CN111235163B - Rice meiosis development related gene OsMFS1 and application thereof - Google Patents

Abstract

The invention discloses a rice meiosis development related gene OsMFS1 and application thereof, wherein the DNA sequence of the gene OsMFS1 is shown as SEQ ID No. 1. The invention obtains the gene MFS1 for controlling the meiosis development of rice by researching the male sterile mutant MFS. The discovery can provide an authoritative theoretical basis, is beneficial to accelerating germplasm innovation, and is beneficial to cultivating high-quality rice varieties with high yield, strong stress resistance and wide adaptability, thereby having important application value in plant breeding.

Description

Rice meiosis development related gene OsMFS1 and application thereof

Technical Field

The invention belongs to the technical field of plant genetic engineering, and relates to a rice meiosis development related protein OsMFS1 and application of a coding gene thereof.

Technical Field

Rice fertility is a key factor affecting rice yield, and in flowering plants, the reproductive development of male and female gametes isImportant biological processes are also growth processes commonly experienced by higher plants. In the study of reproductive development in plants, meiotic development, a complex process of cell division that creates haploid gametes, is a sexually reproducing organism, is of great interest. During division of germ cells, chromosomes are replicated once, and cells are divided twice continuously, which is a special division mode with the number of chromosomes halved^[1]. Meiosis is not only a mechanism for ensuring the chromosome number of a species to be stable, but also a mechanism for the species to evolve constantly in response to environmental changes. The meiotic mechanism is widespread in animals, plants and fungi, although there are also significant differences between different species^[2,3]. With the development of molecular biology in recent years, a plurality of meiosis development related genes are cloned in animals and plants such as rice, arabidopsis thaliana, mice and the like, so that the functions of the meiosis genes and the relationship between the meiosis genes and male and female gametes are more clearly known.

For animals and plants, the successful completion of meiosis to form functional male and female gametophytes is the key to ensure the genetic stability and diversity of species. Meiosis is a complex process involving homologous chromosome pairing, association and segregation, with a large number of meiotic proteins recruited to ensure that this process is successfully completed, any "errors" that are fatal to gametogenesis, and therefore the meiotic process carries numerous innovations for the species and is at high risk. Homologous chromosome association forms a special device association complex for completing homologous recombination, and ensures that DSB (double-strand break) of homologous recombination is successfully repaired^[4,5]. The gene MND1 in Arabidopsis thaliana and another meiosis peculiar protein HOP2 form a heterodimer and play a role as a cofactor in the homologous recombination process^[6,7]In the method, the SPO11 protein cuts double chains to form DSB, then restriction enzymes cut the cut to expose 3' single chains, and a large amount of recombinant proteins RAD51 and DMC1 are recruited to complete the repair work of the DSB^[8,9,10]The MND1/HOP2 complex participates in chain invasion reaction, and the main functions are to search a template double chain and activate the repair capacity of RAD51 and DMC1 to ensure that DSB is successfully repaired^[11]. In recent years, MFS1 has been studied more and clearly, and the mutation can cause complete male and female sterility, playing a key role in homologous recombination process^[12]. A series of researches show the importance of the meiotic homologous recombination process, and seem to reflect that the meiotic homologous recombination process is controllable, the genes can be modified, the recombination efficiency can be improved by applying biotechnology, good genes and good characters can be obtained more quickly in the breeding process, but more researches are still needed to comprehensively understand the homologous recombination process.

Reference to the literature

[1]Wang,C.,Wang,Y.,Cheng,Z.,Zhao,Z.,and Chen,J.,et al.2015.The role of OsMSH4in male and female gamete development in rice meiosis.Journal of Experimental Botany[J],67,1447-1459.

[2]Schwacha,A.,and Kleckner,N.1997.Interhomolog bias during meiotic recombination:Meiotic functions promote a highly differentiated interhomolog-only pathway.Cell[J],90,1123-1135.

[3]Wang,Y.,and Copenhaver,G.P.2018.Meiotic recombination:Mixing it up in plants.Annual Review of Plant Biology[J],69.577-609.

[4]Masson,J.Y.,Davies,A.A.,Hajibagheri,N.,Dyck,E.V.,and Benson,F.E.,et al.1999.The meiosis-specific recombinase hDmc1forms ring structures and interacts with hRad51.Embo Journal[J],18,6552-6560.

[5]Schwacha,A.,and Kleckner,N.1997.Interhomolog bias during meiotic recombination:Meiotic functions promote a highly differentiated interhomolog-only pathway.Cell[J],90,1123-1135.

