CN109880832A - Rice leaf Leaf angle mutated gene PLA1 and its application - Google Patents

Abstract

The invention belongs to gene engineering technology fields, more particularly to rice leaf Leaf angle mutated gene PLA1 and its application, the nucleotide sequence of the mutated gene PLA1 is as shown in SEQ ID No.1, the amino acid sequence of its coding protein is as shown in SEQ ID No.2, compared with wild type, mutated gene PLA1 has become A from C in the 453rd bit base of the exon 2 of P450 gene (LOC_Os10g26340), so that the amino acid of coding becomes threonine by proline.Rice after the gene mutation shows as plant dwarfing, blade Leaf angle increases, and the gene can be used for rice variety selective, select the ornamental type rice with ornamental value.

Description

Rice leaf Leaf angle mutated gene PLA1 and its application

Technical field

The invention belongs to gene engineering technology fields, and in particular to rice leaf Leaf angle mutated gene PLA1 and its answer With.

Background technique

Rice is extremely important crops, most as per unit area yield highest, total output highest and consumer groups in China Cereal crops, the important in inhibiting in terms of ensureing China's grain security.Therefore, rice high yield is always breeders in product Most important target in kind breeding.Seed, fringe type and plant type are to influence three big factors of rice yield, are most heavy in genetic breeding The selection traits wanted.The plant type improvement of the rice rice strain significant, compact for rational close planting, the raising efficiency of light energy utilization Type is conducive to the formation of rice high yield.Rice leaf, which is stood upright, angle becomes smaller causes the reflectivity to light smaller, and it is double can to increase blade Face light improves the photosynthetic efficiency of canopy, so that dry-matter accumulation increases, while can also improve lower layer's illumination of group, Be conducive to the raising of root activity, and then achieve the purpose that promote rice yield.Therefore excavate and clone new adjusting and controlling rice leaf The gene of angle has important theory significance and directive significance for improving rice yield.Furthermore researcher is always continuous The unique ornamental value of excavation rice, power seek collect a batch the Different Variations such as leaf color, fringe type, plant height, plant type, seed water Rice.Leaf angle increases, the leaf development period cause in advance stalk raw blade increase and symmetric growth improves plant type of rice Uniqueness, the variation of rice leaf color also increases the ornamental value of rice, provides germplasm money abundant for cultivation " colored rice " Source.

Summary of the invention

It is an object of the present invention to provide rice leaf Leaf angle mutated gene PLA1 and applications.

It is yet a further object of the present invention to provide the primer pairs for detecting rice leaf Leaf angle mutated gene PLA1 And the kit containing the primer pair.

To achieve the goals above, the present invention is western big 1B using the holding of ethylmethane sulfonate (EMS) mutagenesis Indica Rice The rice leaf intersection angle for obtaining an inheritance stability increases mutant (being named as PLA1).Genetic analysis find the mutant character by Recessive genes (being named as PLA1) control.Molecular mapping is the results show that the gene is located at No. 10 chromosome RM25367 Between XJ-1 in the section of 86.7kb.

On the basis of said gene positioning, the present invention passes through candidate gene prediction, Homology search and gene order difference Compare, it is determined that the nucleotide sequence of PLA1 is as shown in SEQ ID No.1, the amino acid sequence of the protein of coding such as SEQ ID Shown in No.2.Compared with wild type, the 453rd of exon 2 in P450 gene (LOC_Os10g26340) of PLA1 gene Base has become A from C, so that the amino acid of coding becomes threonine by proline.Then, the present invention continues to the gene Expression pattern and hormone response are studied, and primarily determine that PLA1 gene may be passed by influencing the BR signals such as BU1 and ILI1 The development of approach middle and upper reaches gene regulation pulvinus adaxial and its surface cell is led to influence the size of rice leaf intersection angle.

The present invention also provides the primer pairs for detecting detection rice leaf Leaf angle mutated gene PLA1, including including just To primer: 5 '-CATGGCTCGCCCACCTCTACG-3 ', reverse primer (5 ' -3 '): 5 '-CTCCCAGAGGACGGCGACCAT- 3′。

The present invention also provides there are also the detection kits of above-mentioned primer pair.

