Modified paired end rapid library preparation protocol for 454 GS Junior 8 kb library preparation using Covaris g-tubes and BluePippin electrophoresis.

Hill, J., Chaban, B.L., Town, J.R., Links, M.G., and Dumonceaux, T.J. (2014). "Modified paired end rapid library preparation protocol for 454 GS Junior 8 kb library preparation using Covaris g-tubes and BluePippin electrophoresis.", Protocol Exchange. doi : 10.1038/protex.2014.028  Access to full text

Abstract

This protocol describes an alternative approach to performing Roche’s Paired End Rapid Library Preparation Method for 8 kb span libraries. This method uses the Corvaris g-tube for DNA fragmentation, eliminating the need for a HydroShear apparatus, and a Sage Science BluePippin electrophoresis platform to size select the 8 kb fragments, eliminating the need for agarose gel electrophoresis and sample recovery using a Whatman Elutrap device. These two modifications allow the protocol to be completed in significantly less time (eliminate an overnight step) and require significantly less capital investment in laboratory equipment. This modified library preparation protocol has yielded paired end libraries meeting all the quality control benchmarks the original protocol stipulates and has been combined with sequencing on a GS Junior instrument to generate excellent sequencing results. The equipment substitutions presented allow paired end library preparation to be practical for smaller research groups.

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