High-Throughput Transposon Mutagenesis of Corynebacterium glutamicum
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 wordsTransposon Mutagenesis Genome Disruptant library TAIL-PCR Corynebacterium glutamicum High throughput Phi29 Rolling circle DNA amplification
We wish to thank Dr. C. Omumasaba for critical reading of the manuscript. This research was partly supported by New Energy and Industrial Technology Development Organization (NEDO), Japan.
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