CN112889663A - Method for polymerizing multiple corn one-way cross incompatible genes - Google Patents
Method for polymerizing multiple corn one-way cross incompatible genes Download PDFInfo
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- CN112889663A CN112889663A CN201911223651.6A CN201911223651A CN112889663A CN 112889663 A CN112889663 A CN 112889663A CN 201911223651 A CN201911223651 A CN 201911223651A CN 112889663 A CN112889663 A CN 112889663A
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Abstract
The invention discloses a method for polymerizing a plurality of corn one-way cross incompatible genes, which solves the risk that a single Ga-S gene is used as an isolation tool to be outcrossed and fructified aiming at the difficulty in polymerizing a plurality of different one-way outcross incompatible genes. The invention is characterized in that Ga1-S and Ga2-S are polymerized by utilizing the affinity of F1 of Ga-S gene, through pollination to F1, progeny self-bred and assisted by closely linked molecular marker selection. The invention reduces the risk of outcrossing by using the unidirectional cross incompatible gene for reproductive isolation in the production process of corn seed production, further promotes the reproductive isolation of different types of corn by using the unidirectional cross incompatible gene in production, and saves a large amount of manpower, material resources and financial resources.
Description
Technical Field
The invention belongs to the field of creation of new crop germplasm. In particular to a method for aggregating a plurality of corn one-way cross incompatible genes.
Background
Maize is a typical cross-pollinated crop, and usually can bear pollen of itself or other maize pollen to produce fruit. However, there are some types of corn in nature that break this rule: they only accept the pollen of the same corn species, but not the pollen of non-same corn species, and at the same time, they can pollinate and seed most of the non-same corn species. This phenomenon we call the one-way outcrossing incompatibility of maize. Since it is involved in the transmission of male and female maize gametes, the gene controlling this phenomenon is called the gaphylate factor, i.e., the unilateralized, incompatible Ga gene.
The earliest Ga genes in nature were only present in popcorn, very few Central America maize and maize ancestors, teosintes, and most dent and hard-kernel cultivated maize did not have Ga genes. Therefore, in production, the Ga gene can be transferred to the common cultivated corn by a backcross transfer method, so that the Ga gene becomes a biological isolation tool among different varieties of corn, and plays an important role in the links of corn hybridization seed production isolation, special corn production isolation, transgenic corn production isolation and the like.
At present, although a plurality of Ga genes are found, only 3 Ga genes are regarded as important due to nearly 100% of one-way cross incompatibility, namely Ga1-S, Ga2-S and Tcb1-S, which are incompatible with each other. Subsequent studies have shown that there are few materials in the common cultivated corn, which can penetrate the incompatibility of Ga1-S, Ga2-S and Tcb1-S, respectively, the existence of this type of material limits the potential of using a single Ga gene in corn production, and the problem can be solved by polymerizing different Ga-S genes in the same material. However, since Ga1-S and Ga2-S are not compatible with each other, i.e., they do not hybridize to each other, it is extremely difficult and unprecedented to polymerize Ga1-S and Ga2-S at two sites in one.
Disclosure of Invention
The invention aims to solve the problem that a single Ga-S gene is outcrossed and fruited when being used as an isolation tool aiming at the difficulty of polymerization of a plurality of different unidirectional outcross incompatibility genes. The invention is characterized in that Ga1-S and Ga2-S are polymerized by utilizing the affinity of F1 of Ga-S gene, through pollination to F1, progeny self-bred and assisted by closely linked molecular marker selection.
The technical scheme adopted by the invention for solving the technical problems is as follows: firstly, F1 of Ga1-S and Ga2-S materials are respectively constructed by utilizing a common inbred line (Zheng 58 and the like), and then hybridization treatment is carried out by three methods, namely, mutual hybridization of two F1, hybridization of F1 of Ga1-S and a material with Ga2-S, and hybridization of F1 of Ga2-S and a material with Ga 1-S. And screening out single plants which are double-heterozygous with Ga1-S and Ga2-S simultaneously from progeny obtained after hybridization by corresponding molecular markers of Ga1-S and Ga2-S, then selfing, continuously screening out single plants which are double-homozygous with Ga1-S and Ga2-S from the selfed plants by using the molecular markers, and then carrying out continuous multi-generation selfing on the single plants, wherein during the selfing process of each generation, incompatible identification and screening are respectively carried out on the common selfing lines, Ga1-S and Ga2-S selfing lines to obtain the single plants which are both incompetent. Finally, through continuous multi-generation selfing and selection, a maize inbred line with double homozygosity of Ga1-S and Ga2-S and uniform properties is generated.
Drawings
FIG. 1 shows the breeding process for creating double-heterozygous strain of Ga1-S and Ga 2-S.
FIG. 2 shows the selection of individuals heterozygous for Ga1-S in passage 3 using Ga1-S marker screen ID4, wherein lanes 1-21 are selected individuals and lanes 22, 23, 24 are Zheng 58, 511L and SDGa25, respectively.
FIG. 3 shows individuals screened for heterozygous Ga2-S in passage 3 using Ga2-S marker SG2, wherein lanes 1-21 are screened individuals and lanes 22, 23, 24 are Zheng 58, SDGa25 and 511L, respectively.
FIG. 4 shows individuals selected for homozygous Ga1-S in the 4 th generation using Ga1-S marker ID4, wherein lanes 1-21 are selected individuals and lanes 22, 23, 24 are Zheng 58, 511L and SDGa25, respectively.
FIG. 5 shows individuals selected for homozygous Ga2-S in the 4 th generation using Ga2-S marker SG2, wherein lanes 1-21 are selected individuals and lanes 22, 23, 24 are Zheng 58, SDGa25 and 511L, respectively.
