CN106497936A - The albumen of control rice male fertility and its encoding gene and application - Google Patents
The albumen of control rice male fertility and its encoding gene and application Download PDFInfo
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- 235000007164 Oryza sativa Nutrition 0.000 title claims abstract description 94
- 235000009566 rice Nutrition 0.000 title claims abstract description 74
- 108090000623 proteins and genes Proteins 0.000 title claims abstract description 69
- 230000035558 fertility Effects 0.000 title claims abstract description 18
- 240000007594 Oryza sativa Species 0.000 title claims description 21
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- 108091033409 CRISPR Proteins 0.000 claims description 10
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Classifications
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/415—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from plants
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H1/00—Processes for modifying genotypes ; Plants characterised by associated natural traits
- A01H1/02—Methods or apparatus for hybridisation; Artificial pollination ; Fertility
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
- C12N15/82—Vectors or expression systems specially adapted for eukaryotic hosts for plant cells, e.g. plant artificial chromosomes (PACs)
- C12N15/8241—Phenotypically and genetically modified plants via recombinant DNA technology
- C12N15/8261—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield
- C12N15/8287—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield for fertility modification, e.g. apomixis
- C12N15/8289—Male sterility
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- Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
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- General Engineering & Computer Science (AREA)
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- Biotechnology (AREA)
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- Wood Science & Technology (AREA)
- Bioinformatics & Cheminformatics (AREA)
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- Developmental Biology & Embryology (AREA)
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- Cell Biology (AREA)
- Gastroenterology & Hepatology (AREA)
- Medicinal Chemistry (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention discloses a kind of albumen of control rice male fertility and its encoding gene and application.The albumen has SEQ ID NO in sequence table：Aminoacid sequence shown in 3, the genomic dna sequence of its encoding gene and cDNA sequence are respectively such as SEQ ID NO in sequence table：1 and SEQ ID NO：Shown in 2.By rite-directed mutagenesises and functional verification, the present invention proves that the gene has the function of control rice male fertility, be mutated the gene, can obtain male sterile rice strain in wild rice, and there is in rice breeding important using value.
Description
Technical field
The invention belongs to field of plant genetic, and in particular to a kind of gene of control rice male fertility and should
The purposes of gene.
Background technology
Oryza sativa L. (Oryza sativa L.) is one of most important cereal crops.The population of whole world half or so is with water
Rice exceedes the countries in Asia of total world population's number 60% as staple food, especially population.The yield for improving Oryza sativa L. is full generation
The long term object of boundary rice breeding man.
Rice breeding has two kinds of main paths at present.One kind be by hybridize and backcrossing etc. means, breeding high-yield stable yields
Conventional rice variety.Conventional Rice is without the need for hybrid seeding, convenient and strength-saving；Usually, the rice matter of conventional Rice is better than common hybrid paddy rice.But
Variet complexity decline of often produce that to be conventional Rice plant from being reserved seed for planting year after year, affects rice yield.And hybrid paddy rice is another
The successful methods of rice breeding.Hybrid rice refers to the parental rices different with two genetic background, through hybridizing the F for obtaining1
The seed in generation, by F1Peasant planting is supplied directly to for seed.F1In generation, has obvious hybrid vigor, shows after field planting
The merits such as high yield, high-quality and multi-resistance.
Conventional hybrid rice system has two kinds of ternary hybrid rice and two-line hybrid rice at present.Ternary hybrid rice includes sterile
System, maintainer and restorer three are.The male organs development of male sterility line is abnormal, it is impossible to which self-fertility, female organ is but
Development normal energy receives foreign pollen and fertilization.Male sterility line receives after the pollen of restorer awards, solid normal, and
And the new first generation of hybrid fertility restorer for producing is normally, energy self-fertility, and with stronger advantage.Sterile line selfing is shaky,
It is that its pollination is solid to need by maintainer, and offspring still keeps the sterile gene of male.At present, series of three-series hybrid rice kind is not
It is to be cytoplasmic male sterile line to educate, wherein Yebai, red lotus type and bag bench-type be internationally recognized 3 kinds of cytoplasmatic males not
Educate Oryza sativa L. main Types.
