CN107217070A - One kind is based on TALENs gene editing peanut breeding methods - Google Patents
One kind is based on TALENs gene editing peanut breeding methods Download PDFInfo
- Publication number
- CN107217070A CN107217070A CN201710363687.9A CN201710363687A CN107217070A CN 107217070 A CN107217070 A CN 107217070A CN 201710363687 A CN201710363687 A CN 201710363687A CN 107217070 A CN107217070 A CN 107217070A
- Authority
- CN
- China
- Prior art keywords
- sequence
- peanut
- seed
- right arms
- oleic acid
- 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
Classifications
-
- 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
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/0004—Oxidoreductases (1.)
- C12N9/0071—Oxidoreductases (1.) acting on paired donors with incorporation of molecular oxygen (1.14)
- C12N9/0083—Miscellaneous (1.14.99)
-
- 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
- C12N15/8242—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits
- C12N15/8243—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits involving biosynthetic or metabolic pathways, i.e. metabolic engineering, e.g. nicotine, caffeine
- C12N15/8247—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits involving biosynthetic or metabolic pathways, i.e. metabolic engineering, e.g. nicotine, caffeine involving modified lipid metabolism, e.g. seed oil composition
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y114/00—Oxidoreductases acting on paired donors, with incorporation or reduction of molecular oxygen (1.14)
- C12Y114/99—Miscellaneous (1.14.99)
- C12Y114/99033—DELTA12-fatty acid dehydrogenase (1.14.99.33)
-
- 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
- C12N2800/00—Nucleic acids vectors
- C12N2800/80—Vectors containing sites for inducing double-stranded breaks, e.g. meganuclease restriction sites
Abstract
The present invention discloses one kind and is based on TALENs gene editing peanut breeding methods, comprises the following steps：1) design of TALENs left and right arms sequence；2) it is assembled to expression vector；3) genetic transformation and 4) seed selection：According to objective trait seed selection.The TALENs left and right arms sequence pair peanut gene editings that the present invention is obtained, obtained peanut seed oleic acid content is high, and the high oleic acid content heritability is stable.
Description
Technical field
TALENs gene editing peanut breeding methods are based on the present invention relates to one kind, belong to agro-biological engineering technology neck
Domain.
Background technology
Peanut also known as peanut, belong to annual herb plant, are the important oil plant of China and industrial crops.The peanut of China
Cultivated area and production are at the forefront in the world to have very strong competitiveness in the world.Oleic acid is main in peanut grease
Aliphatic acid because oleic acid has preferable oxidation stability, so as to reduce lipid oxidation, thus solve (Groundnut products) in production and
The problems such as bad smell, taste and reduced shelf-life for being produced during storage due to lipid oxidation.Oleic acid or a kind of list
Unrighted acid can reduce body low-density lipoprotein (LDL) level, maintain HDL (HDL) level.So high
Oleic acid peanut not only can effectively extend shelf-life and the shelf life of (Groundnut products), while being also advantageous for the healthy of people.
Breeding High-oil acid new peanut variety is one of important goal of peanut breeding.Being currently known FAD2 (fatty acid dehydrogenase of Δ 12) is
Oleic acid is catalyzed in fatty acid biosynthetic pathway in the 12nd carbon potential dehydrogenation, the key enzyme that double bond changes to linoleic acid base is formed.Pass
System breeding strategy is the high oleic acid come out by natural mutant (F435 that such as U.S. is screened) or by mutagenesis screening
Mutant carry out it is miscellaneous become, backcross improvement, it is low that these breeding methods all deposit big breeding efficiency, the shortcomings of time is longer.
Peanut is G at 448bp after FAD2A gene start codons in allotetraploid plant, high oleic acid peanut:C→A：T
Replacement cause the variation of D150N amino acid, 442bp places A is inserted after FAD2B gene start codons so that terminator codon
Occur in advance, so that the expression reduction of fad2 genes or function are lost.Traditional cross breeding method needs to be difficult to accomplish
Precisely mutation, and wasting time and energy.
