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以3份大白菜杂交种及其亲本为试验材料,利用改良CTAB法提取基因组DNA,采用正交设计对Mg2+、dNTPs、引物、模板DNA和Taq酶5因素4水平进行优化,结果表明,适用于大白菜纯度鉴定EST-SSR检测体系为:在10μL反应体系中包含3.0mmol·L-1MgCl2、400μmol·L-1dNTPs、0.20μmol·L-1SSR引物、4ng模板DNA和0.05UTaq酶。利用该优化的体系,对3份大白菜杂交种进行单株育苗试验,每份杂交种取单株140株进行纯度鉴定。结果显示,扩增带型清晰、稳定,纯度检测结果与田间试验结果基本吻合,可用于大白菜种子纯度分子标记的进一步研究。
Three Chinese cabbage hybrids and their parents were used as experimental materials, genomic DNA was extracted by modified CTAB method, and the levels of Mg2 +, dNTPs, primers, template DNA and Taq enzyme 5-factor 4 were optimized by orthogonal design. The purity of the Chinese cabbage EST-SSR detection system: 3.0mmol·L-1MgCl2, 400μmol·L-1dNTPs, 0.20μmol·L-1SSR primer, 4ng template DNA and 0.05UTaq enzyme in 10μL reaction system. Using the optimized system, three Chinese cabbage hybrids were tested for their single seedling growth, and 140 hybrid plants were taken for purity identification. The results showed that the amplified bands were clear and stable, and the results of the purity test were in good agreement with those of the field test, which could be used to further study the molecular marker of seed purity in Chinese cabbage.