The present invention also provides application of the rice leaf Leaf angle mutated gene PLA1 in rice breeding, using the gene Select the rice varieties of great ornamental value.

The invention has the benefit that the present invention provides rice leaf Leaf angle mutated gene PLA1, and pass through experiment Preliminary identification, the gene may be provided by the development of BR signal transduction path adjusting and controlling rice Leaf angle for improvement plant type of rice Direction.

Detailed description of the invention

Fig. 1 is the histological observation of wild type and mutant PLA1 Leaf angle；Wherein, A is seedling stage wild type and mutant The Leaf angle size statistical chart of PLA1 three pieces functional leaf；B is the leaf folder of tillering stage wild type and mutant PLA1 three pieces functional leaf Angle size statistical chart；C is the Leaf angle size statistical chart of maturity period wild type and mutant PLA1 three pieces functional leaf；D is maturation Phase wild type and mutant single plant tiller；E is that wild type and mutant fall a leaf Leaf angle；F is that wild type and mutant fall two Leaf Leaf angle；G is that wild type and mutant fall three leaf Leaf angles；H is that wild type falls two leaf pulvinus cells；I is that mutant falls two Leaf pulvinus cell；A is a regional enlarged drawing in figure H；B is b regional enlarged drawing in figure I；C is c regional enlarged drawing in figure H；D is figure I Middle d regional enlarged drawing；J is wild type adaxial and its surface and abaxial side cell length；K is that mutant adaxial and its surface and abaxial side cell are wide Degree；

Fig. 2 is the histological observation of wild type (WT) and mutant (PLA1) blade；Wherein, A is seedling stage wild type and dashes forward Variant blade；B is seedling stage wild type and mutant middle part of blade enlarged drawing；C is wild-type leaves sectional view；D is mutant leaf Piece sectional view；E is wild-type leaves middle arteries cell；F is mutant blade middle arteries cell；A is a regional enlarged drawing in figure C；B is Scheme b regional enlarged drawing in C；C is the enlarged drawing for scheming the region c in D；D is the enlarged drawing for scheming the region d in D；G is wild type and mutation Body great vascular bundle quantity statistics figure；H is wild type and mutant small bundle quantity statistics figure；I is that wild type and mutant are big Vascular bundle length and width statistical chart；J is wild type and mutant small bundle length and width statistical chart；

Fig. 3 is the histological observation of wild type and mutant stalk；Wherein, A is that the wild type in maturity period and mutant are planted Strain；B is the spike length of wild type and mutant, falls an internode to four internode length；C is the sectional view between wild type falls two successively；D Sectional view between falling two successively for mutant；E is the enlarged drawing of sectional view between wild type falls two successively；F is that mutant falls two sections The enlarged drawing of internode sectional view；G is longitudinal sectional figure between wild type falls two successively；H is longitudinal sectional figure between mutant falls two successively；I is open country Number of vascular bundles statistical chart in raw type and mutant stalk sectional view；J is cellular layer in wild type and the longitudinal sectional figure of mutant stalk Number statistical chart；K is cell length statistical chart in wild type and the longitudinal sectional figure of mutant；L is thin in wild type and the longitudinal sectional figure of mutant Born of the same parents' width statistical chart；

Fig. 4 is that the molecule of PLA1 gene positions figure；Wherein, the molecule that A is PLA1 positions；B is wild type and mutant Base variation；

Fig. 5 is expression pattern of the PLA1 gene in wild type；

Fig. 6 is response and analysis of related genes result of the mutant PLA1 to brassinosteroid BR；Wherein, A is different BR QPCR in wild type and mutant PLA1 under handling duration analyzes result；B is wild type and mutant Leaf angle to external source The sensitivity Detection result of BR；C is the Leaf angle result of variations of wild type and mutant PLA1 external source BR before and after the processing；D is BR The expression analysis result of biological signal conduction and synthesis related gene.

Specific embodiment

The present invention is clearly and completely described below by specific embodiment, it is clear that described embodiment is only It is a part of the embodiments of the present invention, instead of all the embodiments.Based on the embodiment of the present invention, ordinary skill people Member's every other embodiment obtained, belongs to protection scope of the present invention.

