In this report we present an experimental scheme that facilitates the study of homologous recombination between closely linked genes in cultured mammalian cells. Two different Xho I linker insertion mutants of the herpes simplex virus type 1 thymidine kinase (HTK) gene were introduced into mouse LTK- cells as direct repeats on a plasmid carrying a dominant selectable marker. Following stabilization of these sequences in the recipient cell, selection for TK+ was applied to detect recombinational events between different TK- genes. TK+ segregants were observed at a frequency of 10-4-10-5 in lines harboring both mutant genes. Control lines carrying only one type of mutant HTK gene yielded TK+ cells at frequencies of 10-7 or less. Physical analysis of the TK+ segregants has revealed the presence of an apparently normal HTK gene that is resistant to Xho I endonuclease digestion in each TK+ line examined. Analyses of the TK gene pairs before and after recombination suggest that at least 50% of the recombinants are the result of nonreciprocal exchanges of genetic information, or gene conversion events.
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)