FIG. 6 shows the incompatible identification of double homozygous Ga1-S and Ga2-S individuals in the 4 th generation with common inbred lines, Ga1-S and Ga2-S inbred lines and mixed pollen of Ga1-S and Ga2-S inbred lines.
FIG. 7 shows the incompatible identification of double homozygous Ga1-S and Ga2-S lines in the 9 th generation with common inbred lines, Ga1-S and Ga2-S inbred lines and mixed pollen of Ga1-S and Ga2-S inbred lines.
Detailed Description
The experimental procedures used in the following examples are conventional unless otherwise specified.
Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.
The maize inbred lines SDGa25 (genotype: Ga1-SGa1-Sga2Ga 2) and Zheng 58 (genotype: Ga1Ga1Ga2Ga 2) were collected and stored in the laboratory. Inbred 511L (genotype Ga1Ga1Ga2-SGa 2-S) was maintained by Maize Genetics collaboration storage Center, Co-op ID: 511L; description: 511L Ga2-S (mexicana).
The present invention will be described in detail below with reference to polymerization examples of Ga1-S and Ga 2-S.
1) In the winter of 2013, Hainanle southeast breeding base takes Zheng 58 (with the genotype of Ga1Ga2Ga 2) as the female parent and SDGa25 (with the genotype of Ga1-SGa1-Sga2Ga 2) and 511L (with the genotype of Ga1Ga2-SGa 2-S) as the female parent respectively to obtain corresponding F1.
2) In summer 2014, in Beijing Changping farm, two F1 were crossed.
3) In winter 2014, Hainanle southeast breeding base, individuals of double-hybrid Ga1-S and Ga2-S (the genotypes are Ga1-Sga1Ga2-Sga 2) were identified by using markers of Ga1-S and Ga2-S which are closely linked respectively and selfed (FIG. 2 and FIG. 3). Wherein the mark of Ga1-S close linkage is ID4, and the sequence is as follows: forward primer CCCCTATGATAGAAATGTAGCAC, reverse primer CCCACTTGATGTCACCACC; wherein the mark of Ga2-S close linkage is SG2, and the sequence is: forward primer GTGTGGTGTGGAAATCGTGG, reverse primer ATGTAGGGCGTACAACAATAGGT.
4) In summer 2015, in Beijing Changping farm, single plants of double homozygous Ga1-S and Ga2-S (genotype Ga1-SGa1-SGa2-SGa 2-S) were identified by using markers in close linkage with Ga1-S and Ga2-S, respectively, and selfed (FIG. 4 and FIG. 5).
5) In winter 2015, Hainanle southeast breeding base, each double homozygous line was divided into 4 rows, 2 rows were used for selfing, and the other 2 rows were pollinated to double homozygous single plants with mixed pollen of Zheng 58 (genotype: Ga1Ga1Ga2Ga 2), SDGa25 (genotype: Ga1-SGa1-Sga2Ga 2), 511L (genotype: Ga1Ga2-SGa 2-S) and both SDGa25 and 511L, statistical maturing was performed, and the lines to which Zheng 58, SDGa25 and 511L were not bearing were kept (FIG. 6).
6) In 2016-2019 in summer and winter, in Beijing Changping farm and Hainanle southeast breeding base, 4 double homozygous lines are all subjected to the strategy of (5), 2 lines are subjected to selfing, 2 lines are subjected to incompatible identification, and finally, 9 generations of selfing are performed to obtain 4 lines (with the genotype of Ga1-SGa1-SGa2-SGa 2-S) which are double homozygous and have uniform properties (the genotype is Ga1-SGa1-SGa2-SGa 2-S) (figure 7).
Sequence listing
<110> institute of genetics and developmental biology of Chinese academy of sciences
<120> a method for aggregating multiple corn one-way cross incompatibility genes
<160> 4
<170> SIPOSequenceListing 1.0
<210> 1
<211> 23
<212> DNA
<213> corn (Zea mays)
<400> 1
cccctatgat agaaatgtag cac 23
<210> 2
<211> 19
<212> DNA
<213> corn (Zea mays)
<400> 2
cccacttgat gtcaccacc 19
<210> 3
<211> 20
<212> DNA
<213> corn (Zea mays)
<400> 3
gtgtggtgtg gaaatcgtgg 20
<210> 4
<211> 23
<212> DNA
<213> corn (Zea mays)
<400> 4
atgtagggcg tacaacaata ggt 23
Claims (5)
1. A method for aggregating multiple corn single cross incompatible genes is characterized in that F1 of the corn single cross incompatible genes has affinity, and the possibility of aggregating different single cross incompatible genes is realized by pollinating F1, selfing progeny and assisting in closely linked molecular marker selection.
2. The method of claim 1, wherein the corn one-way cross incompatibility gene is Ga1-S and Ga2-S, or a gene having the same or similar function as Ga1-S/Ga2-S gene.
3. The method according to claim 2, wherein the number of the genes having the same or similar functions as those of Ga1-S/Ga2-S genes polymerized is 2 or more.
4. A primer pair for identifying the molecular marker of claim 1, wherein the nucleotide sequence of the primer pair is as shown in ID4 and SG 4.
5. The molecular marker of claim 1 is applied to identify the maize one-way cross incompatibility gene.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113557955A (en) * | 2021-07-19 | 2021-10-29 | 中国农业大学 | Haploid induction line genetic purification method based on reproductive isolation traits |
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- 2019-12-04 CN CN201911223651.6A patent/CN112889663A/en active Pending
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113557955A (en) * | 2021-07-19 | 2021-10-29 | 中国农业大学 | Haploid induction line genetic purification method based on reproductive isolation traits |
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