摘要
以桑科榕属植物为研究对象,利用改进的CTAB法提取基因组DNA,并通过单因素多水平梯度试验,筛选了模板、Mg2+、Taq酶、dNTPs和随机引物的浓度及用量,建立了榕属植物RAPD技术分析体系。反应体系总体积为25μL,各组分浓度为:10×Buffer 2．5μL,Mg2+ 2．5 mmol/L,Taq DNA polymerase 0．06 U/μL,dNTPs 0．2 mmol/L,Primer 0．2 μmol/L．Template DNA 1．2 ng/μL。PCR循环程序为:94 C预变性4 min;94 C循环变性 1 min,36 C退火1 min,72 C延伸2 min,40个循环;最后72 C延伸8 min。
An improved DNA extraction protocol with CTAB was developed for Ficus species. The genomic DNA was employed to establish optimization of RAPD. The RAPD system for Ficus L. （Moraceae） was optimized including program,template ,primer,dNTPs ,Taq polymerase and Mg^2＋. Through ladder experiments,the reliable RAPD analysis system is established. The total reaction volume is 25 μL and reaction mixture consist of 2.5 μL 10×Buffe,1.2 ng/μL Template DNA,0. 2 mmol/L dNTPs,2.5 mmol/L Mg^2＋ ,0. 2 μmol/L random Primer and 0.06 U/μL Hotstar Taq DNA polymerase. RAPD program is 4 min at 94 C for predenaturation,then 40 cycles of 1 min at 94 C for denaturation,of 1 min at 36 C for annealing,of 2 min at 72 C for extension,finally extension at 72 C for 8 min.
出处
《西北农林科技大学学报（自然科学版）》
CSCD
北大核心
2006年第5期147-152,共6页
Journal of Northwest A&F University(Natural Science Edition)