In double-linear hybrid rice, sterile line is usually photo-thermo-sensitive genetic male sterile line.Under the external conditions such as some light temperature, sterile line
Fertility be restored, selfed seed can be harvested, for the production of hybrid seeds of next year, thus extra maintainer is no longer needed.Three are
The sterile line of hybrid paddy rice is selected generally from cytoplasmic male sterility strain, and photo-thermo-sensitive genetic male sterile line is generally karyogene control fertility.
Hybrid rice is extensively applied in production.Through effort for many years, science are in rice cytoplasmic male sterile
Regulatory mechanism in terms of have been achieved with more progress, while also having cloned dozens of regulates and controls male sterile karyogene.Reflect
The male sterile karyogene of the adjusting and controlling rice made mainly acts on the middle and late stage of paddy rice stamen development, have impact on including subtrahend point
The development and degraded of the process, tapetum split, the deposition of exposore, the redox state in the cracking and coyote hole of pollen bag
Deng in interior multiple processes.Although achieved with the studies above be in progress, control rice fertility molecule mechanism still not it is clear that
Especially the gene of adjusting and controlling rice early stage stamen development, is even more rarely reported.
Existing research is had shown that, is combined the Plant Genome editing technique that develops recently using transgenic technology, will be helped
In the gene function for disclosing control Important Agricultural character, be potentially possibly used for breeding reality so as to contribute to more rapidly excavating
The gene that tramples, efficiently cultivates the excellent new rice variety being worth with production application.But existing achievement is also few
Can be directly thrown in production application, more participate in the male sterile gene of adjusting and controlling rice also to be found.
Content of the invention
It is an object of the invention to provide a kind of albumen of control rice male fertility and its encoding gene, for illustrating water
The male sterile regulatory mechanism of rice, the cultivation for hybrid rice provide potential breeding resources.
The present invention is found that the albumen of an adjusting and controlling rice stamen early development by genetic research, when the encoding gene strikes
The stamen development exception of Oryza sativa L. after removing, can be made, and pollen can not be produced completely, ultimately result in male sterility.
The albumen of control rice male fertility provided by the present invention, is named as OsMPL, from Oryza sativa L. (Oryza
Sativa) kind Japan is fine, with one of following aminoacid sequences：
1) the SEQ ID NO in sequence table：3；
2) the SEQ ID NO in sequence table：Replacement, disappearance of 3 aminoacid sequences through one or more amino acid residues
Or add and the derivative aminoacid sequence with equal function.
SEQ ID NO in sequence table：3 are made up of 349 amino acid residues.Amino acid residue is carried out replacing, is lacked
Or the method that adds is well-known to those skilled in the art, typically its encoding gene is entered using engineered means
Row mutation, then gives expression to corresponding albumen again.By expressing the albumen in plant, and test these plants flower pesticide and
The developmental state of pollen, it can be determined that the function whether albumen after these changes also has control rice male fertility occurs.
The present invention provide control rice male fertility albumen OsMPL expression and/or activity can affect flower pesticide and
The development of pollen, so as to the male fertile of controllable Oryza sativa L..
The encoding gene OsMPL for encoding above-mentioned control rice fertility albumen can be the cDNA sequence of the gene, also may be used
Being the genomic dna sequence of the gene, or there is more than 90% homology with these sequences and encode identical function
The DNA sequence of albumen.Such as in sequence table SEQ ID NO：Genomic dna sequence and SEQ ID NO shown in 1：Shown in 2
CDNA sequence.
The nucleotide sequence of the encoding gene is knocked out or changed, the albumen that can not express the control rice fertility is allowed to
Or reduce expression, or the aminoacid sequence of expression disappearance is occurred or is made a variation, and then cause the aminoacid sequence corresponding
Polypeptide loss of activity, may result in pollen and anther development exception.