[6]Patrick,S.,Lumir,K.,Stephen,V.K.,and Sehorn,M.G.2003.Rad51recombinase and recombination mediators.Journal of Biological Chemistry[J],278,42729-42732.

[7]Yi-Kai,C.,Chih-Hsiang,L.,Heidi,O.,Ming-Hui,L.,and Yuan-Chih,C.,et al.2004.Heterodimeric complexes of Hop2and Mnd1function with Dmc1to promote meiotic homolog juxtaposition and strand assimilation.Proceedings of the National Academy of Sciences of the United States of America[J],101,10572-10577.

[8]Shehre-Banoo,M.,Ramesh,M.A.,Hulstrand,A.M.,and Logsdon,J.M.2007.Protist homologs of the meiotic Spo11gene and topoisomerase VI reveal an evolutionary history of gene duplication and lineage-specific loss.Molecular Biology&Evolution[J],24,2827-2841.

[9]Farah,J.A.,Gareth,C.,Steiner,W.W.,and Smith,G.R.2005.A novel recombination pathway initiated by the Mre11/Rad50/Nbs1complex eliminates palindromes during meiosis in Schizosaccharomyces pombe.Genetics[J],169,1261-1274.

[10]Hong,E.L.,Shinohara,A.,and Bishop,D.K.2001.Saccharomyces cerevisiae Dmc1protein promotes renaturation of single-strand DNA(ssDNA)and assimilation of ssDNA into homologous super-coiled duplex DNA.Journal of Biological Chemistry[J],276,41906-41912.[11]Petukhova,G.V.,Romanienko,P.J.,and Camerini-Otero,R.D.2003.The hop2protein has a direct role in promoting interhomolog interactions during mouse meiosis.Developmental Cell[J],5,927-936.

[12]Clemens,U.,Arnaud,R.,Mona,V.H.,Arnaud,D.M.,and Daniel,V.,et al.2013.Sufficient amounts of functional HOP2/MND1complex promote interhomolog DNA repair but are dispensable for intersister DNA repair during meiosis in Arabidopsis.Plant Cell[J],25,4924-4940.

Disclosure of Invention

It is an object of the present invention to provide a meiotic helix-helix protein OsMFS 1.

The second purpose of the invention is to provide a rice meiosis development key gene OsMFS 1.

The invention also aims to provide application of a meiosis development key gene OsMFS 1.

The purpose of the invention is realized by the following technical scheme:

the invention provides a rice meiosis development key protein OsMFS1, which is an accessory protein OsMFS1 in a DSB repair process. In the meiotic recombination process, the repair of the DSB depends on a complex consisting of OsMFS1 and OsHOP2 to search for a homologous template chain and promote chain invasion reaction so as to ensure that the DSB is successfully repaired. OsMFS1 has a protein size of 23.9kD and contains a domain that is highly conserved across species: a coiled coil structure domain, the amino acid sequence of which is shown in SEQ ID NO. 2.

The invention also provides a rice complete sterility mutant Osmfs1 obtained by EMS mutagenesis, which shows that the rice complete sterility mutant Osmfs1 does not bloom in the flowering phase, has no vitality of pollen, has no fertile embryo sac and is completely fruitless in the mature phase.

The invention also provides a rice meiosis development key gene OsMFS1, the DNA sequence of which is shown in SEQ ID NO. 1. The CDS of OsMFS1 has a full length of 624bp, and comprises 10 exons and 9 introns. The Osmfs1 mutant is characterized in that a single-base mutation from G to A exists on the ninth exon of the OsMFS1 gene, so that the translation of the protein is terminated early.

The invention also provides an exploration on the action mechanism of the OsMFS1 gene and the protein coded by the gene in the meiosis process, particularly in the process of pollen mother cell chromosome development and embryo sac development.

The invention also provides application of the protein OsMFS1 or OsMFS1 gene or rice male sterile mutant Osmfs1 in rice breeding work.

The invention also provides application of the protein OsMFS1 or OsMFS1 gene or rice male sterile mutant Osmfs1 in rice male sterile breeding research. Specifically, the OsMFS1 gene is knocked out from normal wild rice to form male sterile rice.

The invention also provides application of the protein OsMFS1 or OsMFS1 gene in rice breeding with panicle closed flowers and no blooming. More particularly to the application in the rice breeding of the developmental disorder in the meiosis stage caused by the chromosome pairing abnormality.

