河南农业科学2018，Vol.47Issue(1)：7-11,5.DOI:10.15933/j.cnki.1004-3268.2018.01.002
旱稻Tail-PCR技术主要影响因素的优化和验证
Optimizing and Confirming of Main Influence Factors of Tail-PCR for Upland Rice
摘要
Abstract
Tail-PCR about transgenic upland rice was analyzed and optimized,including the quality of DNA samples,dilution degree of amplification products,enzyme types,degenerate primer combinations,primer preferences,etc.The results showed that the quality of DNA sample was an important factor for the success of the experiment.Low-purity sample led to an abortive amplification.The result of the next round amplification was affected by the dilution of the product of the previous round,and appropriately increasing product diluted multiples could increase the belts in the next round of amplification.The effect of LA Taq enzyme in this experiment was not significantly different from that of ordinary Taq enzyme,therefore the normal Taq enzyme can be used to amplify effectively.The effect of amplification with mixed degenerate primers in upland rice was not better than that of separate amplification.Moreover,it was found that degenerate primers had preference in different species.The conventional four kinds of degenerate primers could all effectively combine with the genome of the upland rice.The optimized Tail-PCR about upland rice in our studies is of stability and good repeatability.It can be accurate to obtain flanking sequence of insertion site in transgenic upland rice.关键词
旱稻/Tail-PCR技术/优化/影响因素Key words
Upland rice/Technology of Tail-PCR/Optimizing/Influence factors分类
农业科技引用本文复制引用
朱昀,李朝炜,刘嗣凡,刘颖,魏景芳..旱稻Tail-PCR技术主要影响因素的优化和验证[J].河南农业科学,2018,47(1):7-11,5.基金项目
国家转基因生物新品种培育科技重大专项(2016ZX08001003-006) （2016ZX08001003-006）
河北科技大学五大平台开放基金课题(SW21,2015PT51) （SW21,2015PT51）
河北省高等学校科学技术研究项目(QN2017071) （QN2017071）
