枣树基因组DNA提取及其RAPD体系优化

Genomic DNA Extraction and RAPD System Optimization of Chinese Jujube

以不同生长期的枣和酸枣为研究对象,采用CTAB法提取枣基因组DNA;通过单因素5水平试验,筛选模板、Mg2+、Taq酶、dNTPs和随机引物的浓度及其用量,建立了枣和酸枣RAPD技术最优体系,即25μl反应体系中各组分含量为10×Taq Buffer+KCl 2.5μl;Taq DNA聚合酶0.04 U/μl;MgCl22.0 mmol/L;模板DNA2 ng/μl;引物0.3μmol/L;dNTP为0.2 mmol/L。适宜的扩增程序为:94℃预变性5 min;94℃循环变性50 s,36℃退火50 s,72℃延伸1.5 min,38个循环;72℃延伸10 min。

Genomic DNA was isolated from different growth stages of Ziziphusjujuba Mill. and Ziziphus acidojujuba C. Y. using CTAB method. Content and dosage of template, primer, dNTPs, Mg^2＋ and Taq DNA polymerase were selected by single factor five level tests. The optimum RAPD systems for Ziziphusjujuba Mill, and Ziziphus acidojujuba C. Y. were established, The total reaction volume was 25 μl containing 2.5 μl 10×Taq Buffer with KCl, 0.04 U/μl Taq DNA polymerase, 2.0 mmol/L MgCl2, 2 ng/μl template DNA, 0.3 μmol/L primer and 0.2 mmol/L dNTPs, The appropriate PCR procedure was 94 ℃ 5 min,1 cycle; 94℃ 50 s, 36 ℃50 s, 72 ℃1.5 min,38 cycles; 72℃ 10 min.