CN114164208A - Gene editing sequence and method for creating soybean cell nucleus male sterile line - Google Patents
Gene editing sequence and method for creating soybean cell nucleus male sterile line Download PDFInfo
- Publication number
- CN114164208A CN114164208A CN202110871824.6A CN202110871824A CN114164208A CN 114164208 A CN114164208 A CN 114164208A CN 202110871824 A CN202110871824 A CN 202110871824A CN 114164208 A CN114164208 A CN 114164208A
- Authority
- CN
- China
- Prior art keywords
- soybean
- gene editing
- sequence
- male sterile
- creating
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12Q1/6888—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms
- C12Q1/6895—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms for plants, fungi or algae
-
- 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
- C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
- C12N15/113—Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing
-
- 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
-
- 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)
- C12N15/8216—Methods for controlling, regulating or enhancing expression of transgenes in plant cells
- C12N15/8218—Antisense, co-suppression, viral induced gene silencing [VIGS], post-transcriptional induced gene silencing [PTGS]
-
- 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
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6844—Nucleic acid amplification reactions
- C12Q1/686—Polymerase chain reaction [PCR]
-
- 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
- C12N2310/00—Structure or type of the nucleic acid
- C12N2310/10—Type of nucleic acid
- C12N2310/20—Type of nucleic acid involving clustered regularly interspaced short palindromic repeats [CRISPRs]
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/13—Plant traits
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/156—Polymorphic or mutational markers
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Genetics & Genomics (AREA)
- Engineering & Computer Science (AREA)
- Organic Chemistry (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Molecular Biology (AREA)
- Biotechnology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Biophysics (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Physics & Mathematics (AREA)
- Microbiology (AREA)
- Analytical Chemistry (AREA)
- Plant Pathology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Botany (AREA)
- Immunology (AREA)
- Cell Biology (AREA)
- Mycology (AREA)
- Gastroenterology & Hepatology (AREA)
- Medicinal Chemistry (AREA)
- Virology (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The invention relates to the technical field of genetic breeding, in particular to a gene editing sequence for creating a soybean cell nucleus male sterile line by using a CRISPR/Cas9 gene editing technology and a creating method using the sequence. The CRISPR-Cas9 gene editing technology is utilized to edit GmNACK2 through a specific target spot, a nuclear male sterile line is created, and the significant breeding problems of difficult soybean hybridization, increase of natural outcrossing rate and the like are effectively solved. Has important functions in the genetic improvement of soybean and the improvement of soybean yield.
Description
The application claims the priority of 'an InDel marker for soybean nuclear male sterility and a method for creating a male sterile line' with application number 202110426981.6 applied on 20.4.2021, original acceptance organization is China.
Technical Field
The invention relates to the technical field of genetic breeding, in particular to a method for editing a target gene by using a CRISPR/Cas9 gene editing technology and creating a soybean cell nucleus male sterile line by using a gene editing sequence.
Technical Field
The soybean is one of the most major economic crops in the world, is rich in nutrition, has about 40 percent of protein content which is obviously higher than the level of other major crops, and has about 20 percent of oil content. The soybean also contains soybean isoflavone, dietary fiber and other essential nutrients. However, the low yield per unit of soybean in China still is a problem which needs to be broken through urgently by the soybean industry at present. The annual demand of soybeans in China exceeds 1 hundred million tons, and the domestic yield can only meet about 10 percent. Therefore, increasing yield is currently the primary task of soybean research.
The heterosis has obvious promotion effect on the improvement of the crop yield, and the yield of the hybrid seeds of most crops can be increased by about 20 percent compared with the conventional seeds. The male sterility of soybean is the most important agronomic character used in the cross breeding, and is also the most character used in the cross breeding in the current production. Meanwhile, the soybean male sterile line has wide application prospect, effectively solves the problem of difficult soybean hybridization and increases the natural outcrossing rate. Therefore, the research on the male sterility character of the soybean, the cloning of the sterile gene, the analysis of the sterile molecular mechanism and the creation of the sterile line are the working bases of the application of the soybean hybridization, the utilization of the new generation hybridization technology and the improvement of the soybean yield.
