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Bulletin of Botanical Research ›› 2010, Vol. 30 ›› Issue (5): 576-581.doi: 10.7525/j.issn.1673-5102.2010.05.010

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Establishment and Optimization of ISSR-PCR System for Hybrid Rapeseed

PENG Bing-Yu;HU Yan-Ping;GONG Ai-Qi;WANG Huan-Qiang;WANG Li;BAO Tian-Zhong;LI Rui-Xin;LI Yi*   

  1. 1.Qinghai Seed Management Station,Xining 810000;2.Key Laboratory of Adaptation and Evolution of Plateau Biota,Northwest Institute of Plateau Biology,Chinese Academy of Sciences,Xining 810001;3.Department of Food Science and Engineering,East China University of Science and Technology,Shanghai 201424
  • Received:1900-01-01 Revised:1900-01-01 Online:2010-09-20 Published:2010-09-20
  • Contact: LI Yi
  • Supported by:

Abstract: An orthogonal design was used to optimize the ISSR-PCR system for hybrid rapeseed at three levels of four factors (Mg2+, dNTP, primer and Taq DNA polymerase). Then, based on the optimal ISSRPCR amplification system, the concentration of DNA template and annealing temperature were selected. As a result, the optimal PCR (20 μL) mixture contained 1.50 mmol·L-1 Mg2+, 0.125 mmol·L-1 dNTP, 2.00 μmol·L-1 primer, 0.50 U Taq DNA polymerase, 2.5 μL 10×buffer and 40ng template DNA. The suitable annealing temperature of primer UBC891 was 54.2℃. With the optimal system of ISSR-PCR reaction, clear and steady bands were obtained in different individuals of Qingza No.3 and its male parent. The establishment of ISSR-PCR system could favor the studies on the purity and genuineness of hybrid rapeseed varieties by using molecular marker techniques.

Key words: Hybrid Rapeseed, ISSR-PCR, reaction conditions, orthogonal design

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