The invention also provides the value of the protein OsMFS1 or OsMFS1 gene in the pharmaceutical and chemical industries.

Through the research on the complete sterile mutant Osmfs1, the meiotic development key gene OsMFS1 is obtained. The discovery and utilization of the rice meiosis gene deepens the understanding of the homologous recombination process, the homologous recombination process of meiosis can generate the exchange of homologous chromosome segments in a mode of generating Cross (CO), and a plurality of variations or innovations can be generated inevitably, but in the meiosis process, the quantity of CO is extremely limited, the gene of the patent can be utilized in the future, the quantity of CO generated can be increased or controlled probably by coordinating other genes for specifically regulating and controlling homologous recombination, so that the generation of variations or innovations can be accelerated, the breeding efficiency is greatly improved, the excellent seed source can be created, the hybridization efficiency is improved, high-quality rice varieties with high yield, strong stress resistance and wide adaptability can be cultivated, and therefore, the rice meiosis gene has important application value in plant breeding.

The helix-helix protein OsMFS1 is a heterodimer formed by OsHOP2, directly participates in single-chain invasion reaction mediated by OsRAD51 and OsDMC1, and determines the formation of CO and NCO, so that research and genetic modification can be carried out on the helix-helix protein OsMFS1 to achieve specific production value.

Drawings

FIG. 1 morphological comparison between wild type and Osmfs1 mutant;

FIG. 2 observation of anther development in wild type and Osmfs1 mutant in semi-thin sections;

FIG. 3 Transmission/scanning electron microscopy of pollen of wild type and Osmfs1 mutant;

FIG. 4 shows embryo sac development staining of wild type and Osmfs1 mutant;

FIG. 5 chromosome PI staining observation of pollen mother cells of wild type and Osmfs1 mutant;

fine positioning of OsMFS1 of fig. 6;

fig. 7 CRISPR-Cas9 knock-out vector;

FIG. 8 transgene knockout (CRISPR/Cas9) phenotypic identification;

FIG. 9 spatiotemporal expression analysis and subcellular localization of OsMFS 1;

FIG. 10 OsMFS1 and OsHOP2 interact;

FIG. 11 phenotypic identification of single and double knockout transgenic plants;

FIG. 12 chromosome PI staining observation of single and double knockout transgenic plants;

Detailed Description

The present invention will be further described with reference to the following examples, which are intended to illustrate the present invention and not to limit the scope of the present invention, and all simple modifications of the preparation method of the present invention based on the idea of the present invention are within the scope of the present invention. The following examples are experimental methods without specifying specific conditions, and generally follow the methods known in the art. The test materials used in the following examples were purchased from a conventional biochemical reagent store unless otherwise specified.

Example 1 phenotypic Observation of Osmfs1 mutant and map-based cloning of OsMFS1 Gene

