DNA-mediated gene transfer: recombination between cotransferred DNA sequences and recovery of recombinants in a plasmid.

Abstract

Ltk- aprt- mouse L cells were transformed to the tk+ phenotype with 10 ng of the herpes simplex virus-1 thymidine kinase (tk) gene and 20 .mu.g of pBR322 or SV40 DNA. DNA from 5 cloned cell lines show restriction endonuclease fragments that hybridize to both tk and pBR322 or SV40 DNA. In all of the cell lines some of these fragments also contain cellular DNA sequences. The use of carrier DNA with defined sequences enabled demonstration that the joining of carrier and selectable gene sequences occurs in mouse cells. In 1 case the ampicillin resistance marker of pBR322 was used to rescue a recombinant plasmid. An analysis of the junction between pBR322 and tk in this plasmid suggests that a small area of homology (16 of 19 base pairs) might be involved in the recombination process.