Construction of a novel expression system on a human artificial chromosome

Akihiro Otsuki, Candice G.T. Tahimic, Nozomi Tomimatsu, Motonobu Katoh, David J. Chen, Akihiro Kurimasa, Mitsuo Oshimura

Research output: Contribution to journalArticlepeer-review

19 Scopus citations

Abstract

Efficient regulation of transgene would greatly facilitate the analysis of gene function in biological systems for basic research and clinical applications. The tetracycline-regulatable system (TRS) has proven to be a promising tool for such purposes. Despite their widespread application, a number of challenges are still associated with the use of TRS, including clonal variability in the regulation and copy number. We have recently constructed a novel human artificial chromosome (HAC) called 21ΔqHAC. By housing a TRS-based DNA-PKcs expression cassette in this HAC, we were able to circumvent the problems associated with conventional TRS-based vectors. We achieved tight control of DNA-PKcs expression and rescued the radiosensitive phenotype of DNA-PKcs-deficient CHO cells. The combined use of HAC and the TRS serves as a model for controllable and fixed copy number expression vectors. Our study also demonstrates the suitability of the HAC to accommodate multi-subunit constructs such as that of the TRS.

Original languageEnglish (US)
Pages (from-to)1018-1025
Number of pages8
JournalBiochemical and Biophysical Research Communications
Volume329
Issue number3
DOIs
StatePublished - Apr 15 2005
Externally publishedYes

Keywords

  • DNA repair
  • DNA-PKcs
  • Expression control
  • Human artificial chromosome
  • Insulator
  • Microcell-mediated chromosome transfer
  • Tetracycline-regulatable system
  • V3

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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