Targeted Integration of the Ren-1D Locus in Mouse Embryonic Stem Cells

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 89; no. 11; pp. 5020 - 5024
• Main Authors: Christopher C. J. Miller, McPheat, Jane C., Potts, Wendy J.
• Format: Journal Article
• Language: English
• Published: Washington, DC National Academy of Sciences of the United States of America 01.06.1992; National Acad Sciences; National Academy of Sciences
• Subjects: Animals; Base Sequence; Biological and medical sciences; Biotechnology; Blotting, Southern; Cells, Cultured; Cellular biology; DNA; Embryonic stem cells; Exons; Fundamental and applied biological sciences. Psychology; Gene Conversion; Gene targeting; Genes; Genetic engineering; Genetic loci; Genetic technics; Genetic vectors; Genetics; In Vitro Techniques; Methods. Procedures. Technologies; Mice; Miscellaneous; Molecular Sequence Data; Oligodeoxyribonucleotides - chemistry; Polymerase chain reaction; Recombination, Genetic; Renin - genetics; Rodents; Stem Cells; Synthetic digonucleotides and genes. Sequencing; Transfection; Transgenes; Vertebrata; Embryo; Mammalia; Renin; Gene targeting; Mouse; Enzyme; Stem cell; Rodentia; Homology
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.89.11.5020

We have introduced a Ren-1D targeting vector into embryonic stem cells containing the two highly homologous mouse renin genes Ren-1D and Ren-2. Using a polymerase chain reaction (PCR) screen designed to detect targeted integration at Ren-1D and Ren-2, we isolated 15 targeted embryonic stem cell clones, all of which had undergone a gene conversion event at the Ren-1D locus. We did not isolate any clones in which the incoming DNA had recombined with Ren-2. Over the region encompassed by our transgene, Ren-1D and Ren-2 display >95% homology. Our results suggest that the machinery driving gene targeting by means of homologous recombination in mammalian cells is capable of distinguishing between these two sequences. Construction of transgenic mice with the embryonic stem cells reported here carrying a mutated renin gene will permit a greater understanding of the functions of the Ren-1D and Ren-2 gene products and their relative contribution to cardiovascular homeostasis.