The content of the invention
In order to overcome the deficiencies in the prior art, it is an object of the invention to provide it is a kind of improve arachic acid based on
TALENs gene editing peanut breeding methods.
Realize that the purpose of the present invention can reach by adopting the following technical scheme that：
One kind is based on TALENs gene editing peanut breeding methods, comprises the following steps：
1) design of TALENs left and right arms sequence：The sequence of the fad2 genes of peanut varieties is analyzed, and designs left and right arms sequence
Row；
2) it is assembled to expression vector：By step 1) obtained left and right arms sequence assembling is to plant expression vector and transfects agriculture bar
Bacterium, obtains the Agrobacterium containing conversion plasmid；
3) genetic transformation：By step 2) the obtained Agrobacterium genetic transformation containing conversion plasmid is to peanut seed；
4) seed selection：According to objective trait seed selection.
Preferably, step 1) in, left arm sequence is SEQ ID No.1 in the left and right arms sequence, and right arm sequence is SEQ
ID No.2。
Preferably, step 1) in, left arm sequence is SEQ ID No.3 in the left and right arms sequence, and right arm sequence is SEQ
ID No.4。
Preferably, step 2) in, the expression vector is pCAMBIA3301 or pCAMBIA1301.
Preferably, step 3) in, the concrete operations of genetic transformation are：
A) explant is made with peanut seed；
B) will the Agrobacterium inoculation containing conversion plasmid to YEP medium cultures, collect thalline, by thalline be resuspended in containing
In the liquid MS medium of acetosyringone, obtain infecting liquid；
C) explant leaching is floated on and infects liquid, co-cultured, explant after being co-cultured.
Preferably, in step c), before co-cultivation, to the silweet L-77 for infecting liquid addition 0.1wt%.
Preferably, step 4) in, the objective trait is that oleic acid content is more than 60%.
The formulation Design Principle of the present invention is as follows：
The present invention carries out the fad2 gene orders of issued peanut in ncbi database and peanut wild species gene pool
Compare, in order that obtaining fad2 genes loses function, while fad2A and fad2B SNP site are avoided, in fad2 gene orders
Near initiation codon, the recognition sequence of following several or so the two-arm of design, is found optimal by way of combination of two altogether
Gene editing.
Wherein, wherein L1R1 spacing distance is 13bp, and L1R2 spacing distance is 14bp, and L2R1 spacing distance is
16bp, L2R2 spacing distance are 17bp, and L3R1 spacing distance is 14bp, and L3R2 spacing distance is 15bp.
The present invention carries out the fad2 gene orders of issued peanut in ncbi database and peanut wild species gene pool
Compare, in order that obtaining fad2 genes loses function, while fad2A and fad2B SNP site are avoided, in fad2 gene orders
Near initiation codon, the recognition sequence of following several or so the two-arm of design, is found optimal by way of combination of two altogether
Gene editing.
Compared with prior art, the beneficial effects of the present invention are：Present invention offer one kind can effectively improve arachic acid and contain
The gene editing method of amount, the character of the high oleic acid content can be stably hereditary to filial generation.
Brief description of the drawings
Fig. 1 is that L1R1 is the carrier editor Guangdong oil T0 of No. 7 for seed oleic acid content.
Fig. 2 is that L2R2 is the carrier editor Guangdong oil T0 of No. 7 for seed oleic acid content.
Embodiment
Below, with reference to accompanying drawing and embodiment, the present invention is described further：
Embodiment 1：The design of TALENs left and right arms genes
The present invention carries out the fad2 gene orders of issued peanut in ncbi database and peanut wild species gene pool
Compare, in order that obtaining fad2 genes loses function, while fad2A and fad2B SNP site are avoided, in fad2 gene orders
Near initiation codon, the recognition sequence of following three left arm genes and two right arms is designed altogether, as shown in SEQ ID No.1
L1, the L2 as shown in SEQ ID No.3, the L3 as shown in SEQ ID No.5, R1 and such as SEQ as shown in SEQ ID No.2
R2 shown in ID No.4.
