Lack of site specific recombination of exogenous DNA in mouse L cells

Plasmids were constructed containing the HSV thymidine kinase gene and two copies of X. borealis 5S rDNA. Mouse L TK- cells were transformed with these DNAs, with selection for the TK+ gene. Transformed cells were then analyzed by Southern blot hybridization and hybridization in situ to determine whether integration of the exogenous DNA occurred at regions of chromosomal homology i.e., at the 5S rDNA regions. Four cell lines were analyzed by Southern blots. Differences in restriction endonuclease specificity strongly suggested that integration was at a different site in each cell line. Two cell lines were further analyzed by hybridization in situ; each showed a single integration site, both different from each other and different from the mouse L cell 5S rDNA sites. Therefore, the presence of two copies of the 5S rDNA gene in the DNA introduced by gene transfer and approximately 300-350 copies of the mouse 5S rDNA gene was not sufficient in these experiments to produce homologous integration into a specific site.