(1) Phenotypic identification of Osmfs1 mutant

The Osmfs1 mutant is Ningjing No. 4 obtained by EMS mutagenesis, the mutant has no difference between the vegetative growth stage and the wild type, but shows that spikelet is closed and bloomed in the reproductive growth stage, and the plant is not fruitful at all in the maturation stage. To determine the sterility type of the plant, the pollen and embryo sac fertility of the mutant was identified by I₂KI staining, finding that the mutant is completely free of viable pollen and exhibits complete classical abortion (FIG. 1, A, plant phenotype of wild type and mfs1 mutant after ear emergence; B, spikelet of wild type and Osmfs1 mutant at maturity; C, D, iodine-potassium iodide for anthers of wild type and Osmfs1 at ear emergence (I)₂KI) solution staining). And wild-type pollen is awarded to the mutant plants, and the mutants are found to still show complete sterility. To this end we determined mfs1 to be a male-female gamete total sterile mutant. In order to clarify the cause of the male sterility of the mutant, morphological characteristics of pollen development stages were analyzed, and half-thin sections of anther development stages showed that, at the late stage of microspore, wild type pollen appeared to be round and regularly arranged on the anther wall, whereas mutant vacuolated pollen was not full and regularly arranged, and at the mature stage, mutant pollen was completely free of starch filling and had irregular pollen shape and shriveled (FIG. 2, half-thin sections of wild type anthers A-C and G-I, half-thin sections of mutants anthers D-F and J-L mfs1, half-thin sections of A and J-L mfs1D meiosis stage, early development of B and E microspores, C and F vacuolated pollen stage, G and J diplospora pollen stage, H and K mature pollen stage, I and L anther dehiscence stage, E, epidermis; en, inner layer; ML, middle layer; t, a tapetum layer; dy, dichotomous somatic cell; msp, microspore; BP, pollen diplospora; AP, abnormal pollen; MP, mature pollen). The results of scanning electron microscopy and transmission electron microscopy were consistent with those of semi-thin section, and the mature mutant pollen was abnormal in morphology, shriveled and surface sporopouenin was not dense, completely free of starch grain filling, but was filled with degradants (FIG. 3, wild type pollen scanning/transmission electron microscopy for A-C and G-H, mutant pollen scanning/transmission electron microscopy for D-F and J-K mfs1, mature pollen morphology for A, B, D and E, pollen grain surface for C and F, mature pollen grain cross-section for G, H, J and K transmission electron microscopy, anther wall for I and L wild type and mutant, C, center pillar, st, starch grain, I, pollen inner wall, F, base layer, te, pollen outer wall, E, epidermis, En, inner layer, ML, middle layer, T, tapetum layer). We observed the development of the mutant embryo sac with staining, and found that the mutant embryo sac develops abnormally during meiosis, and finally forms a completely non-fertile embryo sac (FIG. 4, A-J, the wild type embryo sac develops, and the characteristics of normal seven-cell eight-nucleus, K-O, the mutant dysplastic embryo sac can be observed). Although the results of the half-thin section show that no obvious difference is observed between the mutant and the wild type in the meiosis stage of anther development, the mutant is suspected to have abortion in the early stage of pollen development due to the abnormal morphology of microspores, and in order to verify the hypothesis, PI staining observation is carried out on the chromosome development in the meiosis stage of pollen development, and the fact that chromosomes cannot be paired are observed in the even line stage and the pachytene stage is found, the defect is more obvious in the final stage, the wild type can accurately identify twelve condensed point chromosomes, but the number of chromosomes in the mutant is far more than twelve, the phenomenon is that homologous recombination is disturbed due to abnormal chromosome pairing, a large number of chromosome fragments finally appear, and the chromosome fragments can not move to two stages along with the traction of spindle silk, in an episomal state (FIG. 5, development of wild-type chromosomes A-D and I-L, E-H and M-P mutationsSomatic chromosome development, blue arrows point to unpaired chromosomes, white arrows point to episomal chromosome fragments). To this end, we believe that the root cause of male sterility in mutants is a developmental disturbance during the meiotic phase of pollen development.

(2) Map-based cloning of OsMFS1 Gene

The mutant is obtained by EMS mutagenesis of Ningjing No. 4, the mutant mfs1 shows male and female gamete total sterility, and F for constructing positioning₂In the population, a hybrid plant is used as a female parent, an indica rice variety N22 is used as a male parent, hybridization pairing is carried out, and the obtained F is subjected to map-based cloning₂The population (about 1500 strains) was genetically mapped, the interval was determined by the initial linkage of the whole genome at chromosome nine markers RM23662 and RM23916, and then primers were designed for fine mapping (primer design website: http:// ricevarmap. ncpgr. cn/v1/), and finally the interval was mapped to 282kb interval of markers NN9S-L4 and NN9S-L7, at which time we performed genetic prediction of the interval, where LOC _ Os09G10850 was found to encode a meiosis-associated gene, which was then OsMFS1 as a candidate gene, sequenced at the eighth exon with a single base mutation from G to A, resulting in premature termination of protein translation, thus OsMFS1 was used as the phenotype target gene leading to complete sterility of the mutant (FIG. 6). The DNA sequence of OsMFS1 is shown in SEQ ID NO. 1; the protein is 23.9kD in size and contains a domain highly conserved in various species: a coiled coil structure domain, the amino acid sequence of which is shown in SEQ ID NO. 2.

Example 2: CRISPR/Cas9 technology and transgene phenotype identification

To further confirm that the OsMFS1 mutation caused the mutant phenotype, we knocked out the gene using CRISPR/Cas9 technology, and targeted knock out of the gene OsMFS1 by website design of specific knock out primers (http:// cbi. hzau. edu. cn/cgi-bin/CRISPR) (FIG. 7). In the obtained transgenic plants, two homozygous mutant lines named as Osmfs1-1 and Osmfs1-2 are identified by sequencing, the two knockout lines are subjected to single base insertion on the third exon, the Osmfs1-1 is inserted base A, and the Osmfs1-2 is inserted base T, which result in early termination of protein translation. Morphological observation is carried out on the plants of the transgenic families, the phenotype of the plants is consistent with that of the mutant Osmfs1, the plants and the mutant Osmfs1 are completely normal in vegetative growth, but the reproductive growth later stage shows that spikelets are completely sterile (figure 8, A, the phenotype of the plants of the wild type and the Osmfs1-1 mutant after the spike emergence; B, the gene editing mode of the Osmfs1-1 and Osmfs1-2 plants) so that the mutant completely sterile phenotype is determined to be caused by the mutation of the OsMFS 1.