CRISPR (clustered Regularly interrupted Short Palindromic repeats)/Cas system is the most popular in recent years, Chinese is translated into clustered Regularly-spaced Short Palindromic repeats by using the most extensive gene editing tool, and the CRISPR/Cas system is used as acquired immunity of bacteria and is used for resisting phage invasion. Cas9 has the ability to cleave DNA like other common restriction endonucleases, except that most restriction endonucleases recognize fixed DNA sequences, whereas recognition of DNA by Cas9 requires a sgRNA to specifically pair with target DNA, bringing Cas9 to the target site, and cleavage of the target site by Cas9 requires recognition of a PAM sequence on the target DNA.
At present, no method for effectively editing a certain gene target by using a gene editing technology to generate a soybean male sterile line is established. However, although methods for molecular screening and gene editing of other traits of soybean plants exist in the prior art, most of the methods have the problems of unstable technology, long time, complicated technology and the like, and solution is needed urgently.
Disclosure of Invention
The invention aims to provide a gene editing sequence for creating a soybean plant cell nucleus male sterile line by using a gene editing technology CRISPR/Cas9, wherein the gene editing sequence is sgRNA of a soybean plant cell nucleus male sterile gene target spot, and the nucleotide sequence of the sgRNA is SEQ ID NO. 1.
The second purpose of the invention is to provide a method for creating a soybean plant cell nucleus male sterile line by using the gene editing sequence, which comprises the following steps:
s1, constructing a gene editing sequence described by SEQ ID NO.1 on a knockout expression Vector pMDC123, and naming the knockout Vector as Vector-GmNACK 2;
s2, editing a Vector-GmNACK2, introducing an allelopathic strain, and performing soybean genetic transformation of direct organ regeneration by using a method of mediating soybean cotyledon node transformation by the allelopathic strain to obtain a transgenic soybean positive plant;
s3, detecting a mutant sequencing result, namely performing mutant verification on a transgenic positive plant of the transformed Vector-GmNACK2, performing PCR sequencing, and preliminarily judging that the gene is a mutant if a set peak appears at the sgRNA; performing monoclonal sequencing on the plant which is preliminarily judged as the mutant, and performing final judgment according to a sequencing result; the primer pair used for PCR sequencing and monoclonal sequencing is the same, the forward sequence of the primer pair is shown as SEQ ID NO.2, and the reverse sequence is shown as SEQ ID NO. 3.
Preferably, the competent strain is agrobacterium GV 3101.
The invention has the beneficial effects that:
1. according to the invention, the GmNACK2 is edited by using a CRISPR-Cas9 gene editing technology through a specific target point, a nuclear male sterile line is created, and the significant problems of difficult soybean hybridization, increase of natural outcrossing rate and the like in breeding are effectively solved.
2. The method for generating the nuclear male sterile line by editing the specific target of the GmNACK2 by using the gene editing CRISPR-Cas9 technology provides a method basis for breeding new soybean varieties, and has an important role in soybean genetic improvement in order to improve the soybean yield.
Drawings
FIG. 1 is a comparison of natural male sterile and fertile plants of soybean of example 1, wherein FIG. 1A is a comparison of plant types and pod bearing of sterile and fertile plants, the sterile plant showing pod abortion and normal pod bearing of fertile plant, and FIG. 1A is an enlarged view of the pod bearing part of sterile plant; FIG. 1B is a comparison of pollen dispersal characteristics of sterile and fertile plants, where the sterile plants show pollen abortion and failure to disperse pollen, while fertile plants normally disperse pollen; FIG. 1C shows the contrast of the morphology of the sterile anthers with the fertile plants under the body mirror, the sterile plants are smooth and no pollen is produced, while the fertile plants have pollen.
