EXTRACHROMOSOMAL UNEQUAL HOMOLOGOUS RECOMBINATION AND GENE CONVERSIONIN SIMIAN KIDNEY-CELLS - EFFECTS OF UV DAMAGE

Shuttle plasmid Vectors containing the SV40 origin of replication and tandem neo genes with distally placed non-overlapping deletions were u sed to study the effects of DNA damage on extrachromosomal homologous recombination in simian kidney cells. DNA was introduced into COS7 cel ls by a lipofectin-mediated transfection procedure and recombination w as assessed by analyzing the structure of plasmids. Recombinational ev ents observed included unequal homologous recombination (triplication) , gene conversion, double reciprocal recombination, deletion (pop-outs ), gene amplification (4-6 copies), and multimerization. Triplication, an event that previously had not been reported in association with ex trachromosomal recombination, predominated in experiments with undamag ed vectors. The recombination frequency (Neo(R)/Amp(R)) of vectors ran domly damaged by UV irradiation was essentially unchanged; however, th e relative number of triplication events decreased significantly. Sele ctive damage in one of the two reo genes increased the relative freque ncy of gene conversion. The experimental system developed for use in t his study detects all major homologous recombination events observed i n chromosomal direct repeat sequences in mammalian cells and yeast and should prove valuable for future studies of homologous recombination in mammalian cells. (C) 1998 Elsevier Science B.V.