运用多种策略改良差异显示PCR

Several strategies used to improve differential display PCR

目的 :改进差异显示PCR技术 ,提高其在筛选差异表达基因方面的效率。方法 :①采用单碱基铆钉引物 ;②增加引物长度 ;③提高PCR反应的严谨性 ;④应用同步重复测序电泳。结果 :减少了经典方法的工作量 ,降低了非特异性和假阳性率。用改良技术研究热适应大鼠下丘脑基因的差异表达 ,发现了一个差异表达基因片段 ,斑点杂交证实为阳性片段。结论 :改进后的差异显示PCR技术是一种研究基因差异表达从而发现新基因或基因新功能的有效方法。

To improve the effciency of differential display PCR(DD PCR) in screening differentially expressed genes.Four strategies are used to achieve the aim including using one-base anchored primer,increasing the length of primers,carrying out PCR with high-stringency condition and resolving PCR samples repeatedly on the DNA sequencing gel simultaneously.Redundancy of DD PCR is relieved,non-specificity decreased and false positives reduced.A differentially expressed gene was identified from hypothalamus of heat acclimated rats using the improved technique,which is verified by dot blot.Improved differential display PCR is an efficient method for discovering new genes or new functions of genes.