FIG. 2 shows the verification of the obtained transgenic soybean positive plant, wherein WT represents a wild type plant, and it can be seen that compared with wild type WT, no anther is generated between the soybean stigma ms1-Cas9 and mutant ms1 after gene editing.
FIG. 3 shows the result of the mutant verification of the transgenic positive plant (ms1-Cas9) transformed with Vector-GmNACK2 in the invention, and the editing effect of the gene can be seen from the figure.
Detailed Description
For the convenience of understanding, the technical scheme of the invention is described in more detail in the following with reference to the attached drawings:
as shown in figure 1, 1 natural soybean male sterile plant found in the field at the early stage is utilized to be hybridized with other soybean varieties in 2013, so that a sterile population is constructed. And planting and harvesting hybrid seeds in 2014. The harvested seeds of generation F1 were then planted at experimental sites to obtain population F2. The population is continuously propagated in 2016 and 2019 and planted in a high and new technology industrial park of agriculture university in Anhui province.
In this example, genetic analysis was performed on the F2 population, shape survey and statistics were performed on 892 soybean plants in the F2 sterile population, and it was found that 694 were the sterile plants and 198 were fertile plants, and the data obtained by the SPSS12.0 software were subjected to chi-square test with the segregation ratio of 3: 1, as shown in Table 1, χ2 3:1<χ2 0.053.841, indicating that the sterile trait of the population is controlled by a single recessive nuclear gene; the field character survey result shows that the character segregation ratio in the backcross progeny is 1: 1. From the data of the field population segregation condition investigated above, it is indicated that the fertility of the male sterile mutant is controlled by 1 pair of recessive nuclear genes.
TABLE 1F2Population genetics analysis
The results of the re-sequencing analysis of the sterile individuals and the fertile individuals show that the sterile individuals have a structural variation on chromosome 13, the variation causes the deletion of a chromosome fragment of chromosome 13 with 38.7kb (22776268-.
The method for creating the soybean nuclear male sterile line by using the CRISPR/Cas9 gene editing technology and taking the primer sequence as a gene editing sequence comprises the following steps:
s1, constructing a gene editing sequence described by SEQ ID NO.1 on a knockout expression Vector pMDC123, and naming the knockout Vector as Vector-GmNACK 2;
s2, editing a Vector-GmNACK2, introducing an allelopathic strain, and performing soybean genetic transformation of direct organ regeneration by using a method of mediating soybean cotyledon node transformation by the allelopathic strain to obtain a transgenic soybean positive plant;
the competent strain of the application uses agrobacterium GV3101, and the cotyledonary node is converted into a conversion method in the prior art, and the specific steps are as follows:
s11, agrobacterium detection: marking resistance YEP plate by agrobacterium, detecting agrobacterium activity, resistance, whether pollution and carrier resistance, selecting successfully expressed colony for PCR identification, and storing in-80 deg.C refrigerator after detecting without error.
S12, activating and culturing agrobacterium: taking required strains (containing herbicide-resistant plant expression Vector1) in a refrigerator at-80 ℃, scratching inoculated ring picking strains on a YEP flat plate, inverting the flat plate, and culturing in the dark at 28 ℃ for 2 days; 2-4 cultured single colonies are selected and inoculated in 10mL YEP liquid culture medium, proper antibiotics are added into the culture medium, and shaking culture is carried out at 28 ℃ and 250rpm for 16h overnight; taking the once activated bacterial liquid, mixing the once activated bacterial liquid with the ratio of 1: 1000 adding fresh YEP liquid culture medium, continuing to culture at 28 ℃ and 250rpm for 16h with shaking, and detecting the bacterial concentration in the culture medium until OD is 1.0-1.2.
