Novel use of synthetic oligonucleotide insertion mutants for the study of homologous recombination in mammalian cells.

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Abstract
Thymidine kinase-deficient mouse L cells were transformed with plasmid DNA carrying 8 base-pair Xho I linker insertion mutations in the coding region of the herpes simplex virus type 1 thymidine kinase gene. When the mutant plasmids are introduced individually into LTK- cells, transformation efficiencies are greatly reduced relative to the wild type. When 2 mutant plasmids are cotransferred into the same LTK- recipients, significantly higher frequencies of transformation are observed (30-300 times). The usefulness of linker insertions for the study of homologous recombination in detecting the existence of normal thymidine kinase gene sequences (i.e., sequences lacking the insertions after recombination are substantiated by DNA .cntdot. DNA hybridization) is demonstrated. The frequencies of recombination of the various crosses are consistent with the known positions of the mutations.