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

作者：Garza Erin; Finan Chris; Iverson Andrew; Zhou Shengde^*

来源：World Journal of Microbiology and Biotechnology, 2011, 27(2): 449-451.

DOI：10.1007/s11274-010-0451-2

• Biobutanol
• Clostridium acetobutylicum
• Low GC microorganism
• PCR

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 (> 5 kb) from low GC microorganisms using the standard PCR protocols. In this brief communication, we report a modified PCR method with an extension temperature of 60A degrees C, which efficiently amplified a 5.3 and a 5.5 kb DNA fragment (an extension time of 6 min) from a low GC bacterium Clostridium acetobutylicum (similar to 30% GC). A lower than normal extension temperature (72A degrees C) approach may also facilitate PCR amplification of large DNA fragments (> 5 kb) from other low GC microorganisms.

• 出版日期2011-2