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：dNTPs 0．50μL，Buffer 2．50μL，TrnM 1．25μL，TrnV 1．25μL，DNA 1．00μL，rTaqE 0．15μL， ddH2 O 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 GenBank 的 BLAST 比对，确定为TrnV－M基因片段。
Abstract
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, ddH2 O 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.
基金项目
山东省自然科学基金面上项目(ZR2010 CM056)
国家科技期刊平台
NSTL
维普
万方数据