Optimization for ISSR-PCR System of Polygonum Capitatum Buch-Ham. ex D. Don by Orthogonal Design

ZHOU Tao1; WU Yu1; JING Yan-lei1; JIANG Wei-ke1; CHEN Mei-lan2

doi:10.13422/j.cnki.syfjx.2010.06.004

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Optimization for ISSR-PCR System of Polygonum Capitatum Buch-Ham. ex D. Don by Orthogonal Design

更新时间：2024-09-23

- Optimization for ISSR-PCR System of Polygonum Capitatum Buch-Ham. ex D. Don by Orthogonal Design
- Chinese Journal of Experimental Traditional Medical Formulae Vol. 16, Issue 6, Pages: 50-53(2010)
- 作者机构：
  
  1. 贵阳中医学院
  2. 中国中医科学院中药研究所
- 作者简介：
- 基金信息：
- DOI：10.13422/j.cnki.syfjx.2010.06.004
  CLC： S567.239
- Published：2010
- 稿件说明：
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[1]周涛,吴钰,金艳蕾,江维克,陈美兰.头花蓼重复片段多态性分析-多聚酶链式反应体系建立与正交优化[J].中国实验方剂学杂志,2010,16(06):50-53.

ZHOU Tao1, WU Yu1, JING Yan-lei1, et al. Optimization for ISSR-PCR System of Polygonum Capitatum Buch-Ham. ex D. Don by Orthogonal Design[J]. Chinese journal of experimental traditional medical formulae, 2010, 16(6): 50-53.
[1]周涛,吴钰,金艳蕾,江维克,陈美兰.头花蓼重复片段多态性分析-多聚酶链式反应体系建立与正交优化[J].中国实验方剂学杂志,2010,16(06):50-53. DOI： 10.13422/j.cnki.syfjx.2010.06.004.

ZHOU Tao1, WU Yu1, JING Yan-lei1, et al. Optimization for ISSR-PCR System of Polygonum Capitatum Buch-Ham. ex D. Don by Orthogonal Design[J]. Chinese journal of experimental traditional medical formulae, 2010, 16(6): 50-53. DOI： 10.13422/j.cnki.syfjx.2010.06.004.

摘要

目的:建立头花蓼ISSR的反应体系。方法:通过正交试验

研究了Mg2+浓度、dNTP浓度、Taq DNA聚合酶浓度、引物浓度这4个因素在3个水平上对ISSR-PCR的影响。结果:确立了适合于头花蓼ISSR PCR的优化体系:25μL PCR反应体系中含有1×buffer缓冲液（10 mmol.L-1 KC1

8 mmol.L-1（NH4）2SO4

10 mmol.L-1 Tris.HC1

pH 8.0）

3.0 mmol.L-1 MgC12

d NTP 200μmol.L-1

2.0 U.25μL-1 Taq酶

0.25 mmol.L-1引物

40 ng.L-1模板DNA。利用温度梯度PCR

确定了最适宜的退火温度为48℃。结论:该优化体系的建立为进一步对头花蓼进行种质资源的遗传多样性分析奠定了基础。

Abstract

Objective:To establish the optimum ISSR-PCR system of Polygonum capitatum.Method:The Mg2 +

dNTPs

primers and Taq DNA polymerase

were investigated to optimize the ISSR-PCR reaction system of Polygonum capitatum by orthogonal tests.Result:The optimized ISSR-PCR system of P.capitatum included 1 × DNA polymerase buffer（10 mmol.L-1 KCl

8 mmol.L-1（NH4）2SO4

10 mmol.L-1 Tris.HC1

pH 8.0）

3.0 mmol.L-1 MgCl2 200 μmol.L-1 dNTPs

2.0 U.25 μL-1 Taq DNA polymerase

0.25 mmol.L-1 primer

40 ng.L-1 DNA template

in 25 μL PCR reaction systerm.By temperature gradient PCR

the optimum annealing temperature was determined as 48 ℃.Conclusion:The optimized system laid the groundwork for ISSR molecular analysis of P.capitatum Buch-Ham.ex D.Don.

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Related Institution

Dongguan Hospital of Traditional Chinese Medicine

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