Provided by the present invention control rice male fertility albumen or its encoding gene purposes, including but not limited under
State several aspects：
1) fertility of rice male organ is controlled；
2) as the molecular marker of rice male fertility；
3) initiative rice male-sterile plants system；
4) new heterosis utilization approach is created, cultivates new varieties.
The method of present invention initiative rice male-sterile plants system, including：The OsMPL genes of purpose Oryza sativa L. are knocked out, or makes mesh
Oryza sativa L. the gene mutation cause express protein inactivation, or suppress purpose Oryza sativa L. in OsMPL genes expression, obtain water
Rice male sterility strain；Wherein described purpose Oryza sativa L. is the Oryza sativa L. for carrying OsMPL genes.
Knockout or mutation that OsMPL genes can achieve using conventional gene engineering method, and suppress or reduce
The expression of OsMPL genes.Can be realized by any known site-directed point mutation or EVAC, such as CRISPR/
The skills such as Cas9 systems, TALEN systems, zinc finger enzyme system, RNA interference and artificial tiny RNA technology (artificialmicroRNA)
Art means.
In some embodiments of the invention, by the CRISPR/Cas9 vector introduction plant cells of target OsMPL genes,
Tissue or organ, then the plant cell being converted, tissue or organ are cultivated into plant, obtain male sterile transgenic and plant
Thing.
The expression vector of rite-directed mutagenesises, expression cassette, transgenic can be carried out on rice genome to the OsMPL genes
Cell line and Host Strains fall within protection scope of the present invention.
The element of rite-directed mutagenesises paddy gene OsMPL of the present invention or its homologous sequence can be imported by plant expression vector plants
Thing tissue, cell or organ.Agrobacterium tumefaciens can be can be used for for any one for building the carrier of the plant expression vector
Or Agrobacterium rhizogenes convert the binary vector of plant or can be used for carrier of plant micropellet bombardment etc., can also be can be in prokaryote
The carrier of middle duplication, such as pUC serial carriers or pBluescript serial carriers etc..
For the ease of being identified to transgenic plant cells or plant and being screened, plant expression vector used can be carried out
Processing, such as add the coding that can be expressed in plant can produce the enzyme or luminophor of color change gene (gus gene,
GFP genes, luciferase genes etc.), have resistance antibiotic marker (hygromycin phosphotransferase gene, carboxylic benzyl penicillium sp
Plain label or kanamycin label etc.) or anti-chemical reagent marker gene (such as anti-herbicide gene) etc..
The plant expression vector for carrying the paddy gene OsMPL of mutation present invention control male fertile can be by using original
Raw plastid-chemistry mediation method (Ca2+, PEG), Ti-plasmids, Ri plasmids, plant viral vector, directly delivered DNA, pollen tube import,
The combination conversion of any one of microinjection, electric shock, particle gun, the conventional biology methods such as agriculture bacillus mediated or several method
Plant cell, tissue or organ, and the plant cell of conversion, tissue or organ are cultivated into plant；The tissue and organ can
Fruit pod, calluss, stem apex, blade and seed including host plant etc..
Additionally, the transfer-gen plant by the paddy gene OsMPL for being mutated present invention control male fertile carries out subculture training
After supporting, the mutant plant of gene pure therefrom can be further filtered out.
The rice male-sterile plants system that the present invention is obtained can be cross-breeding as female parent.Using conventional genetic means
The OsMPL genes are proceeded in the cell or tissue of the rice male-sterile plants system, is regenerated, recovery male can be cultivated into and educated
The plant of property.
Present invention finds regulating and controlling the albumen and its encoding gene OsMPL of its male fertile in Oryza sativa L., it is mutated the gene and obtains
Obtain male sterile phenotype.Therefore, OsMPL genes according to the present invention have potential application valency in plant hybridization breeding
Value.
Description of the drawings
Fig. 1 shows the sequence variation situation for passing through CRISPR/Cas9 system rite-directed mutagenesises OsMPL genes in embodiment 3,
Compared with wild type, in the representational strain of two target sites, the difference that there is 1 base at target site is inserted
Enter, cause translation frameshift.
