鹰嘴豆SSR-PCR反应体系优化  

Optimization of SSR-PCR Reaction System of Chickpea

作　　者：李辉玲[1] 李利民[2] 张金波[2] 

机构地区：[1]石河子大学生命科学学院,新疆石河子832000 [2]新疆农业科学院农作物品种资源研究所,新疆乌鲁木齐830091

出　　处：《贵州农业科学》2014年第3期10-13,共4页Guizhou Agricultural Sciences

基　　金：新疆维吾尔自治区自然科学基金"鹰嘴豆种质资源遗传多样性研究"(2011211A089)

摘　　要：为了进行鹰嘴豆(Cicer arietinum L.)遗传多样性分析,以鹰嘴豆叶片基因组DNA为模版,通过Mg2+、DNA聚合酶、模版DNA、dNTPs以及引物的用量,设计5因素4水平的正交试验对SSR-PCR反应体系进行优化,并利用梯度PCR对退火温度进行选择。结果表明:优化的SSR-PCR扩增10μL体系为1.5μL 1×PCR Buffer、1.5mmol/L Mg2+、1.5U TaqDNA聚合酶、0.75μmol/L SSR引物、15ng模版DNA、250μmol/L dNTPs。扩增程序:95℃预变性5min;94℃变性30s,52.3℃退火30s,72℃延伸90s,共35个循环;最后72℃延伸5min。所有SSR引物在优化的反应体系中基本能有效扩增,得到特异性。The SSR-PCR reaction system of chickpea was optimized by studying the optimum concentration of Mg2＋ ,TaqDNA polymerase, template DNA, dNTPs and prime to analyze genetic diversity of chickpea,and the annealing temperature was selected by using gradient PCR.The results showed that the optimized SSR-PCR system for 10μL amplification included 1＆#215;PCR Buffer,1.5 mmol/L Mg2＋ ,1.5 U TaqDNA polymerase,0.75 μmol/L SSR primer,15 ng template DNA and 250 μmol/L dNTPs,and the amplification procedure included 35 circulations of pre-denature at 95℃ for 5min,at denature at 94℃ for 30s,annealing at 52.3℃ for30s and extension at 72℃ for 90s and final extension at 72℃ for 5min.All SSR primers all can be effectively amplified under the optimized system basically.

关 键 词：鹰嘴豆 PCR 扩增 扩增产物 

分 类 号：S643.5[农业科学—蔬菜学]