珊瑚菜叶绿体基因TrnV-M序列的PCR扩增与分析  

PCR Amplication and Analysis of Glehnia littoralis Using TrnV-M Gene Fragments

作　　者：慕春歌[1] 

机构地区：[1]鲁东大学资产处,山东烟台264025

出　　处：《安徽农业科学》2016年第33期135-136,154,共3页Journal of Anhui Agricultural Sciences

基　　金：山东省自然科学基金面上项目(ZR2010CM056)

摘　　要：[目的]对珊瑚菜TrnV-M基因片段PCR扩增条件进行优化和测序分析,为濒危物种珊瑚菜的种质资源保护和遗传多样性研究提供理论依据。[方法]探索并优化珊瑚菜叶绿体基因片段TrnV-M的PCR扩增条件,并对扩增产物进行测序分析。[结果]珊瑚菜TrnV-M基因片段的PCR最佳反应体系:d NTPs 0.50μL,Buffer 2.50μL,Trn M 1.25μL,TrnV 1.25μL,DNA 1.00μL,r Taq E 0.15μL,dd H2O 18.35μL,总体积25.00μL。最佳扩增程序:94℃预变性3.0 min;94℃变性50 s,58℃退火45 s,72℃延伸1 min 25 s,38个循环;72℃延伸8 min。[结论]在最优体系下获得了清晰、稳定的目的条带,校正后条带长度约为757 bp,通过Gen Bank的BLAST比对,确定为TrnV-M基因片段。[Objective] The aim was study PCR amplication and analysis of Glehnia littoralis using TrnV-M gene fragments, to provide a tech-nical basis for germplasm resources protection and genetic diversity study of Glehnia littoralis.[Methods] The PCR amplification conditions for TrnV-M gene fragments of Glehnia littoralis were explored and optimized, and the PCR products were sequenced.[Result] The optimized PCR system was acquired as follows： dNTPs 0.50 μL, Buffer 2.5 μL, TrnM 1.25 μL, TrnV 1.25 μL, DNA 1.00 μL, rTaqE 0.15 μL, ddH2O 18.35 μL,the total volume was 25 μL.The optimized thermal cycling profile was as follows：initial template denaturation at 94 ℃ for 3 min, 94 ℃ for 50 s, 58 ℃ for 45 s, and 72 ℃ for 1 min 25 s, followed by 38 cycles, with a final extension of 72 ℃ for 8 min.[ Conclusion] The total alignment of TrnV-M sequences was 757 bp in length after sorting by hand.

关 键 词：珊瑚菜 TrnV—M PCR扩增 最佳反应体系 

分 类 号：S188.1[农业科学—农业基础科学] Q789[生物学—分子生物学]