芹菜RAPD反应体系的优化研究  

The Optimization of RAPD Reaction System in Celery

作　　者：马建华[1] 杨艳君[1] 郭生金[1] 胡变芳[1] 

机构地区：[1]晋中学院,山西榆次030600

出　　处：《中国农学通报》2011年第13期204-207,共4页Chinese Agricultural Science Bulletin

摘　　要：本试验以一般RAPD反应程序为基础,采用单因素递进筛选方法,针对TaqDNA聚合酶、Mg2+、dNTPs、随机引物和DNA模板5个主要影响因素,分别设置5个不同浓度梯度,对芹菜进行RAPD-PCR扩增,建立了芹菜RAPD技术最优体系。结果表明:25μL反应体系中含TaqDNA聚合酶1.0U、Mg2+3.0mM、dNTPs0.2mM、引物28ng、模板DNA70ng,10×PCRBuffer2.5μL;扩增程序为:94℃预变性5min,94℃变性1min,36℃退火1min,72℃延伸1min,进行42个循环,最后72℃延伸10min。In the present study,we set up separately five different concentration gradient to five main influence factors （dNTPs,Taq DNA polymerase,Mg 2＋,random primer and DNA template）.Then based on the common RAPD reaction system,using the method of single factor progressive screening,we conducted the RAPD-PCR amplification of celery.Finally,we got the optimization of RAPD reaction system in celery.Conclusion of the optimized contents of 25 μL RAPD reaction system was as follows：Taq DNA polymerase 1.0 U,Mg 2＋ 3.0 mM,dNTPs 0.2 mM,primer 28 ng,DNA templet 70 ng,10×reaction Buffer 2.5 μL.The optimized PCR cycle program was as follows：94℃ for 5 min,94℃ for 1 min,36℃ for 1 min,72℃ for 1 min,42 cycles and at final 72℃ for 10 min.

关 键 词：芹菜 RAPD 优化 反应体系 

分 类 号：S636.3[农业科学—蔬菜学]