Application of molecular marker MtD4 in identifying carrot petal male sterility
A technique for male sterility and carrots, applied in the field of breeding, can solve the problems of verification, inability to use multiple genotypes of petal flower type sterile lines and maintainer lines, complex structure, etc., and achieve the effect of accelerating the pace of breeding.
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Embodiment 1
[0067] Example 1, Development of mitochondrial haplotype-specific (Mitotype-specific sequence, MSS) molecular markers and polymorphism detection
[0068] 1. Development of MSS molecular markers
[0069] 1. Extract the mitochondrial DNA of carrot 50119 and carrot OK respectively, and then perform mitochondrial genome sequencing.
[0070] 2. After completing step 1, compare and analyze the mitochondrial genome sequencing results of carrot 50119 and carrot O K, and obtain 3 MSS sequences (named MSS sequence 2-MSS sequence 4 in turn). The nucleotide sequence of MSS sequence 2 is shown in sequence 1 in the sequence listing. The nucleotide sequence of MSS sequence 3 is shown as sequence 2 in the sequence listing. The nucleotide sequence of MSS sequence 4 is shown as sequence 3 in the sequence listing.
[0071] 3. Based on the two variant forms of MSS sequence 2 in the carrot mitochondrial genome, the molecular marker MtD2 was developed.
[0072] The nucleotide sequence of the am...
Embodiment 2
[0113] Embodiment 2, the application of MSS molecular marker
[0114] The PCR reaction system in this embodiment is all 10 μ L, consists of 1 μ L genomic DNA (concentration is 50-100 ng / μ L) of young carrot leaves to be tested, 1 μ L 10 × PCR reaction buffer (containing Mg 2 +), 1 μL dNTPs (2.5 mM concentration), 0.2 μL upstream primer (10 μM concentration), 0.2 μL downstream primer (10 μM concentration), 0.2 μL Taq DNA polymerase (2.5 U / μL concentration), and 6.4 μL ddHO composition.
[0115] 10× PCR reaction buffer (containing Mg 2 +) is a product of Tiangen Biochemical Technology (Beijing) Co., Ltd.
[0116] The PCR reaction procedures in this example are: pre-denaturation at 94°C for 5 min; denaturation at 94°C for 30 s, annealing at 55°C for 30 s, extension at 72°C for 30 s, and 35 cycles; extension at 72°C for 5 min.
[0117] 1. Application 1
[0118] The carrots to be tested were 16 carrot materials imported and preserved from the Vegetable Research Center of the Un...
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