High-frequency transformation of human repair-deficient cell lines by an Epstein-Barr virus-based cDNA expression vector

Gene. 1991 Nov 15;107(2):279-84. doi: 10.1016/0378-1119(91)90328-9.

Abstract

We constructed a human cDNA expression vector by combining an episomal Epstein-Barr virus (EBV) vector with the expression cassette from the transient-expression vector, pCDM8. This new vector, designated pEBS7, exhibited high-level expression of reporter genes in normal and repair-deficient xeroderma pigmentosum cell lines. Reconstruction experiments indicated that marker genes diluted to a frequency of 10(-5) can be rescued on a single transfection dish. Moreover, derivative cell lines that constitutively express the gene encoding EBV nuclear antigen 1 exhibited a tenfold enhancement in the frequency of rescue of marker genes. The feasibility of preparing large-scale directional or nondirectional cDNA libraries in pEBS7 was demonstrated and reconstruction experiments indicated that marker genes could be rescued from either library with equal efficiency. These results establish a high-efficiency system for the isolation of genes by direct phenotypic selection in human mutant cell lines.

MeSH terms

  • Antigens, Viral / genetics
  • Base Sequence
  • Cell Line
  • DNA Repair / genetics*
  • Epstein-Barr Virus Nuclear Antigens
  • Gene Expression / genetics
  • Gene Library
  • Genetic Vectors / genetics*
  • Herpesvirus 4, Human / genetics*
  • Herpesvirus 4, Human / immunology
  • Humans
  • Molecular Sequence Data
  • Phenotype
  • Plasmids / genetics
  • Transfection*
  • Xeroderma Pigmentosum / genetics*

Substances

  • Antigens, Viral
  • Epstein-Barr Virus Nuclear Antigens