L1：CAAAAGAAGCCTCTTT
L2：GAAGCTCAAAAGAAGCCTC
L3：CAAAAGAAGCCTCTT
R1：TTCAAACCCTCCATTC
R2：TCAAACCCTCCATTC
Optimal gene editing is found by way of combination of two again.The spacing distance that its L1 and R1 are combined into L1R1 is
The spacing distance that 13bp, L1 and R2 are combined into L1R2 is that the spacing distance that 14bp, L2 and R1 are combined into L2R1 is 16bp, L2 and R2
The spacing distance for being combined into L2R2 is that the spacing distance into L3R1 of 17bp, L3 and R1 combination is that 14bp, L3 and R2 are combined into
L3R2 spacing distance is 15bp.
Embodiment 2：It is cloned into expression vector
By the TALEN kit kits of Si Dansai companies, the TALENs left and right arms sequences L1R1 that embodiment 1 is obtained,
L1R2, L2R1, L3R1 and L3R2 are assembled respectively, then are subcloned respectively to plant expression vector, and transfect Agrobacterium, are selected
Positive strain, obtains the Agrobacterium containing conversion plasmid.
Embodiment 3：Genetic transformation
A) explant is made with peanut seed：Oily No. 7 peanut seeds in Guangdong are taken, peanut seed is numbered, control is set
Group # is wt, and peanut seed is shelled, with distilled water flushing, then immersion 30-60 minutes in distilled water；Then sterilize, go
Skin is planted, is cut seed along gap between cotyledon, takes the half granule seed containing embryo, then plumule is cut along the cotyledon node of embryo, obtains waiting to turn
The explant of change；
B) by embodiment 2) the obtained Agrobacterium inoculation containing plasmid to be transformed is into YEP liquid mediums, and 28 DEG C are shaken
Swing overnight incubation；Agrobacterium thalline is collected by centrifugation in YEP liquid mediums, and Agrobacterium thalline is resuspended in containing acetyl cloves
In the liquid MS medium of ketone, obtain infecting liquid；
C) infected what the explant leaching to be transformed that step a) is obtained floated on that step b) obtains in liquid, add 0.1wt%
Silweet L-77, infect 10-20 minutes, then by explant and infect liquid and stand 10-20 minutes under vacuum；Take
Go out explant, be filtered dry after bacterium solution and be laid in co-cultivation culture medium, dark culturing 2-3 days；Trained again under conditions of illumination 16h/d
Support 7-10 days, the explant after being co-cultured；
Shown in the situation of change following table of the sequence of the cog region of explant after co-cultivation,
The L1R1 of table 1 edits the sequence variation situation of seed cog region
The L2R2 of table 2 edits the sequence variation situation of seed cog region
D) explant after co-cultivation is moved in sterilizing sand, after culture domestication, moves into greenhouse and breed.
Embodiment 4：Oleic acid is detected
The T0 of the explant of embodiment 3 is harvested for seed, the oleic acid content for the explant seed that embodiment 3 is obtained is detected, greatly
It is considered as positive strain in 60%.The conversion strain number of each TALENs left and right arms, positive strain number and frequency statistics are as shown in the table：
The seed selection result of table 3 is counted
Oleic acid content detection is carried out for seed to T0 using non-damage type infrared radiation detection apparatus, as a result shown in Fig. 1 and 2, passed through
L1R1 and L2R2 enter the peanut that edlin FAD2 genes can obtain high oleic acid, wherein, L2R2 conversion ratio is high compared with L1R1, this
May be relevant with the spacing distance of left and right arms, in peanut, left and right arms spacing distance is detrimental to shearing in 14-16bp distance
Enzyme formation dimeric structure, so that the FAD2 genes of peanut can not be edited, and between effect ratio of the spacing distance for 17bp L2R2
Gauge will get well from the L1R1 for 13bp, wherein, No. 314 oleic acid contents are up to 90%.
Embodiment 5：Seed selection
T0 of the oleic acid content that embodiment 4 is detected higher than 60% is planted for seed, is obtained T1 for seed, is utilized
The detection and analysis result that the near infrared detection instrument of non-damage type carries out oleic acid content is as follows：
The T1 of table 4 is for seed oleic acid content
From the result of upper table, T1 for seed oleic acid content compared with T0 generations, do not reduce, or even be higher than T0
For seed, illustrate, the breeding that the method provided by the present invention is carried out, the character of its high oleic acid content can be stably hereditary.
