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油桐ISSR-PCR最佳反应体系的建立

, PP. 194-199

Keywords: 油桐,ISSR-PCR,单因子试验,正交设计

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Abstract:

在油桐遗传多样性研究中,为了获得清晰可靠、重复性强的ISSR扩增结果,综合采用单因子试验和正交设计两种方法对影响油桐ISSR-PCR反应结果的4个因素( Taq酶、Mg2+ 、dNTP、引物)进行优化试验。单因子试验分别研究各因素对ISSR-PCR反应的影响,找出最佳反应水平。正交设计选用L9 (34 )方案,采用直观分析获得影响因素最佳反应水平。通过综合比较分析,最终建立了油桐ISSR2PCR反应的最佳反应体系,即在20μL反应体系中, Taq DNA聚合酶1. 0 U,Mg2+ 2. 0 mmol?L-1 , dNTP 0. 20 mmol?L-1 ,引物0. 4μmol?L-1 , 1 ×PCR缓冲液, 40 ng模板DNA。

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