An efficient gene-disruption method in Cryptococcus neoformans by double-joint PCR with NAT-split markers

Min Su Kim, Seo Young Kim, Ja Kyung Yoon, Yin Won Lee, Yong Sun Bahn

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51 Citations (Scopus)


Targeted gene disruption via biolistic transformation and homologous recombination is a method widely used to identify and investigate the function of genes in Cryptococcus neoformans that causes fatal fungal meningitis if not timely treated. Currently, most laboratories employ the overlap-PCR method to generate a gene-disruption cassette with dominant selectable markers, such as nourseothricin acetyltransferase (NAT). However, the conventional overlap-PCR method is often found to be inefficient because of the presence of multiple templates and of the long length of the final overlap-PCR products. In this report, we suggested an efficient gene-disruption method for C. neoformans, termed a double-joint PCR with NAT-split markers. Here we demonstrated that the gene-disruption cassette generated using double-joint PCR with NAT-split markers can be used successfully for targeted C. neoformans gene disruption with the advantages of providing a more convenient construction of gene-disruption cassettes and high targeted-integration frequency.

Original languageEnglish
Pages (from-to)983-988
Number of pages6
JournalBiochemical and Biophysical Research Communications
Issue number3
Publication statusPublished - 2009 Dec 18


All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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