Extension temperature of 60 degree C required for PCR amplification of large DNA fragments (>5kb) from a low GC bacterium Clostridium acetobutylicum

PCR amplification of DNA fragments has been routinely used in gene cloning and engineering of microbial strains for biotechnological purposes such as production of biofuels and green chemicals. However, it is often a challenge to amplify large DNA fragments (>5kb) from low GC microorganisms using...

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Bibliographic Details
Published inWorld journal of microbiology & biotechnology Vol. 27; no. 2; pp. 449 - 451
Main Authors Garza, Erin, Finan, Chris, Iverson, Andrew, Zhou, Shengde
Format Journal Article
LanguageEnglish
Published 01.02.2011
Subjects
Bacteria
Clostridium acetobutylicum
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Summary:PCR amplification of DNA fragments has been routinely used in gene cloning and engineering of microbial strains for biotechnological purposes such as production of biofuels and green chemicals. However, it is often a challenge to amplify large DNA fragments (>5kb) from low GC microorganisms using the standard PCR protocols. In this brief communication, we report a modified PCR method with an extension temperature of 60 degree C, which efficiently amplified a 5.3 and a 5.5kb DNA fragment (an extension time of 6min) from a low GC bacterium Clostridium acetobutylicum (~30% GC). A lower than normal extension temperature (72 degree C) approach may also facilitate PCR amplification of large DNA fragments (>5kb) from other low GC microorganisms.
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ISSN:0959-3993
1573-0972
DOI:10.1007/s11274-010-0451-2