Test method without specific conditions in embodiment, usually according to normal condition, such as Molecular Cloning: A Laboratory refers to Condition described in south (third edition, J. Pehanorm Brooker etc. write, and Huang Peitang etc. is translated, Science Press, 2002), or according to system Make condition proposed by manufacturer.

Material used in embodiment: the big 1B in wild type west and rice leaf Leaf angle increase mutant PLA1 by southwestern big Rice research is learned to be provided；M-MLV reverse transcriptase, high-fidelity DNA polymerase PFU, T4 DNA ligase, Trizol kit, DNA gel QIAquick Gel Extraction Kit, plasmid extraction kit are purchased from TaKaRa company；Ampicillin (Ampicillin, Amp) is Sigma Products；Primer synthesis and DNA sequencing are completed by the handsome Bioisystech Co., Ltd in Shanghai；Other chemical reagent purchases From Beijing Ding Guo biotechnology Co., Ltd；Bacillus coli DH 5 alpha is provided by Southwest University's rice research.

Embodiment 1

Southwest University's rice research keep the system big 1B in west to obtain an inheritance stability using EMS mutagenesis Indica Rice Rice leaf intersection angle increases mutant (being named as PLA1), shows as plant dwarfing, blade Leaf angle increases.It has been observed that plant Dwarfing is to see Fig. 3 B since PLA1 spike length, the section of falling 1-4 internode length shorten, and reach the level of signifiance, and " * " is indicated in 0.05 water Flat upper significant difference, " * * " are indicated in 0.01 horizontal upper significant difference；Counted respectively under the conditions of field planting seedling stage, tillering stage, The Leaf angle of maturity period wild type and mutant finds that the Leaf angle of three pieces functional leaf is greater than wild type in each period, sees Figure 1A- C, wherein 1st indicates that a leaf Leaf angle, 2nd indicate that two leaf Leaf angles, 3rd indicate three leaf Leaf angles.In addition to this it dashes forward Variant is also embodied by narrow leaf and stalk attenuates.

In order to further analyze mutant Leaf angle increase, blade narrows, stalk attenuates the reason of, using paraffin section into The eucaryotic cell structure discovery mutant adaxial and its surface cell of row cytological observation, observation pulvinus position is elongated relative to wild type, reaches Extremely significant level, the extremely significant shorter than wild type of mutant adaxial and its surface cell width；The cell length and width difference of abaxial side are equal It is not significant.That is mutant Leaf angle increase is caused since adaxial and its surface cell is elongated, sees Fig. 1 D-K, wherein ad indicates adaxial and its surface, Ab indicates abaxial side；The blade that seedling stage starts mutant PLA1 seriously narrows compared with wild type, and this character is continued for To the maturity period, paraffin section is shown in Fig. 2 the results show that it is that number of vascular bundles tails off and causes that PLA1 blade, which narrows,；Count wild type and The crosscutting vascular bundle number of mutant PLA1 stalk, cell length and width find that mutant PLA1 vascular bundle number is extremely significant and subtract It is few, and cell length and width be without significant changes, and under identical amplification factor, the crosscutting structure chart of mutant PLA1 stalk is less than Wild type, considerably less than wild type, i.e. stalk becomes the cellular layer number of the longitudinal sectional cell number of plies discovery mutant PLA1 of statistics stalk It is carefully mainly influenced by cell division, sees Fig. 3.

Positioning, sequencing and the functional verification of 2 rice leaf Leaf angle of embodiment increase mutated gene PLA1

1, the positioning of rice leaf Leaf angle mutated gene PLA1

Hybridized with the normal red silk of phenotype extensive No. 10 with mutant, F1 generation phenotype is normal, illustrates the mutant by recessive gene Control.F2 shows parents' character for occurring apparent separation in group respectively.Through Chi-square test, normal Zhu ﹕ mutant strain meets 3: 1 segregation ratio shows that mutant is controlled by Recessive genes (being named as PLA1), carries out the assignment of genes gene mapping using F2 recessiveness group, altogether Obtain 947 mutant strains.The DNA in parental gene pond and mutated gene pond is extracted according to CTAB method；

