油楠基因组DNA提取·ISSR反应体系优化及引物筛选  

DNA Extraction,Optimization of ISSR-PCR System and Primers Screening of Sindora glabra

作　　者：吴忠锋 杨锦昌[2] 尹光天[2] 李荣生[2] 邹文涛[2] 袁洁[2] 

机构地区：[1]广州市林业和园林科学研究院,广东广州510405 [2]中国林业科学研究院热带林业研究所,广东广州510520

出　　处：《安徽农业科学》2015年第26期13-16,35,共5页Journal of Anhui Agricultural Sciences

基　　金：热带林业研究所基本科研业务费专项(RITFYWZX201206);林业行业专项(201304604);广州市林业和园林局科研项目

摘　　要：[目的]对油楠基因组DNA提取、ISSR反应体系优化及引物筛选进行探讨。[方法]运用改良CTAB法对油楠树皮进行基因组DNA提取,利用单因素试验,对油楠ISSR-PCR反应各因素水平进行优化。[结果]运用改良CTAB法对油楠树皮进行基因组DNA提取效果最佳;最优体系:20μl反应体系中,模板用量20 ng,引物浓度0.6μmol/L,d NTPs浓度0.2 mmol/L,Mg2+浓度2.0 mmol/L,Taq酶用量1.5 U,10倍反应缓冲液2μl,dd H2O补至20μl;以该体系为基础进行引物筛选,在100条ISSR随机引物中筛选出13条多态性较高、扩增条带清晰、重复性好的引物。[结论]该研究丰富了油楠遗传学基本资料,为油楠遗传多样性研究奠定基础。[Objective] DNA extraction,optimization of ISSR-PCR system and primers screening of Sindora glabra were investigated.[Method]Using the modified CTAB method for genonmic DNA extraction of Sindora glabra leather,using the method of single factor experiment,ISSR-PCR reaction factors level was optimized. [Result]The results showed that： the extraction effect for genomic DNA of Sindora glabra bark was the best. The optimal reaction system in 20 μl reaction system was 20 ng DNA,0. 6 μmol / L ISSR primer,0. 2 mmol / L d NTPs,2. 0 mmol / L Mgk D2 ＋,1. 5 U Taq DNA polymerase and 2 μl tenfold reaction buffer solution with dd H2 O added to 20 μl of total solution; According to this reaction system,13 primers in 100 primers were chosen for high polymorphism,their charity and repetition.[Conclusion]The study enriched genetics basic information of Sindora glabra,laid the foundation for the research of genetic diversity.

关 键 词：油楠 DNA提取 ISSR-PCR 优化 

分 类 号：S166[农业科学—农业气象学]