Fig. 2 shows the phenotype of wild type (left side) and Osmpl-1 afunction mutant (right side) outward appearances in embodiment 3,
Osmpl-1 shows as bearing seeds, but other organs do not have notable difference with wild type control ratio.Scale=10 centimetre.
Fig. 3 shows the flower pesticide phenotype of Osmpl-1 and Osmpl-2 afunction mutant in embodiment 3.Wherein, A is to sweep
Retouch electromicroscopic photograph：Left side is wild type, and middle is Osmpl-1 mutants, and right side is Osmpl-2 mutants；B is I2- KI is dyeed
Microphotograph afterwards：Left side is wild type, and middle is Osmpl-1 mutants；Right side is Osmpl-2 mutants.Scale=100 are micro-
Rice.
Specific embodiment
In following embodiments, method therefor is conventional method if no special instructions, and concrete steps can be found in：
《Molecular Cloning:A Laboratory Manual》(Sambrook, J., Russell, David W.,
Molecular Cloning:A Laboratory Manual, 3rdEdition, 2001, NY, Cold Spring Harbor).
Embodiment 1, the rite-directed mutagenesises of OsMPL genes
(1) entry vector of OsMPL rite-directed mutagenesises is built：
According to two pairs of specificity spacer sequences for being capable of target OsMPL genes of sequence specific Sexual behavior mode, synthesis following two
To primer：
F1：5’-GGCAAGCAAGAGGGCGCCGCAGCG-3’(SEQ ID NO：4)；
R1：5’-AAACCGCTGCGGCGCCCTCTTGCT-3’(SEQ ID NO：5)；
F2：5’-GGCACGGCGTTGAGGTCGCGGAAC-3’(SEQ ID NO：6)；
R2：5’-AAACGTTCCGCGACCTCAACGCCG-3’(SEQ ID NO：7).
Respectively by two pairs of primer annealings after, be connected into the pOs-sgRNA plasmids after BsaI enzyme action (Miao et al.,
2013, Cell Research 23 (10):1233-1236.) in.The screening correct carrier of spacer direction of insertion (using R1 and
M13R sequencing primers can amplify target stripe) and be sequenced, be sequenced correct plasmid be named as pOs-sgRNA-MPL-1 and
POs-sgRNA-MPL-2 is used for building OsMPL rite-directed mutagenesises end carrier.
(2) structure of OsMPL rite-directed mutagenesises end carrier：
In order to build OsMPL rite-directed mutagenesises end carrier, first pOs-sgRNA-MPL-1 and pOs-sgRNA-MPL-2 is carried
Body and pH-Ubi-cas9-7 carriers (Miao et al., 2013, Cell Research 23 (10):1233-1236.) LR is carried out
Connection product is converted bacillus coli DH 5 alpha competent cell by reaction (Invitrogen companies), enters positive gram of performing PCR screening
Grand, the plasmid containing the element for being capable of target OsMPL genes is extracted, sequence verification is correctly used for the wound healing of Oryza sativa L. afterwards and converts.
Embodiment 2, the acquisition of the transgenic paddy rice of rite-directed mutagenesises OsMPL genes：
(1) rice callus induction：
Full Japanese fine rice paddy seed is chosen, kind of a skin is peelled off, after sterilizing washing, is uniformly clicked and entered with 2 mg/litres
The sterilizing MS solid mediums of 2,4- dichlorphenoxyacetic acids (2,4-D), the callus induction in 5 days of 32 DEG C of continuous lights are formed.
(2) Agrobacterium-mediated Transformation：
The plasmid of OsMPL rite-directed mutagenesises end carrier is converted Agrobacterium EHA105 competent cells, coating zone with electrization
There is the solid LB media of 50 micrograms per litre spectinomycins and 50 micrograms per litre rifampicin, 28 DEG C of dark culturing are after 2 days, special with gene
Different primer U1 and C1 screening positive clones.The positive colony for obtaining is containing 50 micrograms per litre spectinomycins and 50 micrograms per litre rifampicin
Fluid medium in 28 DEG C cultivate to OD600=0.8, for rice conversion.
