A model for testing recombinogenic sequences in the mouse germline.
Homologous recombination is a conserved process of genetic exchange generated by homologous pairing of nucleotides. Species diversity and gene evolution are dependent on the outcomes of recombination during germ cell development, yet systems to study mammalian germline recombination, especially those with applications to human genetics, are not well developed. We report on a transgenic mouse system designed to study recombination within test sequences in the male germline utilizing an intron-interrupted lacZ reporter gene. beta-galactosidase positive sperm are detected and quantitated by flow cytometry using fluorogenic substrates. Examination of recombination within a 1.7 kb repeat of test sequences derived from the human glycophorin breakpoint cluster region detects approximately 0.04-0.09% fluorescent sperm. Confirmation that these sperm result from recombination in the germline comes from histochemical staining of testicular cells, examination of spliced mRNA, and PCR analysis of sorted sperm populations. The system is readily adaptable to studies of other sequences reported to have elevated levels of recombination, including those implicated in human genetic disease. Investigations of the molecular basis for genomic instability at specific chromosomal locations may yield important insights into mechanisms of chromosomal loss and rearrangements.
Moynahan
M E
ME
Department of Medicine, Memorial Sloan-Kettering Cancer Center, New York, NY 10021, USA.
Akgün
E
E
Jasin
M
M
D000818:Animals; D002469:Cell Separation; D003434:Crossing Over, Genetic / Q000235:genetics; D004274:DNA, Recombinant / Q000032:analysis; D005434:Flow Cytometry; D015870:Gene Expression; D017930:Genes, Reporter / Q000235:genetics; D006021:Glycophorins / Q000235:genetics; D006801:Humans; D007438:Introns / Q000235:genetics; D007763:Lac Operon / Q000235:genetics; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D008957:Models, Genetic; D012326:RNA Splicing; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D011995:Recombination, Genetic; D013094:Spermatozoa / Q000737:chemistry; D013728:Terminator Regions, Genetic / Q000235:genetics
D004274:DNA, Recombinant; D006021:Glycophorins; D012333:RNA, Messenger