Single-step overlap-primer-walk polymerase chain reaction for multiple mutagenesis without overlap extension.

Kumar, Raj and Rajagopal, Kammara (2008) Single-step overlap-primer-walk polymerase chain reaction for multiple mutagenesis without overlap extension. Analytical biochemistry, 377 (1). pp. 105-107. ISSN 1096-0309

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Abstract

We present a simple, single-step, single-tube, and rapid method for introducing a series of mutations into cloned DNA. Polymerase chain reaction (PCR)-based mutagenesis methods have become very prevalent due to their simplicity and efficiency for introducing mutations. Our method, overlap-primer-walk PCR, has several advantages over other published methods. It uses two common oligodeoxyribonucleotides and a series of overlapping primers specific for various mutations. Once common flanking primers are selected, two to three mutations require only one additional primer. Therefore, this method is very useful for introduction of multiple mutations in various sites of the target DNA. We illustrate the usefulness of the method by introducing several mutations into the human TNF-alpha encoding gene.

Item Type:	Article
Additional Information:	copyright of this article belongs to Elsevier Science
Subjects:	Q Science > QD Chemistry
Depositing User:	Dr. K.P.S.Sengar
Date Deposited:	30 Nov 2011 11:37
Last Modified:	30 Nov 2011 11:37
URI:	http://crdd.osdd.net/open/id/eprint/610

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