Embodiment 6：Checking
By embodiment 2 obtain containing L1R1 left and right arms sequences conversion plasmid Agrobacterium and containing L2R2 left and right arms
The conversion plasmid of sequence.Gene editing operation is carried out to Guangdong No. 13 peanut seeds of oil, its T0 is for shown in seed oleic acid content following table：
The Guangdong of table 5 No. 13 T0 of oil are for seed oleic acid content
| Sample number | Left and right arms sequence pair | Oleic acid [%] |
| WT | Nothing | 30.94 |
| 1 | L1R1 | 62.78 |
| 5 | L1R1 | 63.31 |
| 8 | L1R1 | 72.91 |
| 12 | L1R1 | 61.08 |
| 11 | L1R1 | 69.19 |
| 13 | L1R1 | 67.28 |
| 21 | L2R2 | 71.98 |
| 99 | L2R2 | 62.31 |
T0 is subjected to genetic breeding for seed, T1 is obtained for seed, its oleic acid content is as shown in the table：
The Guangdong of table 6 oil 13T1 is for seed oleic acid content
| Sample number | Oleic acid [%] | Sample number | Oleic acid [%] | Sample number | Oleic acid [%] | Sample number | Oleic acid [%] |
| 1-1 | 63.21 | 8-1 | 72.21 | 11-2 | 70.32 | 13-3 | 66.83 |
| 1-2 | 62.32 | 8-2 | 72.33 | 11-3 | 69.32 | 21-1 | 69.78 |
| 1-3 | 63.24 | 8-3 | 72.35 | 11-4 | 68.89 | 21-2 | 68.98 |
| 1-4 | 62.87 | 12-1 | 60.92 | 11-5 | 71.23 | 99-1 | 64.22 |
| 5-1 | 63.11 | 12-2 | 61.32 | 11-6 | 69.22 | 99-2 | 63.78 |
| 5-2 | 63.42 | 12-3 | 62.21 | 13-1 | 66.23 | 99-3 | 63.23 |
| 5-3 | 63.23 | 11-1 | 70.12 | 13-2 | 67.34 | 99-4 | 62.43 |
T1 is subjected to further tracing detection for the strain of seed numbering 8, its T2 carries out near infrared spectrometer for seed
Testing result it is as shown in the table：
The Guangdong of table 7 oil 13T2 is for seed oleic acid content
| Sample number | Oleic acid [%] | Sample number | Oleic acid [%] | Sample number | Oleic acid [%] | Sample number | Oleic acid [%] |
| 8-1-1 | 72.23 | 8-1-7 | 71.76 | 8-2-6 | 70.82 | 8-3-3 | 71.20 |
| 8-1-2 | 71.98 | 8-1-8 | 71.89 | 8-2-7 | 71.10 | 8-3-4 | 71.10 |
| 8-1-3 | 72.32 | 8-2-1 | 72.13 | 8-2-8 | 73.19 | 8-3-5 | 70.98 |
| 8-1-4 | 72.42 | 8-2-3 | 72.33 | 8-2-9 | 72.31 | 8-3-6 | 71.34 |
| 8-1-5 | 73.11 | 8-2-4 | 72.11 | 8-3-1 | 72.24 | 8-3-7 | 69.68 |
| 8-1-6 | 72.45 | 8-2-5 | 72.14 | 8-3-2 | 72.43 | 8-3-8 | 70.88 |
From above checking test, L1R1 or L2R2 are as TALENs left and right arms sequences, and it can make the oil of peanut seed
Acid content significantly increases, and the high oleic acid character has preferable genetic stability.
For those skilled in the art, technical scheme that can be as described above and design, make other each
It is kind corresponding to change and deform, and all these change and deformation should all belong to the protection model of the claims in the present invention
Within enclosing.
