Abstract: Miscanthus andresson (Poaceae) has recently become a research focus because it has been defined as a second-generation non-food energy crop. In this study, genomic DNAs of Miscanthus species from China including Miscanthus sinensis, M. floridulus, M. sacchariflorus, M. lutarioriparius, M. nepalensis, M. nudipes, and M. paniculatus were chosen as specimen samples for subsequent assay. The concentration of Mg²⁺, dNTP, primer, DNA template and the number of amplification cycles suitable for ISSR-PCR amplification reaction of Miscanthus species were optimized using the multifactor and multilevel experiments. Finally, an optimal ISSR-PCR amplification reaction system for the Miscanthus of China was established successfully. The 20 μL volumes of reaction system contained 1×PCR buffer, Mg²⁺ 2.5 mmol·L^-1, 0.25 mmol·L^-1 of each dNTP, primer 0.5 μmol·L^-1, 2 μL DNA, and 1 unit of plus Taq DNA polymerase, which undoubtedly supplied the technical reference for further related genetic analysis of Miscanthus in China.

Key words: Miscanthus, ISSR, PCR reaction system, Optimization

摘要： 芒属植物(Miscanthus)被认为适合作为新一代能源作物开发利用而成为当前国内外研究热点之一。为给后续相关研究奠定基础,本研究从中国芒属植物的芒(Miscanthus sinensis)、五节芒(M. floridulus)、荻(M. sacchariflorus)、南荻(M. lutarioriparius)、尼泊尔芒(M. nepalensis)、双药芒(M. nudipes)和红山茅(M. paniculatus)等类群中挑取部分材料,以其基因组DNA为模板,采用同一试验考察多个因素及水平的筛选方式对ISSR-PCR扩增体系中的Mg²⁺、dNTP、引物、模板的浓度以及循环数进行优化,建立适用于中国芒属植物的最佳ISSR-PCR反应体系。该体系为20 μL,含Mg²⁺2.5 mmol·L^-1,dNTP 0.25 mmol·L^-1,引物0.5 μ mol·L^-1,DNA 2 μL及1 U Taq Plus DNA聚合酶和1×PCR buffer。结果将为进一步开展中国芒属植物的相关遗传分析研究提供技术参考。

关键词: 芒属, ISSR, PCR反应体系, 优化