2 by one or several amino acid residues substitution and/or missing and/or
Addition and the protein as derived from SEQ ID NO:2 relevant to cabbage type rape regulation of plant form.
2. encoding the gene of albumen described in claim 1.
3. gene according to claim 2, it is characterised in that: the gene is any in following (a1)-(a3)
DNA molecular；
(a1) DNA molecular shown in SEQ ID NO:1；
(a2) hybridize under strict conditions with (a1) DNA sequence dna limited and encode cabbage type rape regulation of plant form GAP-associated protein GAP
DNA molecular；
(a3) at least have 70% with (a1) DNA sequence dna limited, at least have 75%, at least with 80%, at least with 85%, extremely
Less with 90%, at least with 95%, at least with 96%, at least with 97%, at least with 98% or at least have it is 99% homologous
Property and encode cabbage type rape regulation of plant form GAP-associated protein GAP DNA molecular.
4. expression cassette, recombinant vector, recombinant microorganism or transgenic cell line containing gene described in Claims 2 or 3.
5.(b1) or the application of (b2) or (b3) or (b4):
(b1) albumen described in claim 1, or, gene described in Claims 2 or 3, or, containing gene described in Claims 2 or 3
Expression cassette, recombinant vector, recombinant microorganism or transgenic cell line, regulation cabbage type rape plant type in application；
(b2) albumen described in claim 1, or, gene described in Claims 2 or 3, or, containing gene described in Claims 2 or 3
Expression cassette, recombinant vector, recombinant microorganism or transgenic cell line, cultivate cabbage type rape new varieties in application；
(b3) albumen described in claim 1, or, gene described in Claims 2 or 3, or, containing gene described in Claims 2 or 3
Expression cassette, recombinant vector, recombinant microorganism or transgenic cell line, regulation cabbage type rape stem number in application；
(b4) albumen described in claim 1, or, gene described in Claims 2 or 3, or, containing gene described in Claims 2 or 3
Expression cassette, recombinant vector, recombinant microorganism or transgenic cell line, regulation plant stem number in application；Described
Plant is cabbage type rape, rice, tobacco, soybean, tomato or wheat.
6. a kind of cultivate the method for increasing the genetically modified plants of plant stem number, which is characterized in that improve in purpose plant
The content or activity of albumen described in claim 1, obtain genetically modified plants；The plant stem number of the genetically modified plants is higher than
The purpose plant.
7. a kind of cultivate the method for increasing the genetically modified plants of plant stem number, which is characterized in that using described in claim 6
Method cultivates to obtain genetically modified plants；Expressed in Transgenic Plant cabbage type rape regulation of plant form described in claim 1
Albumen, or contain gene described in claim 2 or 3.
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