十里香茶基因组DNA高效提取与RAPD方法建立

本文以嫩芽和鲜叶为材料,建立了十里香茶高效、微量基因组DNA提取方法,提取的DNAOD260/280在1.7～2.0之间,DNA产率在81.8～483.5 ng/mg之间,能够满足RPAD的需要。同时建立了十里香茶RAPD分子标记方法,25μL PCR反应体系中包含:50 ng DNA模板,1.2μL引物,2.5μL 10×PCR Buffer,2.5μL 25 mMMgCl2,2μL 10 mM dNTP,14.8μL ddH2 O1,U Taq聚合酶。PCR扩增程序为94℃预变性3 min,94℃变性1 min,36℃退火1 min 20 s,72℃延伸2 min4,0个循环后72℃延伸10 min。15个RAPD随机引物以十里香1号DNA为模板,分别扩增出1～7条带。

Genomic DNA extraction and establishment of RAPD reaction conditions for Camellia sinensis var.Shilixiang

Shilixiang tea is an ancient and famous tea plant(Camellia sinensis) variety in Kunming City of Yunnan Provience in China,which has been cultivated since Tang Dynasty.In this paper,the authors established an efficient and mini-preparation method for extracting genomic DNA from shoots and fresh leaves of Shilixiang tea.The OD260/280of the extracted DNA were 1.7~2.0 with 81.8~483.5 μg/mg of yields.The established RAPD reaction condition was in 25μL PCR reaction system included:50 ng genomic DNA,1.2 μL primer,2.5 μL 10×PCR Buffer,2.5 μL 25 mM MgCl2,2 μL 10 mM dNTP,14.8 μL ddH2O and 1 U Taq polymerase.The amplification procedure condition were predenature at 94℃ for 1 min,annealing at 36℃ for 1 min and 20 s,extension at 72℃ for 2 min,40 for cyclings,last extension 72℃ for 10 min.1~7 bands were amplified by 15 primers.