Molecular markers for identifying the presence or absence of trichomes in cabbage and their development methods and applications
A technology of molecular markers and cuticles, applied in the field of molecular biology, can solve the problems of low efficiency, large land space, and the gene of wild cabbage cuticles has not been reported, so as to improve selection efficiency, reduce land space, and shorten breeding The effect of years
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Embodiment 1
[0038] A method for developing a molecular marker Bol.trichome-CAPS.02 for identifying the presence or absence of cabbage epidermis of the present invention, comprising the following steps:
[0039] 1) F 1 Generation, F 1 The generation was re-selfed and planted in the field, and 1063 F 2 Generation plants (including 149 clones F 2 population and 914 individual F 2:3 pedigree), in F 2 In the fourth to fifth leaf stages of generation plants, the presence or absence of epidermal hairs was observed with the naked eye, and F 2 Among the generation plants, 778 were hairless and 285 were hairy, conforming to the segregation ratio of 3:1 (chi-square value = 1.628, P > 0.05), indicating that the trichomes of wild cabbage were controlled by a recessive single gene;
[0040] 2) Using 149 clones F 2 The population constructed the genetic linkage map of cabbage (Mei et al., 2013, TAG 126:549-556), and mapped the epidermis gene to a 4.3cM (17.56MB) interval on the C01 chromosome of c...
Embodiment 2
[0046] The application process of a molecular marker Bol.trichome-CAPS.02 for identifying the presence or absence of cabbage epidermis in the breeding of cabbage varieties with different epidermis traits of the present invention, the specific steps are as follows:
[0047] (1) Using the DNA of a leaf of cabbage at the seedling stage as a template, amplifying the molecular marker Bol.trichome-CAPS.02, the primers have the nucleotide sequences shown in SEQ ID No.2 and SEQ ID No.3;
[0048] (2) Each 10 μl of the PCR amplification product was treated with 2.5 U of restriction enzyme HinfI at 37°C for 1 h. -five specific bands of bp, 131-bp and 27-bp;
[0049] (3) if Figure 4 As shown, the cabbage plants with four specific bands of 271-bp, 158-bp, 131-bp and 27-bp are homozygous hairy individuals, and three bands of 271-bp, 185-bp and 131-bp appear Those with bands are homozygous hairless plants, and those with five specific bands at the same time are heterozygous individuals. T...
Embodiment 3
[0051] The application process of a molecular marker Bol.trichome-CAPS.02 for identifying the presence or absence of cabbage epidermis in backcross breeding of the present invention:
[0052] Such as figure 1 As shown, the F 1 Continue to backcross with C41 and get 80 copies of BC 1 f 1 The backcross population was used as the research material, and the backcross breeding research of transferring the hairy trait (single recessive gene) to the disease-resistant Brassica oleracea C41 was carried out. Wherein 80 parts of materials all show hairless, utilize the molecular marker developed in embodiment 1 to carry out genotype identification, the steps are as follows:
[0053] 1) Take 80 BC 1 f 1 The leaf DNA of the seedling stage of the backcross population is used as a template;
[0054] 2) PCR amplification was carried out using the following primers:
[0055] The forward primer sequence is 5'-AAACCTTCTCAACAAAGCCCA-3', the reverse primer sequence is 5'-CCACTTTACGCTACACAC...
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