SEQUENCE LISTING
<110>Crop Institute, Guangdong Academy of Agricultural Sciences
<120>One kind is based on TALENS gene editing peanut breeding methods
<130>One kind is based on TALENS gene editing peanut breeding methods
<160> 5
<170> PatentIn version 3.3
<210> 1
<211> 16
<212> DNA
<213>Artificial sequence
<400> 1
caaaagaagc ctcttt 16
<210> 2
<211> 16
<212> DNA
<213>Artificial sequence
<400> 2
ttcaaaccct ccattc 16
<210> 3
<211> 19
<212> DNA
<213>Artificial sequence
<400> 3
gaagctcaaa agaagcctc 19
<210> 4
<211> 15
<212> DNA
<213>Artificial sequence
<400> 4
tcaaaccctc cattc 15
<210> 5
<211> 15
<212> DNA
<213>Artificial sequence
<400> 5
caaaagaagc ctctt 15
Claims (7)
1. one kind is based on TALENs gene editing peanut breeding methods, it is characterised in that comprise the following steps：
1) design of TALENs left and right arms sequence：The sequence of the fad2 genes of peanut varieties is analyzed, and designs left and right arms sequence；
2) it is assembled to expression vector：By step 1) obtained left and right arms sequence assembling is to plant expression vector and transfects Agrobacterium,
Obtain the Agrobacterium containing conversion plasmid；
3) genetic transformation：By step 2) the obtained Agrobacterium genetic transformation containing conversion plasmid is to peanut seed；
4) seed selection：According to objective trait seed selection.
2. according to the method described in claim 1, it is characterised in that step 1), left arm sequence is in the left and right arms sequence
SEQ ID No.1, right arm sequence is SEQ ID No.2.
3. according to the method described in claim 1, it is characterised in that step 1), left arm sequence is in the left and right arms sequence
SEQ ID No.3, right arm sequence is SEQ ID No.4.
4. according to the method described in claim 1, it is characterised in that step 2), the expression vector be pCAMBIA3301 or
pCAMBIA1301。
5. according to the method described in claim 1, it is characterised in that step 3), the concrete operations of genetic transformation are：
A) explant is made with peanut seed；
B) Agrobacterium inoculation containing conversion plasmid is collected thalline, thalline is resuspended in containing acetyl to YEP medium cultures
In the liquid MS medium of syringone, obtain infecting liquid；
C) explant leaching is floated on and infects liquid, co-cultured, explant after being co-cultured.
6. method according to claim 5, it is characterised in that in step c), before co-cultivation, adds 0.1wt%'s to liquid is infected
silweet L-77。
7. according to the method described in claim 1, it is characterised in that step 4), the objective trait is more than for oleic acid content
60%.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710363687.9A CN107217070A (en) | 2017-05-22 | 2017-05-22 | One kind is based on TALENs gene editing peanut breeding methods |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710363687.9A CN107217070A (en) | 2017-05-22 | 2017-05-22 | One kind is based on TALENs gene editing peanut breeding methods |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN107217070A true CN107217070A (en) | 2017-09-29 |
Family
ID=59944238
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201710363687.9A Pending CN107217070A (en) | 2017-05-22 | 2017-05-22 | One kind is based on TALENs gene editing peanut breeding methods |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN107217070A (en) |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1724669A (en) * | 2005-03-17 | 2006-01-25 | 东北师范大学 | Method in increasing oleic acid content of soybean and peanut seed by applying gene silent technology |
| CN101445808A (en) * | 2008-11-24 | 2009-06-03 | 广东省农业科学院作物研究所 | Agrobacterium-mediated genetic transformation method with peanut seed domant bud hypocotyl as explant |
| CN102191269A (en) * | 2011-03-28 | 2011-09-21 | 广东省农业科学院作物研究所 | Non tissue culture gene transferring method by using half of peanut seed as acceptor |
| CN104780756A (en) * | 2012-09-07 | 2015-07-15 | 美国陶氏益农公司 | Fad2 performance loci and corresponding target site specific binding proteins capable of inducing targeted breaks |