(1) it is attached separately to after shredding 10 plants of parent's rice leafs and Mutant Rice blade quick-frozen in the centrifuge tube of 2ml In liquid nitrogen, it is broken into powder using tissue grinder, 65 DEG C of warm bath after the CTAB liquid of 60 DEG C of preheatings of 500 μ l are added 30min, period 10min or so shake up once；

(2) 500 μ l chloroforms are added, mixing of turning upside down is placed at room temperature for 5000rpm centrifugation 5min after 2min, by supernatant It is drawn in the centrifuge tube of new 1.5ml, the dehydrated alcohol after twice of supernatant volume frost is added gently shakes up until cotton-shaped Precipitating is united；

(3) 12000rpm outwells supernatant after being centrifuged 2min；

(4) 600 μ l dehydrated alcohols washed once, and outwell supernatant after 12000rpm centrifugation 2min；

(5) ddH of 200 μ l is added after air-drying to no liquid residual₂O, -20 DEG C save backup.

Select 400 pairs of SSR markers being uniformly distributed on 12 chromosome filter out extensive No. 10 and mutant PLA1 of red silk it Between polymorphism mark, mutated gene pond and normal gene pond are analyzed using polymorphism mark (referring to master's degree opinion Text: the genetic analysis and the assignment of genes gene mapping of rice leaf margin albino mutant mal, 2014, page 22), as a result on No. 10 chromosome XYD-3 and XYD-5 and PLA1 performance it is chain.The further design primer between two labels, primer sequence is auspicious to be shown in Table 1, finally will PLA1 is located between RM25367 and XJ-1, and physical distance 246kb is shown in Fig. 4 A.

1 gene finely positioning the primer of table and its sequence

2, the sequencing of rice leaf Leaf angle mutated gene PLA1

On the basis of to PLA1 finely positioning, by candidate gene on-line prediction (http: // ) and Gramene database (http://ensembl.gramene.org/genome- mendel.cs.rhul.ac.uk Browser/index.html the gene order) provided downloads the genomic dna sequence of candidate gene, is compared online using BLAST To and VectorNTI10 software design sequencing primer: forward primer sequence (5 ' -3 ') are as follows: 5 ' - CATGGCTCGCCCACCTCTACG-3 ', reverse primer sequences (5 ' -3 ') are as follows: 5 '-CTCCCAGAGGACGGCGACCAT-3 '.Point Not Yong wild type and mutant DNA profiling expand target fragment, amplification system are as follows: SYBRPremix Ex Taq II 10 μ l, PCR 0.6 μ l, PCR Reverse Primer of Forward Primer, 0.6 μ l, cDNA template, 2 μ l, RNase Free dd H2O 6.8 μ l, 20 μ l of overall reaction system.The single purpose band blob of viscose expanded is cut and recycles purpose using plastic recovery kit Then 16 DEG C of segment are connected on 19-T carrier by segment overnight, convert bacillus coli DH 5 alpha, select after 37 DEG C are cultivated one day Single positive clone, send sequencing company to be sequenced, sequencing result is compared difference on VectorNTI10 software.As the result is shown The nucleotide sequence of PLA1 is as shown in SEQ ID No.1, and the amino acid sequence of coding protein is as shown in SEQ ID No.2. Compared with wild type, PLA1 gene is become in the 453rd bit base of the exon 2 of P450 gene (LOC_Os10g26340) by C In order to which A is shown in Fig. 4 B so that the amino acid of coding becomes threonine by proline.

The expression pattern of 3 rice leaf Leaf angle of embodiment increase mutated gene PLA1

To understand the expression pattern that rice leaf Leaf angle increases mutated gene, by real time fluorescent quantitative qPCR to wild Type carries out expression analysis discovery, (referring to Ph.D. Dissertation: the map based cloning and function of rice Spikelet development related gene MFS1 Can analysis, 2013, page 29) rice leaf Leaf angle increases mutated gene in each tissue site has expression, and in blade Middle expression highest, followed by root and sheath, expression quantity is minimum in stem.See Fig. 5, wherein Ro represents root, and St represents stem, and Le is represented Leaf, Sh represent sheath, and Sp-1 represents length as the fringe of 0-2cm, and Sp-2 represents length as the fringe of 2-5cm.