U1：5’-GCGGTCGTTCATTCGTTCTAGATCG-3’(SEQ ID NO：8)；
C1：5’-AGTTGGGCGATCAGATTCTC-3’(SEQ ID NO：9).
(3) rice callus conversion：
The good calluss of upgrowth situation are chosen, is soaked 2 minutes in bacterium solution, is dried on aseptic filter paper afterwards, then
Calluss are moved to the NB containing 100 micromoles per liter acetosyringones and co-cultures culture medium, co-cultured 3 days under 25 DEG C of dark.
(4) rice callus are screened：
After co-culturing 3 days, with aseptic water washing wound healing 5 times, then with 200 milliliters containing 500 mg/litre Carbenicillins
The aseptic washing of sodium one time, carefully removes liquid, is gripped calluss to aseptic filter paper with aseptic nipper, shifts after drying
(the carboxylic benzyl penicillium sp of the 2,4-D containing 2 mg/litres, the hygromycin of 50 mg/litres and 400 mg/litres to NB screening culture medium
Plain sodium), 32 DEG C of continuous lights 2 weeks.
(5) differentiation of positive wound healing：
Positive calluss of the well-grown in bright yellow are chosen, and the pre- division culture mediums of NB are moved to aseptic nipper and (are contained 1
Mg/litre naphthalene acetic acid (NAA), 5 mg/litre abscisic acid (ABA), 2 mg/litre kinetins (kinetin), the tide of 25 mg/litres
Enzyme element and the carbapen of 200 mg/litres), 32 DEG C of continuous light cultures.Eugonic calluss are selected after 2 weeks
MS division culture mediums are proceeded to (containing 0.02 mg/litre NAA, 2 mg/litres kinetin, the damp enzyme element of 50 mg/litres and 200 millis
The carbapen of grams per liter), 32 DEG C of continuous light cultures.The seedling length for breaking up out is waited to 2 to 5 millimeters, is proceeded to without sharp
The week of MS culture medium culturings 2 to 3 of element and antibiotic, move into afterwards be placed in soil growth in greenhouse (temperature 28-30 DEG C, 16 hours
Illumination/8 hour are dark).
Embodiment 3, the Molecular Identification of OsMPL afunction mutants and phenotypic evaluation
(1) in transgenic paddy rice OsMPL genes rite-directed mutagenesises：
The blade of clip Transgenic Rice Seedlings, extracts plant with plant genomic DNA extracts kit (QIAGEN companies) total
DNA.Gene-specific primer MPL-F and MPL-R are designed according to the locus DNA sequence of OsMPL genes and enters performing PCR reaction, expanded
Increase the fragment containing OsMPL targeted mutagenesis site, and the catastrophe for checking OsMPL genes is sequenced：
MPL-F：5'-GGAGACAGCCTCCCTTCGAGTACACTA-3'(SEQ ID NO：10)；
MPL-R：5'-GGCAGGAGGAGGAGGAAGAAGAAGA-3'(SEQ ID NO：11).
The catastrophe of sequencing detection OsMPL genes, finds the different insertions for detecting a base at target site
The two class genes mutant, be respectively designated as Osmpl-1 and Osmpl-2, as shown in Figure 1.Osmpl-1 and Osmpl-2 are removed
Do not bear seeds, outward appearance does not have a notable difference with the Oryza sativa L. of wild type, as shown in Figure 2.
(2) observation of OsMPL rite-directed mutagenesises transgenic paddy rice flower pesticide phenotype：
This research adopts scanning electron microscopic observation flower pesticide outward appearance, and Taking Pictures recording；Use I2Inside-KI dyeing observation pollen bags
Dividing condition；And the pollen with wild rice is awarded to OsMPL afunction mutants, whether observation mutant ovary just develops
Often.As shown in figure 3, scanning electron microscope analysis show that Osmpl-1 and Osmpl-2 afunction mutant flower pesticide outward appearances are all distorted, with
Quite different (see the A in Fig. 3) of wild type.I2After-KI dyeing, micro- sem observation shows Osmpl-1 and Osmpl-2 afunction
The inside of mutant flower pesticide does not contain pollen grain, and full of can be by I in the flower pesticide of wild type2The mature flower powder of-KI dyeing is (see figure
B in 3).When awarding with wildtype pollens, Osmpl-1 and Osmpl-2 mutants can be normally solid, and the function of OsMPL is described
Disappearance causes male sterility of rice, and Stamen development is normal, illustrates that knocking out OsMPL may result in male sterility of rice.
