Ligation-anchored PCR: a Simple Amplification Technique with Single-sided Specificity

Overview

Journal Proc Natl Acad Sci U S A

Specialty Science

Date 1992 Oct 15

PMID 1409706

Citations 74

Authors

A B Troutt

M G McHeyzer-Williams

B Pulendran

G J Nossal

Affiliations

Soon will be listed here.

Abstract

A simple, efficient, and sensitive technique has been developed for amplification of cDNAs encoding molecules with 5' regions of unknown sequence. In this ligation-anchored PCR, T4 RNA ligase is used to covalently link an "anchor" oligonucleotide to first-strand cDNAs. These anchored cDNAs are then amplified by using one PCR primer specific for the anchor and another specific for a sequence within the molecule of interest. The anchor oligonucleotide has been especially designed to facilitate subsequent analysis and cloning of the resultant PCR products. This three-stage procedure does not require purification of product between steps and avoids many of the technical difficulties associated with established anchored PCR protocols. The efficacy of ligation-anchored PCR was demonstrated by amplification of a specific IgG1 cDNA; total RNA equivalent to as few as 100 cells yielded the expected PCR product.

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