The hormone of 4 rice leaf Leaf angle mutated gene PLA1 of embodiment responds experimental analysis

(1) by mutant PLA1 and wild type seeds vernalization, selection shows money or valuables one carries unintentionally consistent seed sowing in Nutrition Soil, places It is cultivated in constant incubator, with 10 μm of ol L when mutant PLA1 and wild type length to three one heart stage of leaf^–1BR (rape element Lactone) wild type and mutant PLA1 are handled respectively, and handling duration is 0h, 0.5h, 1h, 2h, 4h, 6h.Furthermore with difference The 24-eBL processing wild type and mutant PLA1 of concentration, the results showed that with the increase of external source 24-eBL concentration, wild type and The Leaf angle of mutant PLA1 constantly increases, when concentration is increased to 0.01 μm of ol L^–1When wild type Leaf angle reach maximum, with Concentration increase Leaf angle and tend towards stability, and mutant Leaf angle constantly increases, 10 μm of ol L^–1When Leaf angle reach maximum value and see Fig. 6 B；The variable quantity of Leaf angle is increased to 56.5 ° of mutant PLA1 (△ S) by 7 ° of wild type (△ B), is risen 87.6%, reach extremely significant level of difference and sees Fig. 6 C.It takes wild type and mutant PLA1 under each processing to extract RNA, and inverts CDNA is recorded into, is saved backup.

(2) quantitative PCR analysis

Take 10 μm of ol L^–1The different wild type and mutant PLA1 of BR (brassinosteroid) handling duration aerial part In in the centrifuge tube of the nuclease free of 2.0ml, it is placed in liquid nitrogen at once.It is (total referring to Tiangeng plant that RNA is extracted according to the following steps RNA extracts kit specification):

The quick-frozen centrifuge tube after liquid nitrogen is placed in tissue grinder 45HZ by a, and 60s is smashed completely；

RNA lysate is added in b, and the dilution of addition equivalent after 3-5min is placed at room temperature for after being mixed with liquid-transfering gun；

Room temperature 12000rpm is centrifuged 5min, careful Aspirate supernatant after c Tissue Lysis mixes；

The dehydrated alcohol piping and druming that 0.5 times of supernatant volume is added in d is fully transferred in centrifugal column after mixing, 12000rpm from Supernatant is outwelled after heart 1min；

600 μ lRNA washing lotions are added in e, abandon filtrate after 12000rpm centrifugation 45s, and 50 μ lDNA enzymes I are added, and (10 ╳ DNA enzymatics I are slow 5 μ l of fliud flushing, DNA enzymatic I 5 μ l, 40 μ l of nuclease free Shui) to adsorbed film center, it is placed at room temperature for 15min；

F is added 600 μ lRNA washing lotion 12000rpm and is centrifuged 45s, supernatant is outwelled, by centrifuge tube weight after repetitive operation is primary For new placement on collecting pipe, 12000rpm is centrifuged 2min；

Centrifugal column is transferred on elution pipe by g, and 50-200 μ l free nucleic acid Shui is added in film center, is stored at room temperature 2min, RNA can be obtained in 12000rpm centrifugation 1min.

According to following system reverse transcription at cDNA (sending out transcript reagent box specification referring to TaKaRa) after RNA extraction: 5 ╳ 2 μ l, gDNA Ereasar of gDNA Ereasar Buffer, 1 μ l, RNA 7,4 DEG C of preservation 1min after μ l, 42 DEG C of reaction 2min；So I 1 μ l, RT Primer Mix of Primefxcript Enzyme Mix 1 μ l, 5 ╳ is added in above-mentioned 10 μ l system afterwards 4 μ l, RNase Free ddH of Primescript Buffer₂4 DEG C of preservation 1min after O 4 μ l, 37 DEG C of reactions 15min, 85 DEG C of 5s Products therefrom is cDNA product.

