Optimization of ISSR Reaction System for Pinus armandii
LI Cuixin
HE De
Abstract：To obtain amplification result with clear,stable and rich polymorphism fragment of Pinus ar-mandii,the affection ISSR-PCR by main reaction system elements,such as primer,Mg2+and annealing temperature,was presented.The reaction system and amplified procedure suitable for P.armandii were es-tablished,the main elements include,15 μL amplification reaction mixture containing 1.5 μL 10×reaction buffer,22.5 ng template DNA,2.2 mmol/L Mg2+,0.23 mmol/L dNTPs,0.93 μmol/L primers,0.027 U/μL TaqDNA polymerase.The reaction program was devised for 4 min of predenaturalization at 94℃,33 cycles of 30 s for denaturalization at 94℃,30 s of anneal at Tm-4.8℃,108 s of extension at 72℃,and 10 min of extension at 72℃ after the final cycle.The 12 best primers were selected for ISSR-PCR system of P.ar-mandii from 44 primers,whose results of ISSR amplification were clear and stable.The research on the op-timization of ISSR reaction system for P.armandii will lay a foundation for studying interspecific genetic variation and genetic map construction of P.armandii using ISSR molecular marker technique in the future.
Keywords：Pinus armandiiISSRoptimizationPCR
Publication Date：2018-04-02
Online Publishing Date：2025-08-15(First online date of this platform, not the publication date of the document)
Pages：5( 121-124,155 )