落羽杉属树木基因组总DNA的提取及SRAP反应体系的优化  

Extraction of Genomic DNA and Optimization of SRAP Reaction System in Taxodium Plants

作　　者：於朝广[1] 殷云龙[1] 

机构地区：[1]江苏省中国科学院植物研究所(南京中山植物园),南京210014

出　　处：《基因组学与应用生物学》2009年第1期109-114,共6页Genomics and Applied Biology

基　　金：江苏省高技术(农业)研究项目(BG2007318);国家林业局植物新品种保护办公室植物新品种测试指南及已知品种数据库项目(2007010);江苏省林业三项工程项目(lysx200702)资助

摘　　要：本文针对落羽杉属植物组织中多糖、多酚等次生物质含量高的特点,对其基因组DNA提取方法进行研究,比较了SDS法、CTAB法提取落羽杉属植物基因组DNA的效果,结果表明:CTAB法提取效果较佳。在此基础上,利用正交设计法,对SRAP反应体系中的各个主要影响因子Mg2+、dNTP、引物、Taq DNA聚合酶进行了优化筛选,确立了适合落羽杉属植物SRAP-PCR反应的最佳体系,即10μL体系中含有1μL10×PCR buffer,Mg2+2.0 mmol/L,dNTP 100μmol/L,引物0.3μmol/L,Taq DNA聚合酶0.5U和50ng模板DNA。利用该优化体系,通过48对SRAP引物组合对2个落羽杉属植物(落羽杉和墨杉)及4个杂交后代进行SRAP扩增,结果发现,SRAP引物及优化后的反应体系能够有效地用于落羽杉属植物种质资源鉴定及遗传多样性分析等研究。Study on the extraction of high quality DNA from Taxodium was carried out in allusion to high levels of polysaccharides and poly phenols combined with nucleic acid. We compared the methods of SDS and CTAB extracting DNA from Taxodium, the results showed that CTAB extraction method is more effective. Then the con- centration of Mg^2＋, dNTP, primer and Taq DNA polymerase in the SRAP-PCR system was optimized for genomic DNA in Taxodium, the optimal concentration was 10×PCR buffer, Mg^2＋ 2.0 mmol/L, dNTP 100 μmol/L, Primer 0.3 μmol/L, Taq DNA polymerase 0.5 U, template DNA 50 ng with a total volume of 10μL reaction solution. The molecular identification of two Taxodium plants and 4 hybrids was conducted by 48 SRAP primers with the optimal SRAP marker system. It showed that the SRAP primers and its optimized system can be applied in the germplasm identification and genetic diversity analysis of Taxodium plants effectively.

关 键 词：落羽杉属 DNA提取 SRAP—PCR 体系优化 

分 类 号：Q943[生物学—植物学]