The expression quantity in PCR reaction system analysis wild type and mutant PLA1 is prepared according to following components: II 10 μ l, PCR Forward Primer of SYBRPremix Ex Taq, 0.6 μ l, PCR Reverse Primer, 0.6 μ l, 2 μ l, RNase Free dd H of cDNA template₂6.8 μ l of O, 20 μ l of overall reaction system, amplification are quantitative in Real-Time System It is carried out in PCR instrument.Wild type expression quantity sharply increases when length reaches 1h when treated as the result is shown, when rise is about 0h 120 times, subsequent expression quantity is die-offed, and is tended towards stability with suitable and later period when 0h, is seen Fig. 6 A；It takes each concentration simultaneously treated and is wild Type and mutant PLA1, extract RNA, and reverse transcription carries out qPCR analysis at cDNA, the results showed that, compared with wild type, BR signal Expression quantity of pathway gene BRI1, BZR1, BU1, ILI1, the TUD1 in mutant PLA1 raises, and wherein ILI1 is prominent Expression quantity in variant PLA1 is 76.5 times of wild type, and 8 times；But BR route of synthesis controlling gene D4, D2 and D11 expression quantity Likelihood signal pathway gene variation is not obvious for variation, sees Fig. 6 D, related quantification PCR primer sequence is shown in Table 2.Therefore it primarily determines Rice leaf Leaf angle increases mutated gene PLA1 may be by influencing the BR signal transduction path middle and upper reaches genes such as BU1 and ILI1 The development of regulation pulvinus adaxial and its surface cell is to influence the size of rice leaf intersection angle.

2 quantification PCR primer sequence of table

The foregoing is merely illustrative of the preferred embodiments of the present invention, is not intended to limit the invention, all in essence of the invention Within mind and principle, all any modification, equivalent substitution, improvement and etc. be should all be included in the protection scope of the present invention.

Sequence table

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His Pro Ala Leu Leu Ala Gly Ala Val Leu Leu Ala Phe Ala Thr Ala

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Ala Val Leu Ala Trp Ala Ala Ser Pro Gly Gly Pro Ala Trp Ala His

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Gly Arg Gly Arg Leu Gly Ala Thr Pro Ile Glu Gly Pro Arg Gly Leu

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Pro Val Phe Gly Ser Ile Phe Ala Leu Ser Arg Gly Leu Pro His Arg

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Ala Leu Asp Ala Met Ser Arg Asp Ala Ala Ala Pro Arg Ala Arg Glu

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Leu Met Ala Phe Ser Val Gly Glu Thr Pro Ala Val Val Ser Ser Cys

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Pro Ala Thr Ala Arg Glu Val Leu Ala His Pro Ser Phe Ala Asp Arg

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Pro Leu Lys Arg Ser Ala Arg Glu Leu Leu Phe Ala Arg Ala Ile Gly

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Gln Ala Asp Ala Thr Ala Met Leu Ser Ala Met Ala Ala Glu Gln Ser

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Ala Thr Gly Ala Val Val Leu Arg Pro His Leu Gln Ala Ala Ala Leu

