通过抑制性消减杂交技术鉴定差异基因：V. 基于PCR的DNA斑点杂交

Identification of Differential Genes by Suppression Subtractive Hybridization: V. PCR-Based DNA Dot Blot.

作者信息

Rebrikov Denis V

出版信息

CSH Protoc. 2008 Jul 1;2008:pdb.prot4859. doi: 10.1101/pdb.prot4859.

Abstract

INTRODUCTIONSuppression subtractive hybridization (SSH) is one of the most powerful and popular methods for generating subtracted cDNA or genomic DNA libraries. This technique can be used to compare two mRNA populations and obtain cDNAs representing genes that are either overexpressed or exclusively expressed in one population as compared to another. It can also be used for comparison of genomic DNA populations. This protocol describes a method for the use of PCR-based DNA dot blots in the differential screening of arrayed subtracted DNA clones. For high-throughput screening, a 96-well or 384-well format PCR from one of several thermal cycler manufacturers is recommended. Alternatively, single tubes can be used.

摘要

引言

抑制性消减杂交（SSH）是用于构建消减cDNA或基因组DNA文库的最强大且最常用的方法之一。该技术可用于比较两个mRNA群体，并获得代表与另一个群体相比在一个群体中过度表达或特异性表达的基因的cDNA。它还可用于比较基因组DNA群体。本方案描述了一种在阵列化消减DNA克隆的差异筛选中使用基于PCR的DNA斑点印迹的方法。对于高通量筛选，建议使用来自几家热循环仪制造商之一的96孔或384孔格式PCR。或者，也可以使用单管。