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Bulletin of Botanical Research ›› 2009, Vol. 29 ›› Issue (1): 80-85.doi: 10.7525/j.issn.1673-5102.2009.01.017

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Establishment and Optimization of RAPD-PCR Reaction System for Larix gmelnii(Rupr.) Rupr Using Orthogonal Design

LI Xue-Feng;ZHANG Han-Guo*;GUAN Chun-Yu;ZHANG Yao   

  1. Northeast Forestry University,Key Laboratory of Forest Tree Genetic Improvement and Biotechnology,Ministry of Education,Harbin 150040
  • Received:1900-01-01 Revised:1900-01-01 Online:2009-01-20 Published:2009-01-20
  • Contact: ZHANG Han-Guo
  • Supported by:

Abstract: The orthogonal design was used to optimize RAPD amplification system of Larix gmelnii(Rupr.) Rupr with five factors (Taq polymerase, Mg2+, dNTP, primer, DNA templet) at four levels, respectively. Through the deep analysis, a suitable RAPD-PCR reaction system was established, namely 20 μL reaction system containing 1.0 U Taq DNA polymerase, 2.5 mmol·L-1 Mg2+,0.25 mmol·L-1 dNTP, 0.5 μmol·L-1 primer, 1×PCR buffer, 90 ng DNA template. 20 primers with stable amplification and rich polymorphism for RAPD were screened. The optimal annealing temperature for RAPD-PCR reaction was proposed by gradient PCR. The result provided a standardizing RAPD-PCR program for the analysis of genetic diversity of L. gmelnii(Rupr.) Rupr.

Key words: Larix gmelnii(Rupr.)Rupr, RAPD-PCR, orthogonal design, reaction system

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