S13, soybean explant preparation: selecting disease-free and crack-free plump soybeans, performing moist heat sterilization treatment in a laboratory, inoculating the soybeans in a germination culture medium, and germinating for 1-5 days at 26 ℃ in a dark place.
S14, explant agrobacterium infection and co-culture: cutting the soybean cotyledon node explant, and then putting the cut soybean cotyledon node explant into a culture medium containing an agrobacterium tumefaciens resuspension for dip dyeing; after infection, the explants are inoculated in a co-culture medium with filter paper spread on the surface for dark culture for 3-5 d.
S15, recovery culture: after co-cultivation, cotyledons were inserted into recovery medium for 5-10 days.
S16, bud induction culture: after the recovery culture, transferring the explant to a bud induction solid culture medium, inoculating the explant on the culture medium with an upward incision according to the growth vigor of the explant for 7-10 days, and transferring the explant to a bud induction culture medium containing 6-8mg/l of glufosinate-ammonium for one time for 3 weeks.
S17, bud elongation culture: after the bud induction is completed, cluster buds are left, inoculated into a bud elongation culture medium, and cultured for 3 weeks for subculture once.
S18, rooting culture: when the 3-4cm caulicles of the cluster buds are extracted, the cluster buds are cut off by being close to the base part of the caulicles and are inoculated into a rooting culture medium for 3 weeks of subculture.
S19, hardening and transplanting seedlings: transplanting the explant into a plastic pot of nutrient soil after the explant grows roots, covering and moisturizing the explant by using a transparent plastic bag after the explant is watered thoroughly, and moving the regenerated plant leaves into a greenhouse for management after the regenerated plant leaves are free from fog drops and bag withdrawing; the soybean strain after hardening off can be collected for molecular detection.
S3, detecting a mutant sequencing result: performing mutant verification on a transgenic positive plant (ms1-Cas9) transformed with Vector-GmNACK2, performing PCR sequencing, and performing monoclonal sequencing. As shown in FIG. 2, the sequencing result shows that gene editing occurs at 1bp and 4bp upstream of the PAM target, a single G base is added, TC 2 bases are deleted, and the sequencing peak graph shows different-height set peaks near the target, which indicates that the mutant generates the editing effect on the gene.
The detection primers for the PCR sequencing and the monoclonal sequencing are as follows:
forward sequence: TCGGAAGAACATGGCTTAGCTATTA
Reverse sequence: ACCGAAGGTGTATGGTGTTGTA
As shown in figure 3, compared with wild type WT, no anther is generated in the soybean stigma ms1-Cas9 and mutant ms1 after gene editing, and the gene editing sequence provided by the application can play a gene editing effect, so that the soybean male sterile line can be effectively created by the gene editing target.
The above embodiments are only used to illustrate the technical solutions of the present invention, and do not limit the present invention; although the present invention has been described in detail with reference to the foregoing embodiments, it will be understood by those skilled in the art that: any modification, equivalent replacement, and improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.
Sequence listing
<110> agriculture university of Anhui
<120> gene editing sequence and method for creating soybean nucleus male sterile line
<141> 2021-07-30
<160> 3
<170> SIPOSequenceListing 1.0
<210> 1
<211> 23
<212> DNA
<213> Glycine max
<400> 1
gggtggcccc aaagttcggg agg 23
<210> 2
<211> 25
<212> DNA
<213> Glycine max
<400> 2
tcggaagaac atggcttagc tatta 25
<210> 3
<211> 22
<212> DNA
<213> Glycine max
<400> 3
accgaaggtg tatggtgttg ta 22
Claims (3)
1. A gene editing sequence for creating a soybean plant cell nucleus male sterile line by using a gene editing technology CRISPR/Cas9 is characterized in that the gene editing sequence is sgRNA of a soybean plant cell nucleus male sterile gene target spot, and the nucleotide sequence of the sgRNA is shown in SEQ ID No. 1.
