数字农科院2.0

高GC含量青枯菌aac基因PCR扩增体系的建立与优化

文献类型: 中文期刊

作者: 张杨;徐进;许景升;何礼远;冯洁

作者机构:

关键词: 高GC;PCR;正交试验

期刊名称:植物保护

ISSN: 0529-1542

年卷期: 2008 年 34 卷 02 期

页码: 90-93

收录情况: 北大核心

摘要: 通过添加增效剂、正交试验设计优化PCR反应体系、联合采用多种PCR程序等措施,建立并优化了PCR扩增体系,成功地从高GC青枯菌基因组中扩增出了长度为2 434 bp且GC含量高达70.9%的aac基因。PCR反应体系为20μL包括5%DMSO2、.5 mmol/L MgCl2、500μmol/L dNTP、10 pmol/L引物1、.25 UTaq酶、50 ng模板DNA。首先采用热启动PCR:95℃5 min,保持80℃,加入Taq酶;然后采用二步PCR:5个循环包括变性95℃1 min,65℃1 min;最后采用降落PCR:30个循环为95℃1 min,78℃1 min,每个循环降低0.5℃,72℃3 min;补充10个循环为95℃1 min,63℃1 min,72℃3 min;72℃10 min。该试验体系的建立与优化为研究高GC含量生物的基因功能提供了方法。

分类号: S432.4

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