Multiplex PCR: Critical parameters and step-by-step protocol

被引：732

作者：

Henegariu, O ^{[1
]}

Heerema, NA ^{[1
]}

Dlouhy, SR ^{[1
]}

Vance, GH ^{[1
]}

Vogt, PH ^{[1
]}

机构：

[1] UNIV HEIDELBERG,D-6900 HEIDELBERG,GERMANY

来源：

BIOTECHNIQUES | 1997年 / 23卷 / 03期

关键词：

D O I：

10.2144/97233rr01

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

By simultaneously amplifying more than one locus in the same reaction, multiplex PCR is becoming a rapid and convenient screening assay in both the clinical and the research laboratory. While numerous papers and manuals discuss in detail conditions influencing the quality of PCR in general, relatively little has been published about the important experimental factors and the common difficulties frequently encountered with multiplex PCR. We have examined various conditions of the multiplex PCR, using a large number of primer pairs. Especially important for a successful multiplex PCR assay are the relative concentrations of the primers at the various loci, the concentration of the PCR buffer, the cycling temperatures and the balance between the magnesium chloride and deoxynucleotide concentration. Based on our experience, we propose a protocol for developing a multiplex PCR assay and suggest ways to overcome commonly encountered problems.

引用

页码：504 / 511

页数：8

共 16 条

[1] ANALYSIS OF QUANTITATIVE PCR FOR THE DIAGNOSIS OF DELETION AND DUPLICATION CARRIERS IN THE DYSTROPHIN GENE [J].