-
2017
- 2017-05-22 CN CN201710363687.9A patent/CN107217070A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1724669A (en) * | 2005-03-17 | 2006-01-25 | 东北师范大学 | Method in increasing oleic acid content of soybean and peanut seed by applying gene silent technology |
| CN101445808A (en) * | 2008-11-24 | 2009-06-03 | 广东省农业科学院作物研究所 | Agrobacterium-mediated genetic transformation method with peanut seed domant bud hypocotyl as explant |
| CN102191269A (en) * | 2011-03-28 | 2011-09-21 | 广东省农业科学院作物研究所 | Non tissue culture gene transferring method by using half of peanut seed as acceptor |
| CN104780756A (en) * | 2012-09-07 | 2015-07-15 | 美国陶氏益农公司 | Fad2 performance loci and corresponding target site specific binding proteins capable of inducing targeted breaks |
Non-Patent Citations (3)
| Title |
|---|
| SHIJIE WEN等: "TALEN-mediated targeted mutagenesis of fatty acid desaturase 2 (FAD2) in peanut (Arachis hypogaea L.) promotes the accumulation of oleic acid", 《PLANT MOLECULAR BIOLOGY》 * |
| WILLIAM HAUN等: "Improved soybean oil quality by targeted mutagenesis of the fatty acid desaturase 2 gene family", 《PLANT BIOTECHNOLOGY JOURNAL》 * |
| 单奇伟等: "植物基因组编辑及衍生技术最新研究进展", 《遗传》 * |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Hoffmann et al. | Anther culture as a breeding tool in rape: II. Progeny analyses of androgenetic lines and induced mutants from haploid cultures | |
| Rieseberg et al. | Molecular evidence and the origin and development of the domesticated sunflower (Helianthus annum, Asteraceae) | |
| CN106544357A (en) | A kind of method for cultivating low cadmium-accumulation rice variety | |
| CN105505981B (en) | A kind of resistance glyphosate genetically engineered soybean and the preparation method and application thereof | |
| CN108239674B (en) | Cabbage type rape high oleic acid QTL and molecular marker closely linked with same | |
| CN105210859A (en) | A kind of efficient selection of odor type anti-rice blast rice new varieties | |
| Gamborg et al. | Protoplast and cell culture methods in somatic hybridization in higher plants | |
| Corduan | Regeneration of anther-derived plants of Hyoscyamus niger L. | |
| CN111304237B (en) | High-oil stress-resistant peony breeding method | |
| CN109588309A (en) | A kind of method of quick, efficient Breeding High-oil acid peanut varieties | |
| CN102229950B (en) | Rapid and high-efficiency transgenic method for indica rice | |
| Yemets et al. | Establishment of in vitro culture, plant regeneration, and genetic transformation of Camelina sativa | |
| CN110408652A (en) | A method of based on CRISPR/Cas9 system to Malus sieversii gene multiple target point rite-directed mutagenesis | |
| Grosser et al. | Production of mandarin+ pummelo somatic hybrid citrus rootstocks with potential for improved tolerance/resistance to sting nematode | |
| CN109750061A (en) | One kind overcoming the self-incompatible method of Diploid Potato | |
| Amberger et al. | Analysis of Culture‐Induced Variation in Soybean | |
| Kim et al. | Selection of 5-methyltryptophan resistant rice mutants from irradiated calli derived from embryos | |
| CN107217070A (en) | One kind is based on TALENs gene editing peanut breeding methods | |
| CN105734057B (en) | With the SSR marker and its application of muskmelon downy mildew resistance main effect QTL linkage | |
| CN104593411A (en) | Application of barley Hvsusiba2 genes in reducing release of methane in rice paddies | |
| CN110484559A (en) | The method of BnaC2-GMYB28 gene editing acquisition seed low-sulfur glycosides rape | |
| CN105732786A (en) | Rice gene OsEIN2L and application thereof | |
| CN102816854A (en) | Research method of space mutation mechanism and biogenic mutation molecule reporting model used by same | |
| CN104855278A (en) | Breeding method capable of reducing conversion rate of tobacco nicotine | |
| Tudses et al. | Establishment of method for protoplast fusion with PEG-mediated between Jatropha curcas L. and Ricinus communis L |
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 |