In vivo DNA double-strand breaks enhance gene targeting in cultured silkworm cells

Hiroaki Mon, Takahiro Kusakabe, Jae Man Lee, Yutaka Kawaguchi, Katsumi Koga

Research output: Contribution to journalArticlepeer-review

10 Citations (Scopus)

Abstract

Alteration of genomic information through homologous recombination (HR) is a powerful tool for reverse genetics in bacteria, yeast, and mice. The low frequency of HR is, however, a major obstacle to achieve efficient gene targeting. In this study, we have developed an assay system for investigating the frequency of gene targeting in cultured silkworm cells using a firefly luciferase gene as a reporter. The introduction of a DNA double-strand break (DSB) either in the chromosomal target locus or in the targeting construct drastically increased the frequency of gene targeting. Interestingly, the inhibition of poly(ADP-ribose) polymerase (PARP), a protein known to play an important role in overall suppression of the HR pathway, stimulated the targeting efficiency, whereas the overexpression of two silkworm RecA homologs, BmRad51 and BmDmc1, had no effect. The presently devised assay system may serve as a useful tool to improve the gene targeting efficiency in the silkworm (Bombyx mori).

Original languageEnglish
Pages (from-to)99-106
Number of pages8
JournalComparative Biochemistry and Physiology - B Biochemistry and Molecular Biology
Volume139
Issue number1
DOIs
Publication statusPublished - Sep 1 2004

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Physiology
  • Molecular Biology

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