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OALib Journal期刊
ISSN: 2333-9721
费用：99美元
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东方百合srap体系优化及引物筛选
DOI: 10.3969/j.issn.1000-2006.2013.03.002, PP. 6-10
Keywords: 东方百合,序列相关扩增多态性分子标记,体系优化,引物筛选
Abstract:
以东方百合商品种‘索蚌’(‘sorbonne’)和‘康斯坦撒’(‘constantsa’)及其f1代杂交群体为实验材料,从100对srap引物中筛选出59对扩增产物丰富、具有较高多态性的引物,同时对pcr反应体系进行了优化。采用正交设计,对mg2+、dntps、taq聚合酶及引物浓度进行4因素4水平优化,并在此基础上对mg2+、引物浓度和退火温度进行了单因素优化。结果表明,东方百合srap的最佳体系为:1×buffer,mg2+浓度2.7mmol/l,dntps浓度0.25mmol/l,引物浓度0.8μmol/l,taq聚合酶1.5μmol/min,模板dna60～100ng。srap两步反应最佳退火温度为第1步35℃,第2步退火59.3℃。最后用‘索蚌’、‘康斯坦撒’和部分f1植株对优化体系进行稳定性检测,发现所筛选出的59对引物在优化的体系下均有稳定的扩增产物。
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