山茶属植物体内酚类、多糖等次生代谢物质含量较高,获得高纯度的总DNA比较困难.为筛选出适宜山茶属植物基因组DNA的较优提取方法,并研究不同材料处理方法的差异,采用四因素三水平的正交设计,对山茶属植物基因组DNA的提取方法CTAB法进行优化.最后确定较优提取方法为:老叶用CTAB-free去除多糖并防止酚氧化,用2%CTAB、2%PVP、24:1抽提2次;嫩叶用2×CTAB去除多糖并防止酚氧化,用1%CTAB、2%PVP、24:1抽提1次,并用ISSR-PCR检测.结果显示采用优化方案提取的DNA质量更好,能较好地溶于TE,可用于ISSR-PCR扩增,且扩增出清晰的条带.%It is difficult to get high-quality DNA from Camellia,which is rich in phenolic compound poly-saccharides and other secondary metabolites.In order to select a best optimum method for extracting the genom-ic DNA from different leaves of Camellia,an orthogonal design by four factors at three levels was carried out to optimize the CTAB method for the genomic DNA extraction. Finally,the optimal extraction method was deter-mined that polysaccharide was removed and oxidation of phenol avoided,and CTAB-free,2% CTAB,2% PVP of 24:1 were used to extract two times for old leaves;polysaccharide was removed and oxidation of phenol avoi-ded,and 2×CTAB,1%CTAB,2%PVP of 24:1 were used to extract one time for tender leaves,and it was veri-fied by ISSR-PCR.The results showed that high quality genomic DNA of Camellia L. was obtained,and it was well dissolved in TE.It could be used in amplification of ISSR-PCR,in which clear bands could be detected.
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