Method as well as molecular marker and primer for identifying rice male sterility type and three-line hybrid rice type
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A male sterile line, three-line hybrid rice technology, applied in biochemical equipment and methods, microbial determination/inspection, DNA/RNA fragments, etc., to achieve a wide range of applications and improve efficiency.
Inactive Publication Date: 2013-12-11
WUHAN UNIV
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Mitochondria, although thought to mutate at a slower rate than the nucleus, generate very rich genetic variation due to the susceptibility to recombination or intracellular genome-to-genome communication in mitochondrial intergenic regions
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Embodiment 1
[0028] Example 1 MSS molecular markers and primers
[0029] According to the differences in the mitochondrial genome sequences of different cytoplasmic male sterile rice, MSS molecular markers for identifying rice cytoplasmic male sterile types and three-line hybrid rice types were developed, respectively MSS01, MSS02, MSS03, MSS04, MSS05, MSS06, MSS07, MSS08 , MSS09, MSS10g, MSS11a and MSS12s, the sequences of which are shown in SEQ ID NO.1-12.
[0030] The primers and parameters designed according to the above MSS molecular markers are shown in Table 1.
[0033] Example 2 Detection of cytoplasmic karyotype of rice cytoplasmic male sterile lines
[0034] (1) DNA extraction
[0035]Select 10 different types of rice cytoplasmic male sterile lines: Y4A, Gang 46A (G46A), Ⅱ-32A, Xieqingzao A (XQZA), Maxie A (MXA), K17A, Chujing 23A (CJ23A), Baoyuan A (BYA), Zhenshan 97A (ZS97A) and Yuetai A (YTA), and maintainer lines Baoyuan B (BYB), Ⅱ-32B, Gang 46B (G46B), Y4B. When it grows to two leaves and one heart, take 0.5 grams of seedling leaves for each material and put them in a 2mL centrifuge tube, grind and break the cells with a high-speed oscillator, refer to Cold Spring Harbor Laboratory Press (Cold Spring Harbor Laboratory Press), Nina Irwin and Kaaren The book "Molecular Cloning" edited by A. Janssen extracted the total DNA of rice materials.
[0036] (2) Establishment of PCR amplification system
[0037] The standard PCR amplification reaction system is 25 μL, and the composition is as follows:
[0038] 50ng / μL total DNA 1μL, ...
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Abstract
The invention discloses a method as well as a molecular marker and a primer for identifying a rice male sterility type and a three-line hybrid rice type, belonging to the field of molecular marker detection. Sequences of the molecular marker and the primer for identifying the rice cytoplasmic male sterility type and the three-line hybrid rice type are shown as SEQ ID NO.1-36. The identification method disclosed by the invention comprises the steps of extracting the total DNA (Deoxyribose Nucleic Acid) of a rice material; by taking the extracted total DNA as a template, using the primer for PCR (Polymerase Chain Reaction) amplification; analyzing PCR amplification results, if amplification banding patterns of two materials are completely consistent, the two materials have the same cytoplasm karyotype, namely have the same female parent source, otherwise, the two materials have different cytoplasm karyotypes, namely have different female parent sources. The method as well as the molecular marker and the primer for identifying the rice male sterility type and the three-line hybrid rice type are easy and convenient to operate, stable and reliable, can be used for identifying cytoplasmic male sterile lines and three-line hybrid rice varieties, have the characteristics of accuracy, high efficiency and rapidness, and thus improve the identification efficiency of the sterile lines and new varieties.
Description
technical field [0001] The invention belongs to the technical field of molecular marker detection, and in particular relates to a method for identifying rice male sterility types and three-line hybrid rice types, molecular markers and primers. Background technique [0002] Rice mitochondria are inherited maternally, so the source of rice cytoplasm can be traced by mitochondrial markers. Mitochondria, although thought to mutate at a slower rate than the nucleus, generate very rich genetic variation due to the ease of recombination in the intergenic regions of mitochondria or the exchange of genomes in cells. This variation can serve as a specific molecular signature of the mitochondrial genome at the genetic level. The cytoplasmic karyotype can be marked by using mitochondrial molecular tags, so that different rice cytoplasmic male sterile lines or hybrid rice varieties can be effectively distinguished. Due to the uniqueness of molecular markers, specific genotypes can be t...
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