210 215 220

Asn Asn Ile Met Gly Ser Val Phe Gly Arg Arg Tyr Asp Val Ser Ser

225 230 235 240

Ser Ser Gly Ala Ala Ala Asp Glu Ala Glu Gln Leu Lys Ser Met Val

245 250 255

Arg Glu Gly Phe Glu Leu Leu Gly Ala Phe Asn Trp Ser Asp His Leu

260 265 270

Pro Trp Leu Ala His Leu Tyr Asp Pro Asn His Val Ala Arg Arg Cys

275 280 285

Ala Ala Leu Val Pro Arg Val Gln Ala Phe Val Arg Gly Val Ile Arg

290 295 300

Asp His Arg Leu Arg Arg Asp Ser Ser Ser Thr Ala Ala Asp Asn Ala

305 310 315 320

Asp Phe Val Asp Val Leu Leu Ser Leu Glu Ala His Glu Asn Leu Ala

325 330 335

Glu Asp Asp Met Val Ala Val Leu Trp Glu Met Ile Phe Arg Gly Thr

340 345 350

Asp Thr Thr Ala Leu Val Thr Glu Trp Cys Met Ala Glu Val Val Arg

355 360 365

Asn Pro Ala Val Gln Ala Arg Leu Arg Ala Glu Val Asp Ala Ala Val

370 375 380

Gly Gly Asp Gly Cys Pro Ser Asp Gly Asp Val Ala Arg Met Pro Tyr

385 390 395 400

Leu Gln Ala Val Val Lys Glu Thr Leu Arg Ala His Pro Pro Gly Pro

405 410 415

Leu Leu Ser Trp Ala Arg Leu Ala Thr Ala Asp Val Gly Leu Ala Asn

420 425 430

Gly Met Val Val Pro Ala Gly Thr Thr Ala Met Val Asn Met Trp Ala

435 440 445

Ile Thr His Asp Gly Glu Val Trp Ala Asp Pro Glu Ala Phe Ala Pro

450 455 460

Glu Arg Phe Ile Pro Ser Glu Gly Gly Ala Asp Val Asp Val Arg Gly

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Gly Asp Leu Arg Leu Ala Pro Phe Gly Ala Gly Arg Arg Val Cys Pro

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Gly Lys Asn Leu Gly Leu Ala Thr Val Thr Leu Trp Val Ala Arg Leu

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Val His Ala Phe Asp Trp Phe Leu Pro Asp Gly Ser Pro Pro Val Ser

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Leu Asp Glu Val Leu Lys Leu Ser Leu Glu Met Lys Thr Pro Leu Ala

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Ala Ala Ala Thr Pro Arg Arg Arg Arg Ala Ala

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<213>artificial sequence (Artificial Sequence)

<400> 16

ttacatccct tgcggtaggt 20

<210> 17

<211> 22

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 17

gtagccagct tgatctcatc tc 22

<210> 18

<211> 20

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 18

gggacgactc tactgcatca 20

<210> 19

<211> 20

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 19

gttcgaagtc tcctgcttcc 20

<210> 20

<211> 20

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 20

tgttcccaac agattcctca 20

<210> 21

<211> 20

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 21

ccggtctagg ctctgttctc 20

<210> 22

<211> 20

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 22

acacttgctc cttcagcctt 20

<210> 23

<211> 22

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 23

ttctctccaa gcttcaggcc ct 22

<210> 24

<211> 22

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 24

tgcacgtctc ctgcaaaacc ct 22

<210> 25

<211> 20

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 25

gacccaactc tggtcaatcc 20

<210> 26

<211> 20

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 26

caagttgtgg gaccaaagtg 20

<210> 27

<211> 20

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 27

cacaagagaa tgcctccaga 20

<210> 28

<211> 20

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 28

attgggctct cgtagctcat 20

<210> 29

<211> 20

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 29

cttgcagcct tcctctctct 20

<210> 30

<211> 20

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 30

gcgtggagag aaactgaaca 20

<210> 31

<211> 20

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 31

tatgtgccaa caaaggagga 20

<210> 32

<211> 20

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 32

cctctggcct cctacatcat 20

<210> 33

<211> 20

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 33

caaatcctgg acctcttgct 20

<210> 34

<211> 20

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 34

tcctccctca gttcttggac 20

<210> 35

<211> 20

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 35

gaggtggaag gagaaggaca 20

<210> 36

<211> 20

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 36

ctggtgacca agtggtgaag 20

<210> 37

<211> 20

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 37

tcacctccac aaagctcaag 20

<210> 38

<211> 20

<212> DNA

<213>artificial sequence (Artificial Sequence)

<400> 38

gcaaactttc ttgcctcctc 20

Claims (6)

1. rice leaf Leaf angle mutated gene PLA1, which is characterized in that the nucleotide sequence such as SEQ of the mutated gene PLA1 Shown in ID No.1.

2. the protein of rice leaf Leaf angle mutated gene PLA1 coding described in claim 1, which is characterized in that the egg The amino acid sequence of white matter is as shown in SEQ ID No.2.

3. the primer pair for detecting rice leaf Leaf angle mutated gene PLA1 described in claim 1, which is characterized in that including Forward primer: 5 '-CATGGCTCGCCCACCTCTACG-3 ', reverse primer (5 ' -3 '): 5 ' - CTCCCAGAGGACGGCGACCAT-3′。

4. including the detection kit of primer pair described in claim 3.

5. application of the rice leaf Leaf angle mutated gene PLA1 described in claim 1 in rice variety selective.

6. weighing application of the rice leaf Leaf angle mutated gene PLA1 in ornamental type rice variety selective described in requiring 1.