Example 3: spatiotemporal expression analysis and subcellular localization of gene OsMFS1

Through real-time fluorescent quantitative PCR (qRT-PCR) and GUS staining experiments, the expression mode of the gene OsMFS1 is known, RNA of various tissues of wild plants and various development periods of anthers is extracted by using an RNA extraction kit, and specific quantitative primers are designed through a website for PCR (https:// www.genscript.com/tools/real-time-PCR-taqman-primer-design-tool). Quantitative results showed that OsMFS1 exhibited constitutive expression in each tissue and was highly expressed during the meiotic phase of anther development, indicating that OsMFS1 indeed functioned during the meiotic phase of anther development (fig. 9, a, quantitative expression levels of genes in wild-type tissues including leaves, leaf sheaths, stem nodes, stems, spikelets and roots, B, quantitative expression levels of genes in wild-type spikelet development phases including S5-12). Subsequently, we amplified a 2.6k fragment specific to the upstream promoter of the gene, which was used for starting a GUS reporter gene, and stained spikelets of the obtained GUS transgenic plants, and the result showed that the signal was strongest at the meiosis stage of spikelet development (FIG. 9, C, spikelets at various stages of GUS transgenic plants), which is also consistent with the previous fluorescent quantitative PCR result, and further confirmed that OsMFS1 is specifically expressed at the meiosis stage. In order to determine the functional position of the gene OsMFS1, GFP signals were detected in tobacco cells and rice protoplasts by subcellular localization experiments, and were found to exhibit nuclear localization, indicating that OsMFS1 is a nuclear localization protein (FIG. 9, D-G, fluorescence signal detection in rice protoplasts, H-K, fluorescence signal detection in tobacco cells).

Example 4: OsMFS1 and OsHOP2 interaction validation and double-process phenotype identification

To explore the function of OsMFS1, we found OsMFS1 and OsHOP2 interaction by yeast two-hybrid experiments, and further confirmed that both proteins can interact in vitro by pull-down experiments (fig. 10, a, yeast plasmid co-transformed with AD-OsMFS1 and BD-OsHOP2 using yeast two-hybrid system, observing yeast growth on defective media, B, in vitro protein interaction experiments, testing protein binding using western-blot). Through a CRISPR/Cas9 technology, plants with single Oshop2 and double Osmfs1/Oshop2 are obtained, morphological observation is carried out on the plants of the transgenic families, and the results show that the plants with single Oshop2 and double Osmfs1/Oshop2 are all completely normal in vegetative growth period, but completely sterile in later reproductive growth period, and the plants are obtained through I₂KI staining, which revealed that both single and double processes were completely free of viable pollen, and exhibited complete catastrophe, similar to the phenotype of mfs 1. The sequencing result shows that a single-bulge Oshop2-1 has a single-base A insertion at the third exon, a double-bulge has an A/T insertion at the third exon of the gene OsMFS1, a single-base C insertion is formed in one strand of the OsHOP2 gene, and 5 bases are deleted in the other strand (FIG. 11, A, wild type at maturation stage, Osmfs1-1, Oshop2-1 and Osmfs1/Oshop2 plant phenotype, B, wild type at maturation stage, Osmfs1-1, Oshop2-1 and Osmfs1/Oshop2 spikelet, C-F, potassium iodide staining observation, G, transgenic plant Osmfs1-1, Oshop2 and Osmfs1/Oshop2 gene editing mode). Then, PI staining observation was carried out on the single-and double-bulge chromosomes, and it was found that the development of the single-and double-bulge chromosomes was defective and that a large number of chromosome fragments similar to Osmfs1 were present (FIG. 12, PI staining observation of pachytene, metaphase I and anaphase I of the development of wild-type, Osmfs1-1, Oshop2-1 and Osmfs1/Oshop2 chromosomes). Therefore, we determined that any gene mutation in the complex OsMFS1/OsHOP2 leads to the abnormal development of plant chromosomes, and finally shows complete sterility.

