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Strain Engineering

Volume 765 of the series Methods in Molecular Biology pp 409-417

Date:

High-Throughput Transposon Mutagenesis of Corynebacterium glutamicum

  • Nobuaki SuzukiAffiliated withResearch Institute of Innovative Technology for the Earth (RITE)
  • , Masayuki InuiAffiliated withResearch Institute of Innovative Technology for the Earth (RITE)
  • , Hideaki YukawaAffiliated withResearch Institute of Innovative Technology for the Earth (RITE) Email author 

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Abstract

Construction of gene disruption mutants and analysis of the resultant phenotypes are an important strategy to study gene function. A simple and high-throughput method developed for microorganisms combines two different types of transposons, direct genomic DNA amplification and thermal asymmetric interlaced-PCR. The considerable utility of this approach is demonstrable in Corynebacterium glutamicum, where 18,000 transposon disruptants enabled the generation of an insertion library covering nearly 80% of the organism’s 2,990 ORFs.

Key words

Transposon Mutagenesis Genome Disruptant library TAIL-PCR Corynebacterium glutamicum High throughput Phi29 Rolling circle DNA amplification