摘要
以岷江百合为材料,对影响ISSR-PCR反应的主要参数进行优化,建立了适用于岷江百合的ISSR-PCR反应体系。20μL反应体系中,内含1×PCRbuffer、1.25mmol/LMg2+、0.4mmol/LdNTPs、0.6μmol/L引物、0.5UTaqDNA聚合酶、20ng模板DNA。扩增程序为:94℃预变性5min;94℃变性40s,52.8℃退火45s,72℃延伸1min30s,共45个循环;最后72℃延伸8min。该优化系统的建立为今后利用ISSR标记技术开展岷江百合遗传多样性研究奠定了基础。
The major factors in the reaction system of ISSR-PCR reaction for Lilium regale were optimized. The proper ISSR- PCR reaction system was determined as follows： 20 μL reaction system containing 1 x PCR buffer, 1, 25 mmol/L Mg^2＋ ,0. 4 mmol/L dNTPs, 0. 6 μmol/L primer,0. 5 U Taq DNA polymerase ,20 ng template DNA. The amplification was after 1 cycle initial denaturation at 94℃ for 5 rain, followed by 45 cycles of 40 s at 94℃, 45s annealing at 52. 8℃, and extension of 1.5 rain at 72℃, and a final 8min extension at 72℃, after which the samples were held at 4℃. The establishment of optimal system could settle a favorable basis for the further study on genetic diversity of Lilium regale using ISSR molecular marker.
出处
《林业科技开发》
2007年第6期19-22,共4页
China Forestry Science and Technology
基金
上海市重点攻关"百合种质创新与品种快繁技术研究"项目(沪农科攻字2004
第1-1号)
2005年江苏省高校自然科学基金项目(编号:05KJB180050)
关键词
岷江百合
ISSR
反应体系
优化
Liliurn regale
ISSR
Reaction system
Optimization