Sequence listing

<110> Nanjing university of agriculture

<120> rice meiosis development related gene OsMFS1 and application thereof

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acatattggt attggatgca ttttttgtaa taatgttgtt ttaagggtat gagcattttg 4080

aggccatgct gatgatgtta tttaaaaagc ttgcacttgt ttgctgtttg gactctatac 4140

tgataagcct aaatcatgtc catctgtttt gattagttct tggtgctatt tgtcttctct 4200

tagctatcat accaaacttg ttattctagt tacttctggg attgattata atatccacat 4260

gagtacaata tttcttcatg ttgtaagtta tgctaagcgc cctcaataag gatatccctc 4320

aaacttcctt ggcttgaatg gtcactaaat tgcaagggag caaagccatt gcccagtaga 4380

acacttctac acttcatgtg tctggcttga gtgatcatac cccaacggcc caacccagtc 4440

actttccata tcaggactca aaagcttgtc attctctcgg gttgatagaa gtgttctaga 4500

aatattgata tctttgtgag tgatgatcca atagtttagc ttttgctgtg attagttccc 4560

ttcaaacttc caacttttcc gatcacatca aatgtttgga cacatgcatg gagcattaaa 4620

tgtggacgta aaaaaaacca attgcacagt ttgcatgtaa attgcgagac gaatcttttg 4680

agccttatta cgccatgagt tgacaatgtg gtgctacagt aaacatttgc taatgacgga 4740

ttaattaggc ttaatagatt cgtctcgcag tttacagccg gaatctgtaa tttgttttgt 4800

tattagttta cgtttaatac ttcaaatgtg tatccgtata cgtcaaattt tttttgccaa 4860

aacaactaaa cacggccttt gtccactcca catcaccaca actaaagttt cacatatgtt 4920

gtgctgctgg ttttttgagc atccccaatg cttttccttc acgctagtac cctctgtttg 4980

tgtaacaagg cgttttcact tctcatagtt tcctctgttc tcatgttctg atttactctt 5040

gtgaaatttc aggtggggat aactgaagat ttcgaatatc tgcagtaa 5088

<210> 2

<211> 207

<212> PRT

<213> japonica rice variety NIP (Japan rice variety Nippobrare)

<400> 2

Met Ser Lys Lys Arg Gly Leu Ser Leu Glu Glu Lys Arg Glu Gln Met

1 5 10 15

Leu Gln Ile Phe Tyr Asp Ser Gln Asp Phe Tyr Leu Leu Lys Glu Leu

20 25 30

Glu Lys Leu Gly Pro Lys Lys Gly Val Ile Ser Gln Ser Val Lys Asp

35 40 45

Val Val Gln Ser Leu Val Asp Asp Asp Leu Val Leu Lys Asp Lys Ile

50 55 60

Gly Thr Ser Val Tyr Phe Trp Ser Leu Pro Ser Cys Ala Gly Asn Gln

65 70 75 80

Leu Arg Thr Thr Tyr Ser Lys Leu Glu Ser Asp Leu Ser Ser Ser Lys

85 90 95

Lys Arg Phe Ile Glu Leu Val Glu Gln Arg Glu Asn Leu Lys Arg Gly

100 105 110

Arg Glu Asp Ser Asp Glu Arg Glu Ala Ala Leu Glu Glu Leu Lys Ala

115 120 125

Val Glu Gln His His Lys Lys Leu Lys Glu Glu Leu Ala Ala Tyr Ala

130 135 140

Asp Ser Asp Pro Ala Ala Leu Glu Ala Met Asn Asp Ala Ile Glu Val

145 150 155 160

Ala His Ala Ala Ala Asn Arg Trp Thr Asp Asn Ile Phe Thr Leu Gln

165 170 175

Gln Trp Cys Ser Thr Thr Phe Pro Gln Ala Lys Glu Gln Leu Glu His

180 185 190

Met Tyr Arg Glu Val Gly Ile Thr Glu Asp Phe Glu Tyr Leu Gln

195 200 205

Claims (3)

1, SEQ ID NO.1OsMFS1Or the protein shown as SEQ ID NO.2OsMFS1Application in male sterility breeding of rice.

2, SEQ ID NO.1OsMFS1Or the protein shown as SEQ ID NO.2OsMFS1The application in rice breeding with small ear closed flowers and no blooming.

3, the gene shown in SEQ ID NO.1OsMFS1Application in the research of cultivating rice sterile mutant.

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