限制性酶切片段差异显示及其应用  

A new differential display PCR——Restriction fragment differential display PCR

作　　者：胡岳山[1] 李杰芬[1] 王剑[1] 

机构地区：[1]广州中医药大学生化教研室,中基公共实验室,广州510405

出　　处：《生物技术通讯》2004年第1期65-66,共2页Letters in Biotechnology

摘　　要：差异显示技术(DD)-PCR是一种研究基因表达差异的重要而应用广泛的方法,传统的差异显示法由于在PCR时采用Poly穴T雪引物和随机引物而导致较高的假阳性率和产物的近Poly穴A雪非编码区的大量扩增。改进后的限制性酶切片段差异显示技术(RFDD)-PCR采用TaqI酶切双链cDNA,连上特殊设计的接头,再用经特殊设计的特异性配对于接头的引物来扩增,因此能重点扩增编码区并能极大地消除假阳性率。由于扩增时引物就带有荧光或放射性核素标记,还使得差异显示条带的检测更为方便、灵敏。本文简要介绍了该法的原理、步骤、应用及其优缺点。Differential display(DD)is an important and wide-applied method in the research of gene's expression.Tradi-tional DD has some limitations that a high number of false positives occur and3'-ends of the transcripts are preferentially amplified.The restriction fragment differential display(RFDD)-PCR technique overcomes the limitations of standard differ-ential display.Instead of directly amplifying first-strand cDNA,the RFDD-PCR approach adds adaptors to the ends of the cDNA after reverse transcription.The high stringency PCR amplification step uses a series of specially designed dis-playPROBE primer which anneal to the adaptor junctions.It is convenient to detect of display profile for the primer la-beled fluorescence or radionucide.This paper introduced principle,practice steps,application and attention of this method in brief.

关 键 词：限制性酶切片段差异显示 接头 假阳性率 基因表达差异 

分 类 号：Q78[生物学—分子生物学] Q753