欢迎访问《植物研究》杂志官方网站,今天是 分享到:

植物研究 ›› 2011, Vol. 31 ›› Issue (1): 105-108.doi: 10.7525/j.issn.1673-5102.2011.01.019

• 论文 • 上一篇    下一篇

正交设计优化北重楼ISSR-PCR体系

张龙进1;白成科1,2*   

  1. 1.陕西师范大学生命科学学院,西安 710062;2.西北濒危药材资源开发国家工程实验室,西安 710062
  • 收稿日期:1900-01-01 修回日期:1900-01-01 出版日期:2011-01-20 发布日期:2011-01-20
  • 通讯作者: 白成科
  • 基金资助:
     

Optimization of ISSR-PCR System for Paris vertcillata

ZHANG Long-Jin;BAI Cheng-Ke;*   

  1. 1.College of Life Sciences,Shaanxi Normal University,Xi’an 710062;2.National Engineering laboratory for Resource Development of Endangered Crude Drugs in Northwest of China,Xi’an 710062
  • Received:1900-01-01 Revised:1900-01-01 Online:2011-01-20 Published:2011-01-20
  • Contact: BAI Cheng-Ke
  • Supported by:
     

摘要: 利用正交试验设计L9(34)的方法,最终建立了北重楼ISSR-PCR的优化反应体系,即20 μL反应体系中包含1×buffer,dNTP 175 μmol·L-1,Primer 0.8 μmol·L-1,Mg2+ 1.4 mmol·L-1Taq酶2.5 U及模板DNA 80 ng。适宜的扩增程序为95℃预变性5 min;94℃变性40 sec,55℃退火45 sec,72℃延伸90 sec,40个循环;72℃延伸7 min,4℃保存。该优化体系的建立,将为北重楼及其近缘种的系统学和种质资源鉴定及遗传多样性的研究奠定基础。

关键词: 北重楼, ISSR-PCR, 分子标记, 正交优化

Abstract: The optimal ISSR-PCR system for Paris vertcillata was established with orthogonal design.A total of 20 μL ISSR-PCR system contains 1×buffer, dNTP 175 μmol·L-1,Primer 0.6 μmol·L-1,Mg2+ 1.4 mmol·L-1Taq DNA polymerase 2.5 U and 80 ng template DNA.The suitable PCR procedure is one cycle denaturing at 95℃ for 5 min;40 cycles each involved denaturing at 94℃for 40 s,annealing at 55℃ for 45 s,extending at 72℃ for 90 s,one cycle extending at 72℃ for 7 min,and then remaining at 4℃.This optimized system would play an important role in further research of systematology on P.vertcillata and its related species, as well as identification on germplasm, and genetic diversity by ISSR molecular marker.

Key words: Paris vertcillata, ISSR-PCR, molecular markers, orthogonal optimization

中图分类号: