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. 1992 Aug 11;20(15):3815–3820. doi: 10.1093/nar/20.15.3815

Targeting of the creatine kinase M gene in embryonic stem cells using isogenic and nonisogenic vectors.

J van Deursen 1, B Wieringa 1
PMCID: PMC334053  PMID: 1508665

Abstract

Replacement vectors with genomic DNA originating from different mouse strains were used to introduce site-specific mutations into the creatine kinase M (CKM) gene of mouse embryonic stem (ES) cells. Here we demonstrate that in mouse strain 129-derived ES cells, the gene is at least 25-fold more efficiently targeted with an isogenic, 129-derived vector (129-pRV8.3) than with a nonisogenic, BALB/c-specific vector (BALB/c-pRV8.3). The two targeting constructs were identical except for allelic differences which were typed by partial sequencing. These included base pair mismatches (2%) and a polymorphic [GTC]-repeat length variation. Both in separate transfections as well as in cotransfections with mixed vectors, homologous disruption of the CKM gene resulted uniquely from the 129-isogenic DNA. Our data confirm earlier observations on requirements for homologous recombination in pro- and eukaryotic systems and indicate that targeting of the CKM locus is highly sensitive to small sequence differences between cognate segments in the endogenous and incoming DNA.

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Selected References

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  1. Belt P. B., Groeneveld H., Teubel W. J., van de Putte P., Backendorf C. Construction and properties of an Epstein-Barr-virus-derived cDNA expression vector for human cells. Gene. 1989 Dec 14;84(2):407–417. doi: 10.1016/0378-1119(89)90515-5. [DOI] [PubMed] [Google Scholar]
  2. Charron J., Malynn B. A., Robertson E. J., Goff S. P., Alt F. W. High-frequency disruption of the N-myc gene in embryonic stem and pre-B cell lines by homologous recombination. Mol Cell Biol. 1990 Apr;10(4):1799–1804. doi: 10.1128/mcb.10.4.1799. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. DeChiara T. M., Efstratiadis A., Robertson E. J. A growth-deficiency phenotype in heterozygous mice carrying an insulin-like growth factor II gene disrupted by targeting. Nature. 1990 May 3;345(6270):78–80. doi: 10.1038/345078a0. [DOI] [PubMed] [Google Scholar]
  4. Dorin J. R., Inglis J. D., Porteous D. J. Selection for precise chromosomal targeting of a dominant marker by homologous recombination. Science. 1989 Mar 10;243(4896):1357–1360. doi: 10.1126/science.2538001. [DOI] [PubMed] [Google Scholar]
  5. Hasty P., Rivera-Pérez J., Bradley A. The length of homology required for gene targeting in embryonic stem cells. Mol Cell Biol. 1991 Nov;11(11):5586–5591. doi: 10.1128/mcb.11.11.5586. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Hasty P., Rivera-Pérez J., Chang C., Bradley A. Target frequency and integration pattern for insertion and replacement vectors in embryonic stem cells. Mol Cell Biol. 1991 Sep;11(9):4509–4517. doi: 10.1128/mcb.11.9.4509. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hattori M., Sakaki Y. Dideoxy sequencing method using denatured plasmid templates. Anal Biochem. 1986 Feb 1;152(2):232–238. doi: 10.1016/0003-2697(86)90403-3. [DOI] [PubMed] [Google Scholar]
  8. Jasin M., Berg P. Homologous integration in mammalian cells without target gene selection. Genes Dev. 1988 Nov;2(11):1353–1363. doi: 10.1101/gad.2.11.1353. [DOI] [PubMed] [Google Scholar]
  9. Kim H. S., Smithies O. Recombinant fragment assay for gene targetting based on the polymerase chain reaction. Nucleic Acids Res. 1988 Sep 26;16(18):8887–8903. doi: 10.1093/nar/16.18.8887. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Koller B. H., Marrack P., Kappler J. W., Smithies O. Normal development of mice deficient in beta 2M, MHC class I proteins, and CD8+ T cells. Science. 1990 Jun 8;248(4960):1227–1230. doi: 10.1126/science.2112266. [DOI] [PubMed] [Google Scholar]
  11. Koller B. H., Smithies O. Inactivating the beta 2-microglobulin locus in mouse embryonic stem cells by homologous recombination. Proc Natl Acad Sci U S A. 1989 Nov;86(22):8932–8935. doi: 10.1073/pnas.86.22.8932. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Mansour S. L., Thomas K. R., Capecchi M. R. Disruption of the proto-oncogene int-2 in mouse embryo-derived stem cells: a general strategy for targeting mutations to non-selectable genes. Nature. 1988 Nov 24;336(6197):348–352. doi: 10.1038/336348a0. [DOI] [PubMed] [Google Scholar]
  13. McMahon A. P., Bradley A. The Wnt-1 (int-1) proto-oncogene is required for development of a large region of the mouse brain. Cell. 1990 Sep 21;62(6):1073–1085. doi: 10.1016/0092-8674(90)90385-r. [DOI] [PubMed] [Google Scholar]
  14. Ramírez-Solis R., Rivera-Pérez J., Wallace J. D., Wims M., Zheng H., Bradley A. Genomic DNA microextraction: a method to screen numerous samples. Anal Biochem. 1992 Mar;201(2):331–335. doi: 10.1016/0003-2697(92)90347-a. [DOI] [PubMed] [Google Scholar]
  15. Saunders A. M., Seldin M. F. The syntenic relationship of proximal mouse chromosome 7 and the myotonic dystrophy gene region on human chromosome 19q. Genomics. 1990 Feb;6(2):324–332. doi: 10.1016/0888-7543(90)90573-d. [DOI] [PubMed] [Google Scholar]
  16. Shen P., Huang H. V. Homologous recombination in Escherichia coli: dependence on substrate length and homology. Genetics. 1986 Mar;112(3):441–457. doi: 10.1093/genetics/112.3.441. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Shulman M. J., Nissen L., Collins C. Homologous recombination in hybridoma cells: dependence on time and fragment length. Mol Cell Biol. 1990 Sep;10(9):4466–4472. doi: 10.1128/mcb.10.9.4466. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Soriano P., Montgomery C., Geske R., Bradley A. Targeted disruption of the c-src proto-oncogene leads to osteopetrosis in mice. Cell. 1991 Feb 22;64(4):693–702. doi: 10.1016/0092-8674(91)90499-o. [DOI] [PubMed] [Google Scholar]
  19. Stanton B. R., Reid S. W., Parada L. F. Germ line transmission of an inactive N-myc allele generated by homologous recombination in mouse embryonic stem cells. Mol Cell Biol. 1990 Dec;10(12):6755–6758. doi: 10.1128/mcb.10.12.6755. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Thomas K. R., Capecchi M. R. Site-directed mutagenesis by gene targeting in mouse embryo-derived stem cells. Cell. 1987 Nov 6;51(3):503–512. doi: 10.1016/0092-8674(87)90646-5. [DOI] [PubMed] [Google Scholar]
  21. Thomas K. R., Capecchi M. R. Targeted disruption of the murine int-1 proto-oncogene resulting in severe abnormalities in midbrain and cerebellar development. Nature. 1990 Aug 30;346(6287):847–850. doi: 10.1038/346847a0. [DOI] [PubMed] [Google Scholar]
  22. Thompson S., Clarke A. R., Pow A. M., Hooper M. L., Melton D. W. Germ line transmission and expression of a corrected HPRT gene produced by gene targeting in embryonic stem cells. Cell. 1989 Jan 27;56(2):313–321. doi: 10.1016/0092-8674(89)90905-7. [DOI] [PubMed] [Google Scholar]
  23. Waldman A. S., Liskay R. M. Dependence of intrachromosomal recombination in mammalian cells on uninterrupted homology. Mol Cell Biol. 1988 Dec;8(12):5350–5357. doi: 10.1128/mcb.8.12.5350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Waldman A. S., Liskay R. M. Differential effects of base-pair mismatch on intrachromosomal versus extrachromosomal recombination in mouse cells. Proc Natl Acad Sci U S A. 1987 Aug;84(15):5340–5344. doi: 10.1073/pnas.84.15.5340. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Wallimann T., Wyss M., Brdiczka D., Nicolay K., Eppenberger H. M. Intracellular compartmentation, structure and function of creatine kinase isoenzymes in tissues with high and fluctuating energy demands: the 'phosphocreatine circuit' for cellular energy homeostasis. Biochem J. 1992 Jan 1;281(Pt 1):21–40. doi: 10.1042/bj2810021. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Watt V. M., Ingles C. J., Urdea M. S., Rutter W. J. Homology requirements for recombination in Escherichia coli. Proc Natl Acad Sci U S A. 1985 Jul;82(14):4768–4772. doi: 10.1073/pnas.82.14.4768. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Zijlstra M., Li E., Sajjadi F., Subramani S., Jaenisch R. Germ-line transmission of a disrupted beta 2-microglobulin gene produced by homologous recombination in embryonic stem cells. Nature. 1989 Nov 23;342(6248):435–438. doi: 10.1038/342435a0. [DOI] [PubMed] [Google Scholar]
  28. te Riele H., Maandag E. R., Clarke A., Hooper M., Berns A. Consecutive inactivation of both alleles of the pim-1 proto-oncogene by homologous recombination in embryonic stem cells. Nature. 1990 Dec 13;348(6302):649–651. doi: 10.1038/348649a0. [DOI] [PubMed] [Google Scholar]
  29. van Deursen J., Lovell-Badge R., Oerlemans F., Schepens J., Wieringa B. Modulation of gene activity by consecutive gene targeting of one creatine kinase M allele in mouse embryonic stem cells. Nucleic Acids Res. 1991 May 25;19(10):2637–2643. doi: 10.1093/nar/19.10.2637. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. van Deursen J., Schepens J., Peters W., Meijer D., Grosveld G., Hendriks W., Wieringa B. Genetic variability of the murine creatine kinase B gene locus and related pseudogenes in different inbred strains of mice. Genomics. 1992 Feb;12(2):340–349. doi: 10.1016/0888-7543(92)90383-4. [DOI] [PubMed] [Google Scholar]

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