Claims (9)
1. the application of rice Os MPL albumen or its encoding gene in control rice male fertility, it is characterised in that described
OsMPL albumen has one of following aminoacid sequences：
1) the SEQ ID NO in sequence table：3；
2) the SEQ ID NO in sequence table：3 aminoacid sequences through one or more amino acid residues replacement, lack or add
Plus and the derivative aminoacid sequence with equal function.
2. application as claimed in claim 1, it is characterised in that the sequence of the encoding gene of rice Os MPL albumen is its gene
Group DNA sequence or cDNA sequence, or there is more than 90% homology with these sequences and encode the DNA of identical function albumen
Sequence.
3. application as claimed in claim 2, it is characterised in that the sequence such as sequence of the encoding gene of the rice Os MPL albumen
SEQ ID NO in list：1 or SEQ ID NO：Shown in 2.
4. application as claimed in claim 1, it is characterised in that knock out or change the nucleotide sequence of the encoding gene, make
Can not express the OsMPL albumen or expression reduce, or the aminoacid sequence of expression disappearance is occurred or is made a variation, enter
And the corresponding polypeptide loss of activity of the aminoacid sequence is caused, cause male sterility of rice.
5. a kind of method of initiative rice male-sterile plants system, including：The OsMPL genes of purpose Oryza sativa L. are knocked out, or makes purpose water
The OsMPL gene mutation of rice causes the expression of OsMPL genes in the protein inactivation that expresses, or suppression purpose Oryza sativa L., obtains water
Rice male sterility strain；Wherein described purpose Oryza sativa L. is the Oryza sativa L. for carrying OsMPL genes, the following ammonia of the OsMPL gene codes
Base acid sequence 1) or 2) shown in protein：
1) the SEQ ID NO in sequence table：3；
2) the SEQ ID NO in sequence table：3 aminoacid sequences through one or more amino acid residues replacement, lack or add
Plus and the derivative aminoacid sequence with equal function.
6. method as claimed in claim 5, it is characterised in that SEQ ID in the sequence of the OsMPL genes such as sequence table
NO：1 or SEQ ID NO：Shown in 2.
7. method as claimed in claim 5, it is characterised in that by CRISPR/Cas9 systems, TALEN systems, zinc finger enzyme system
System, RNA interference or artificial tiny RNA technology realize knockout or the mutation of OsMPL genes.
8. method as claimed in claim 7, it is characterised in that the CRISPR/Cas9 vector introductions of target OsMPL genes are planted
Thing cell, tissue or organ, make OsMPL genes undergo mutation, then the plant cell being converted, tissue or organ are cultivated into plant
Strain, obtains male sterile transgenic line.
9. a kind of rice breeding method, is cross-breeding using the rice male-sterile plants system of claim 5 initiative as female parent.
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| CN107164393A (en) * | 2017-07-21 | 2017-09-15 | 东北农业大学 | Cucumber female related gene CsPAP fib and encoding proteins and application |
| CN107338257A (en) * | 2017-08-22 | 2017-11-10 | 广东省农业科学院农业生物基因研究中心 | Applications of the rice glutaredoxin gene OsGrxC2 in breeding |
| CN108220328A (en) * | 2017-12-15 | 2018-06-29 | 中国烟草总公司郑州烟草研究院 | A kind of method for identifying genome editor's Mutants homozygous |
| CN108660150A (en) * | 2018-05-22 | 2018-10-16 | 山东省水稻研究所 | A method of improving Rice Salt ability |
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