Molecular cloning of the human nucleotide-excision-repair gene ERCC4

Larry H. Thompson, Kerry W. Brookman, Christine A. Weber, Edmund P. Salazar, Joyce T. Reardon, Aziz Sancar, Zuoming Deng, Michael J. Siciliano

Research output: Contribution to journalArticlepeer-review

40 Scopus citations

Abstract

ERCC4 was previously identified in somatic cell hybrids as a human gene that corrects the nucleotide-excision-repair deficiency in mutant hamster cells. The cloning strategy for ERCC4 involved transfection of the repair- deficient hamster cell line UV41 with a human sCos-1 cosmid library derived from chromosome 16. Enhanced UV resistance was seen with one cosmid-library transformant and two secondary transformants of UV41. Cosmid clones carrying a functional ERCC4 gene were isolated from a library of a secondary transformant by selecting in Escherichia coli for expression of a linked neomycin-resistance gene that was present in the sCos-1 vector. The cosmids mapped to 16p13.13-p13.2, the location assigned to ERCC4 by using somatic cell hybrids. Upon transfection into UV41, six cosmid clones gave partial correction ranging from 30% to 64%, although all appeared to contain the complete gene. The capacity for in vitro excision of thymine dimers from a plasmid by transformant cell extracts correlated qualitatively with enhanced UV resistance.

Original languageEnglish (US)
Pages (from-to)6855-6859
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume91
Issue number15
DOIs
StatePublished - Jul 19 1994
Externally publishedYes

Keywords

  • cell line UV41
  • DNA repair
  • excinuclease
  • genetic complementation
  • UV resistance

ASJC Scopus subject areas

  • Genetics
  • General

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