2. A method of creating a nuclear male sterile line of a soybean plant using the gene editing sequence of claim 1, comprising the steps of:
s1, constructing a gene editing sequence described by SEQ ID NO.1 on a knockout expression Vector pMDC123, and naming the knockout Vector as Vector-GmNACK 2;
s2, editing a Vector-GmNACK2, introducing an allelopathic strain, and performing soybean genetic transformation of direct organ regeneration by using a method of mediating soybean cotyledon node transformation by the allelopathic strain to obtain a transgenic soybean positive plant;
s3, detecting a mutant sequencing result, namely performing mutant verification on a transgenic positive plant of the transformed Vector-GmNACK2, performing PCR sequencing, and preliminarily judging that the gene is a mutant if a set peak appears at the sgRNA; performing monoclonal sequencing on the plant which is preliminarily judged as the mutant, and performing final judgment according to a sequencing result; the forward sequence of the primer used for PCR sequencing and monoclonal sequencing is shown as SEQ ID NO.2, and the reverse sequence is shown as SEQ ID NO. 3.
3. The method of claim 2, wherein the competent strain is Agrobacterium GV 3101.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2021104269816 | 2021-04-20 | ||
| CN202110426981 | 2021-04-20 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN114164208A true CN114164208A (en) | 2022-03-11 |
Family
ID=80476477
Family Applications (2)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202110871824.6A Pending CN114164208A (en) | 2021-04-20 | 2021-07-30 | Gene editing sequence and method for creating soybean cell nucleus male sterile line |
| CN202110871852.8A Pending CN114164289A (en) | 2021-04-20 | 2021-07-30 | Soybean nuclear male sterility InDel marker and application thereof |
Family Applications After (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202110871852.8A Pending CN114164289A (en) | 2021-04-20 | 2021-07-30 | Soybean nuclear male sterility InDel marker and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (2) | CN114164208A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112391395A (en) * | 2020-12-03 | 2021-02-23 | 吉林省农业科学院 | Soybean sterile gene mutant, application and construction method of soybean sterile line |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101437843A (en) * | 2006-01-23 | 2009-05-20 | 密歇根州立大学评议会 | Methods for breeding glyphosate resistant plants and compositions thereof |
| WO2020069241A1 (en) * | 2018-09-27 | 2020-04-02 | University Of Tennessee Research Foundation | Discovery of soybean cyst nematode resistance genes based on epigenetic analysis |
| CN111574602A (en) * | 2020-05-14 | 2020-08-25 | 中国科学院东北地理与农业生态研究所 | GmAMS1 protein, coding gene and inhibitor thereof, and method for creating plant nuclear male sterile line |
| CN112391395A (en) * | 2020-12-03 | 2021-02-23 | 吉林省农业科学院 | Soybean sterile gene mutant, application and construction method of soybean sterile line |
Family Cites Families (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103160599B (en) * | 2013-04-07 | 2014-08-13 | 吉林省农业科学院 | Method for identifying soybean cytoplasmic male sterile line seed purity through molecular marker |
| CN104131012B (en) * | 2014-07-10 | 2017-02-08 | 安徽农业大学 | Molecular marker for identifying soybean nucleus male sterility line and identification method thereof |
| CN109042299B (en) * | 2018-08-23 | 2020-05-05 | 河北省农林科学院粮油作物研究所 | Stable close linkage marker for soybean sterility gene |
| CN112609016B (en) * | 2019-10-22 | 2022-11-22 | 中国农业科学院作物科学研究所 | Method for identifying or assisting in identifying whether soybean contains male sterile gene or not and application of method |
-
2021
- 2021-07-30 CN CN202110871824.6A patent/CN114164208A/en active Pending
- 2021-07-30 CN CN202110871852.8A patent/CN114164289A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101437843A (en) * | 2006-01-23 | 2009-05-20 | 密歇根州立大学评议会 | Methods for breeding glyphosate resistant plants and compositions thereof |
