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A PCR-based method for isolation of genomic DNA flanking a known DNA sequence

Boulter, CA; Natarajan, D; (1999) A PCR-based method for isolation of genomic DNA flanking a known DNA sequence. Current Issues in Molecular Biology , 1 (1) 47 - 52. Green open access

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Abstract

We describe a simple PCR-based method for the isolation of genomic DNA that lies adjacent to a known DNA sequence. The method is based on the directional cloning of digested genomic DNA into the multiple cloning site of a pUC-based plasmid to generate a limited genomic library. The library is plated onto a number of selective LA plates which are incubated overnight, and recombinant plasmid DNA is then isolated from resistant colonies pooled from each plate. PCR amplification is performed on the pooled recombinant plasmid DNAs using primers specific for the pUC vector and the known genomic sequence. The combination of efficient directional cloning and bacterial transformation gives relative enrichment for the genomic sequence of interest and generates a simple DNA template, enabling easy amplification by PCR.

Type: Article
Title: A PCR-based method for isolation of genomic DNA flanking a known DNA sequence
Location: England
Open access status: An open access version is available from UCL Discovery
Publisher version: http://www.horizonpress.com/cimb/
Language: English
Additional information: This article is published under the Creative Commons Attribution License (CC BY). Under this license, authors retain ownership of the copyright for their article, but authors allow anyone to download, reuse, reprint, modify, distribute, and/or copy articles as long as the original authors and source are credited.
Keywords: DNA, Gene Library, Genome, Plasmids, Polymerase Chain Reaction
UCL classification: UCL
UCL > Provost and Vice Provost Offices
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Population Health Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Population Health Sciences > UCL GOS Institute of Child Health
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Population Health Sciences > UCL GOS Institute of Child Health > Developmental Biology and Cancer Dept
URI: https://discovery.ucl.ac.uk/id/eprint/1314497
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