The dna molecular marker for identifying the quality of rice fatty acid and its application
A technology of DNA molecules and fatty acids, which is applied in the field of DNA molecular markers to identify the quality of rice fatty acids, can solve the problems of difficulty, difficulty in operation, and high cost, and achieve low cost, simple operation methods, and accurate and reliable detection results
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Embodiment 1
[0027] Example 1 Development of rice linolenic acid quality gene markers
[0028] 1. According to the literature
[0029] Kodama H, Akagi H, Kusumi K, Fujimura T, Iba K. (1997). Structure, chromosomal location and expression of a rice gene encoding the microsome ω-3fatty acid desaturase. Plant Molecular Biology, 33: 493-502 report, Rice The protein encoded by the OsFAD3 gene (the gene number corresponding to MSU is Loc_Os11g01340) has the activity of fatty acid dehydrogenase, which can catalyze the conversion of linoleic acid into linolenic acid. In the website ricevarmap (http: / / ricevarmap.ncpgr.cn) and RIGW (http: / / rice.hzau.edu.cn / cgi-bin / gb2 / gbrowse_syn) the gene sequence of the resequenced germplasm OsFAD3 was compared Yes, an 8bp (AGGAAGGA) insertion-deletion variation was found in the 5'UTR region of the OsFAD3 gene.
[0030] 2. Download the OsFAD3 gene sequence from the MSU website (http: / / rice.plantbiology.msu.edu / ), intercept the upstream and downstream 400bp seque...
Embodiment 2
[0031] Example 2 Detection method of gene marker OSFA3 and its polymorphism in rice parents
[0032] 1. Extract the DNA of rice samples to be tested, including rice conventional rice varieties, common wild rice and African cultivated rice
[0033] 2. Using the designed gene marker OSFA3, carry out PCR amplification on the sample DNA, and detect the polymorphism of the gene marker OSFA3.
[0034]PCR reaction system: total volume 20ul including: 100ng DNA template, 2ul 10X PCR buffer, 2ul 2mMdNTP mixture, 0.3ul each of 10uM primers OSFA3F and OSFA3R, 0.1ul rTaq DNA polymerase. Perform PCR reactions on a PCR thermocycler. The PCR amplification program was set as follows: 94°C pre-denaturation for 4 minutes, followed by 94°C denaturation for 30 seconds, 55°C annealing for 30 seconds, 72°C extension for 30 seconds, 32 cycles, and finally 72°C extension for 7 minutes to end the reaction.
[0035] The PCR product was electrophoresed in 4% polyacrylamide gel for 40 minutes (electrop...
Embodiment 3
[0037] Example 3 Application of Gene Marker OSFA3 in Selection of Linolenic Acid Quality in Breeding Populations
[0038] In order to evaluate the effect of molecular marker-assisted selection of rice linolenic acid quality using marker OSFA3, a breeding segregation population constructed by crossing japonica rice Nipponbare (NIP) with indica rice Zhenshan 97 (ZS97) was analyzed for marker OSFA3, and rice linoleic acid / Genotypes with higher linolenic acid content. Said application includes the following steps:
[0039] 1. Extract the DNA of 107 individual plants.
[0040] 2. Use the marker OSFA3 to detect the genotype of the OsFAD3 gene. If it is consistent with the parental ZS97 genotype, it will be recorded as 11 (the size of the amplified product is 182bp), and if it is consistent with the NIP genotype, it will be recorded as 22 (the size of the amplified product is 190bp). The NIP genotype was recorded as heterozygous 12. The genotypes of marker OSFA3 in 107 individual...
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