柱花草单粒种子DNA提取和RAPD扩增  

DNA Extraction from Single Seed and RAPD Procedure for Stylosanthes sp.

作　　者：傅小霞[1] 漆智平[1] 何华玄[1] 

机构地区：[1]中国热带农业科学院热带作物品种资源研究所,海南儋州571737

出　　处：《热带农业科学》2004年第3期28-30,共3页Chinese Journal of Tropical Agriculture

摘　　要：以7个柱花草品种的单粒种子为材料,采用改良的CTAB法提取DNA,进行RAPD扩增。结果表明,采用单粒种子提取的DNA完整性好。采用模板3μL,随机引物2μL(10μmol/L),dNTPs2.5μL(2mmol/L),Taq酶0.2μL(5U/μL),MgCl2溶液2μL(25mmol/L),10×Buffer2.5μL(500mmol/LKCl,100mmol/LTris-Cl,pH9.0,25℃,10mL/LTritonX-100),无菌水12.8μL,总体积为25μL的反应体系,可扩增出谱带清晰的RAPD产物。扩增程序为:94℃预变性4min;94℃变性30s、37℃复性30s、72℃延伸90s,共40个循环;72℃延伸6min。DNA extraction and optimum conditions of RAPD procedure were studied u sing single dry seed from seven Stylosanthes cultivars individual as experimenta l material. The results showed that the effective amplification products could b e obtained with DNA templates extracted from dry seed by modified CTAB. The opti mum reaction system is: 3 ìL template, 2 ìL (10 ìmol/L) random primers, 2 ìL (25 mmol/L) Mg2+, 2.5 ìL (2 mmol/L) dNTPs, 1 U Taq polymerase, 10譈uffer 2.5 ìL, dd H2O 12.8 ìL, totaling 25 ìL. Forty PCR cycles were performed, with 30 s of denaturation at 94癈, 30 s of annealing at 37癈 and 90 s of polymerizatio n at 72癈.

关 键 词：柱花草 单粒种子 DNA提取 RAPD扩增 牧草 

分 类 号：S541[农业科学—作物学]