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北方园艺  2012 

菊花SSR-PCR反应体系的建立和优化

, PP. 127-131

Keywords: 菊花,SSR标记,正交设计,反应体系优化

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Abstract:

为快速确定菊花SSR反应体系,利用正交实验设计L?16(45)对菊花基因组SSR-PCR反应体系的5个因素(模板DNA、Mg2+、dNTP、引物和Taq酶)在4个水平上进行正交设计,筛选出适合菊花的最佳SSR-PCR反应体系,进一步利用单因素完全随机试验筛选各反应因素的最佳水平。结果表明建立菊花基因组DNASSR-PCR反应体系为25μL60ng模板DNA、2.0mmol/LMg2+、0.1mmol/LdNTP、0.3μmol/L引物、1UTaq酶。并对菊花引物进行梯度退火试验,其最佳退火温度在53.1℃;扩增程序是95℃预变性5min;32个循环的94℃变性50s、53.1℃退火50s、72℃延伸50s;72℃延伸8min,4℃保存。该体系的建立为今后菊花SSR分析奠定了基础。

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