Utility of C57BL/6J × 129/SvJae embryonic stem cells for generating chromosomal deletions: tolerance to γ radiation and microsatellite polymorphism

Abstract

We have previously reported a method for making nested deletion complexes in mice by irradiation of ES cells. The key to this technology is that F1 hybrid ES cells (called v17.2) of the genotype (BALB/cTa x 129/SvJae) retain germline colonizing ability after exposure to levels of ionizing radiation that induce chromosomal deletions. In an effort to identify other genotypes of ES cells that are suitable for this technology, the radiation sensitivity of the cell line v6.4, which is of the genotype (C57BL/6J x 129/SvJae), was investigated. After treatment with a range of radiation exposures, the developmental potential of these cells was assayed by injecting them into blastocysts to generate chimeric mice. These experiments showed that while cell lethality increased as the level of radiation increased, the surviving ES cells retained full totipotency at all exposure levels, up to 400 Rads. Because polymorphism between parental microsatellite alleles in the F1 hybrid ES cells is important for ascertaining the sizes of induced deletions, the 129/SvJ and 129/SvJae allele sizes of 48 microsatellite loci on chromosome (Chr) 17 were determined. This revealed a higher level of polymorphism between 129 and C57BL/6J on Chr 17. The radiation tolerance, high polymorphism between parental strains, and presence of the widely used C57BL/6J strain component make v6.4 ES cells an attractive cell line for generating radiation-induced chromosomal deletions.

Genetic Background and the Dilemma of Translating Mouse Studies to Humans

“…Instead, we should expand the use of hybrid ES cell lines. Hybrid ES cell lines derived, for example, from F1 intercrosses of 129 3 C57BL/6 or 129 3 BALB/c mice, retain the 129 characteristics of easy derivation and ability to be manipulated in culture while remaining competent to repopulate the mouse germline (You et al, 1998). They allow study of the effects of genetic heterogeneity observed in the human population.…”

Positional cloning and characterization of mouse mei8, a disrupted allele of the meiotic cohesin Rec8

“…Mutant families carrying infertility mutations were established using chimeric male mice derived from chemically treated v6.4 ES cells of the F1 hybrid genotype C57BL/6J ϫ 129S4/SvJae (You et al, 1998). mei8 segregated as a simple recessive Mendelian trait, yielding ϳ25% affected offspring from intercrosses of heterozygotes.…”

Adipocyte-specific Disruption of Fat-specific Protein 27 Causes Hepatosteatosis and Insulin Resistance in High-fat Diet-fed Mice

“…A neomycin (Neo) cassette flanked by two FRT sites and one loxP site was inserted into the third intron of the Fsp27 gene using the PL451 vector that contains this cassette. The final targeting vector was purified and electroporated into C57BL/6J x 129/ SvJae hybrid ES cells (14). After screening with G418, the correctly targeted ES clones were confirmed by Southern blotting (Fig.…”