河南农业科学2012，Vol.41Issue(1)：65-68,4.
土壤微生物总DNA的V3可变区PCR反应体系优化
Optimization of V3 Region PCR Reaction System with Soil Microbial Total DNA as Template
摘要
Abstract
The optimization of PCR reaction system and condition for 16S rDNA V3 region was carried out with soil microbial total DNA as template,which was extracted using E. Z. N. A. Soil DNA kit. The PCR reaction was optimized from the aspects of DNA template concentration,primer concentration, annealing temperature,and hot-start. According to the results,the most suitable PCR system was established in a 50 μL reaction volume containing 10. 5 ng of DNA template,5μL of 10XPCR buffer,4μL of 2. 5 mmol/L dNTP,1. 5μL of 10μmol/L primers for each,2. 5U of Taq DNA polymerase. The reaction was built up using the conditions:94 ℃ for 2 min;29 cycles of 94 ℃ for 60 s,52 ℃ for 60 s,72 ℃ for 70 s;72 t℃ for 5min. The results demonstrated that hot-start PCR and suitable annealing temperature were important to get high-quality PCR products.关键词
土壤微生物DNA/16S rDNA/V3区/PCR扩增/优化/热启动方式/退火温度Key words
soil microbial DNA/ 16S Rdna/V3 region/PCR amplification/optimization/hot-start PCR/annealing temperature分类
农业科技引用本文复制引用
殷全玉,郭夏丽,赵铭钦,王岩..土壤微生物总DNA的V3可变区PCR反应体系优化[J].河南农业科学,2012,41(1):65-68,4.基金项目
吉林省烟草工业有限责任公司重大科技攻关项目(JY2006012) （JY2006012）
