利用逆转录聚合酶链反应分析单个细胞中的大量信使核糖核酸种类。

The use of reverse transcription polymerase chain reaction to analyse large numbers of mRNA species from a single cell.

作者信息

Toellner K M, Scheel-Toellner D, Seitzer U, Sprenger R, Trümper L, Schlüter C, Flad H D, Gerdes J

机构信息

Department of Immunology and Cell Biology, Forschungsinstitut Borstel, Germany.

出版信息

J Immunol Methods. 1996 May 10;191(1):71-5. doi: 10.1016/0022-1759(96)00006-3.

DOI:10.1016/0022-1759(96)00006-3

PMID:8642203

Abstract

A PCR method is described for determining the expression of multiple heterogeneous mRNAs from single cells. The total mRNA pool of a single selected cell is subjected to reverse transcription and subsequent tailing with poly(dA). This cDNA is preamplified by a sequence non-specific PCR protocol using oligo(dT)-containing primers. The single cell cDNA library obtained permits the analysis of virtually unlimited numbers of mRNA species per cell using sequence-specific PCR. This procedure of multiple mRNA analysis enables phenotyping of any cell for its mRNA composition and could be used to study the cytokine mRNA expression of individual human T cells ex vivo. The method should greatly facilitate the analysis of combinatorial expression of known genes in any cell.

摘要

描述了一种用于确定单个细胞中多种异源mRNA表达的PCR方法。对单个选定细胞的总mRNA库进行逆转录，随后用聚（dA）加尾。使用含寡聚（dT）的引物，通过序列非特异性PCR方案对该cDNA进行预扩增。获得的单细胞cDNA文库允许使用序列特异性PCR分析每个细胞中几乎无限数量的mRNA种类。这种多mRNA分析程序能够对任何细胞的mRNA组成进行表型分析，可用于体外研究个体人类T细胞的细胞因子mRNA表达。该方法应极大地促进对任何细胞中已知基因组合表达的分析。