| WO2020069241A1 (en) * | 2018-09-27 | 2020-04-02 | University Of Tennessee Research Foundation | Discovery of soybean cyst nematode resistance genes based on epigenetic analysis |
| CN111574602A (en) * | 2020-05-14 | 2020-08-25 | 中国科学院东北地理与农业生态研究所 | GmAMS1 protein, coding gene and inhibitor thereof, and method for creating plant nuclear male sterile line |
| CN112391395A (en) * | 2020-12-03 | 2021-02-23 | 吉林省农业科学院 | Soybean sterile gene mutant, application and construction method of soybean sterile line |
Non-Patent Citations (1)
| Title |
|---|
| BINGJUN JIANG ET AL.: "The cloning and CRISPR/Cas9-mediated mutagenesis of a male sterility gene MS1 of soybean", PLANT BIOTECHNOLOGY JOURNAL, vol. 19, pages 1098 - 1099 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112391395A (en) * | 2020-12-03 | 2021-02-23 | 吉林省农业科学院 | Soybean sterile gene mutant, application and construction method of soybean sterile line |
Also Published As
| Publication number | Publication date |
|---|---|
| CN114164289A (en) | 2022-03-11 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN101528934B (en) | Methods for producing transgenic plants | |
| CN107988229A (en) | A kind of method for obtaining the rice that tiller changes using CRISPR-Cas modification OsTAC1 genes | |
| CN105821074B (en) | Application of rice temperature-sensitive male sterility gene TMS10 and fertility restoration method | |
| US20210269816A1 (en) | Method of Obtaining Multileaflet Medicago Sativa Materials by Means of MsPALM1 Artificial Site-Directed Mutants | |
| WO2022135246A1 (en) | R gene for controlling matching of soybean-rhizobium, protein and use thereof | |
| CN104004781A (en) | Preparation method of glyphosate resistant transgenic rice | |
| CN105087641A (en) | Novel agrobacterium mediated brassica pekinensis in-situ transgenosis method | |
| CN113604497B (en) | Genetic transformation method of gramineous plants | |
| CN110408652A (en) | A method of based on CRISPR/Cas9 system to Malus sieversii gene multiple target point rite-directed mutagenesis | |
| CN114164208A (en) | Gene editing sequence and method for creating soybean cell nucleus male sterile line | |
| CN114150000B (en) | Application of rape BnLAC2 gene in improving cold resistance and early flowering | |
| CN107338231A (en) | The application of OsMPK21-1 albumen and its encoding gene in plant drought resistance is regulated and controled | |
| CN105968178B (en) | The application of rice Os RAD1 albumen and its encoding gene in regulation pollen fertility | |
| CN113699165A (en) | Nucleic acid for reducing height of corn strain and application thereof | |
| Webb et al. | Expression of GUS in primary transformants and segregation patterns of GUS, TL-and TR-DNA in the T1 generation of hairy root transformants of Lotus corniculatus | |
| CN100469886C (en) | Non-embryo transgenic process for preparing transgenic cotton | |
| CN112889668A (en) | Populus genetic transformation method | |
| CN1125878C (en) | Method for creating transgenic receptor system of corn and application of same | |
| CN104017823A (en) | Screening method for fast and effectively reducing false positive rate of peanut genetic transformation plant | |
| CN117604027B (en) | Novel efficient genetic transformation method for pineapple | |
| CN1589611A (en) | Method of inducing helero category anti powdery mildew gene into wheat | |
| CN115125252A (en) | Application and method of CsSH1 gene in improvement of excessive growth resistance of cucumber in seedling stage | |
| CN107475283A (en) | A kind of corn gene method of the agriculture bacillus mediated pollen of ultrasonic assistant | |
| CN111778275A (en) | Application and method of fast-cycle rape in rape functional gene research field | |
| CN117821473A (en) | Main gene GhFBA1 of cotton branch included angle and application thereof in regulation and control of cotton plant type |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination |