Comparative study of Epstein-Barr virus and SV40-based shuttle-expression vectors in human repair-deficient cells.
Shuttle vectors and expression vectors have been used in human cells to examine various aspects of DNA repair including effects of DNA damage on mutagenesis, transcription, replication and recombination. A combined shuttle-expression system should provide further advantages for the stable expression of and perhaps selection/rescue strategies for DNA repair genes. We describe 2 such systems. The first is a simian virus 40 (SV40) shuttle system which allows a quasi-stable episomal vector/host relationship in which the shuttle vector may be recovered in extrachromosomal DNA preparations many months after transfection and selection but in which a high proportion of the plasmids rescued in bacteria are heavily mutated and rearranged. Secondly, we describe Epstein-Barr virus-based shuttle-expression vectors which exist as stable, multicopy episomes in human cells. Using a reporter gene and a metal-inducible promoter we have obtained low basal and very high induced expression from episomal vectors in a variety of human cells including xeroderma pigmentosum and ataxia telangiectasia cell lines. This should facilitate many molecular genetic experiments in human cells and may have particular application to molecular cloning, expression and analysis of DNA repair genes.
James
M R
MR
Laboratory of Molecular Genetics, Institut de Recherches Scientifiques sur le Cancer, Villejuif, France.
Stary
A
A
Daya-Grosjean
L
L
Drougard
C
C
Sarasin
A
A
D000818:Animals; D002460:Cell Line; D002522:Chlorocebus aethiops; D004260:DNA Repair; D004261:DNA Replication; D005786:Gene Expression Regulation; D005822:Genetic Vectors; D004854:Herpesvirus 4, Human / Q000235:genetics*; D006801:Humans; D010957:Plasmids; D013539:Simian virus 40 / Q000235:genetics*; D013997:Time Factors; D014162:Transfection; D014779:Virus Replication