利用 L16(4)5正交实验设计,对新疆狗牙根 EST-SSR反应体系进行优化;并将该体系初步应用于14份新疆典型区域的野生狗牙根材料,通过聚类分析研究供试材料的亲缘关系.结果表明,新疆狗牙根 EST-SSR反应最优体系为:20μL反应体系中含模板DNA 100 ng,引物0.5μmol/L,2μL 10×Buffer,MgCl21.5 mmol/L,dNTPs 0.2 mmol/L,Taq DNA聚合酶0.5 U.运用该体系对14份新疆狗牙根材料进行PCR扩增,条带较清晰,可用于新疆狗牙根 EST-SSR标记的进一步研究.对14份狗牙根的多样性及其亲缘关系分析结果表明,狗牙根 Nei’s 基因多样性指数(H)变异范围为0.2077~0.3153,平均为0.2550;Shannon信息指数(I)在0.2540~0.4394.
The best optimized system from Xinjiang Cynodon dactylon L.were as follows:There were template DNA 100 ng,0.5 μmol/L of each primer,2 μL of 10 × Buffer,MgCl2 1.5 mmol/L,dNTPs 0.2 mmol/L,Taq DNA polymerase 0.5 U in 20μL reaction system.The 14 portions of Xinjiang Cynodon dactylon L.were amplified by use of the system,the bands were clearer,which can be used in studying Xin-jiang Cynodondactylon L.EST-SSR marker.The diversity and phylogenetic relationships of 14 portions of Cynodon dactylon L.were analyzed.The results showed that the variance range of diversity indexes (H)of Cynodon dactylon L.Nei’s were as follows:0.2077-0.3153,the average was 0.2550,the information index (I)was 0.2540 to 0.4394.