Germ-line intrachromosomal recombination restores fertility in transgenic MyK-103 male mice.

Wilkie, Thomas M.; Braun, Robert E.; Ehrman, Walter J.; Palmiter, Richard D.; Hammer, Robert E.

doi:10.1101/gad.5.1.38

Cited by 31 publications

(14 citation statements)

References 34 publications

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“…Mitotic (54). Our analysis of the OPP line, in which a prescreening of whole seminiferous tubules was performed, was consistent with jackpot events in germ line precursors.…”

Section: Discussionsupporting

confidence: 76%

“…In those experiments, nearly all of the sperm in a hemizygous male stained positive for human growth hormone. Additional histological and functional evidence for this process has been obtained in studies of the Myk-103 transgenic mice (54) and mice hemizygous for an mPl locus (10). If the lacZ gene product were to behave similarly in our system, we would expect that a single correction event would result in multiple blue cells, depending on the number of cells connected in a syncytium (about 128 [19]) and the distance to which diffusion would detectably occur.…”

Section: Resultsmentioning

confidence: 71%

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High-frequency germ line gene conversion in transgenic mice.

Murti¹,

Bumbulis²,

Schimenti³

1992

Mol. Cell. Biol.

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Gene conversion is the nonreciprocal transfer of genetic information between two related genes or DNA sequences. It can influence the evolution of gene families, having the capacity to generate both diversity and homogeneity. The potential evolutionary significance of this process is directly related to its frequency in the germ line. While measurement of meiotic inter-and intrachromosomal gene conversion frequency is routine in fungal systems, it has hitherto been impractical in mammals. We have designed a system for identifying and quantitating germ line gene conversion in mice by analyzing transgenic male gametes for a contrived recombination event. Spermatids which undergo the designed intrachromosomal gene conversion produce functional 13-galactosidase (encoded by the lacZ gene), which is visualized by histochemical staining. We observed a high incidence of lacZ-positive spermatids (-2%), which were produced by a combination of meiotic and mitotic conversion events. These results demonstrate that gene conversion in mice is an active recombinational process leading to nonparental gametic haplotypes. This high frequency of intrachromosomal gene conversion seems incompatible with the evolutionary divergence of newly duplicated genes. Hence, a process may exist to uncouple gene pairs from frequent conversion-mediated homogenization.

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“…Mitotic (54). Our analysis of the OPP line, in which a prescreening of whole seminiferous tubules was performed, was consistent with jackpot events in germ line precursors.…”

Section: Discussionsupporting

confidence: 76%

Section: Resultsmentioning

confidence: 71%

High-frequency germ line gene conversion in transgenic mice.

Murti¹,

Bumbulis²,

Schimenti³

1992

Mol. Cell. Biol.

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“…In studies of random murine transfection integrants, the majority of stable insertions were H/T, and recombination prior to integration was suggested as an explanation for this nonrandom distribution (15). However, in swine transgenic constructs (20) and several murine transgenic constructs (66,67), complex integrants were found to be present primarily as H/H or T/T insertions. Perhaps the distribution of H/H and T/T insertions in mammalian cells suggests that, as in bacteria (30), inverted repeat structures are unstable and the cellular machinery provides strong negative selection against such insertions when introducing naked DNA.…”

Section: Discussionmentioning

confidence: 99%

Nonhomologous recombination in human cells.

Derbyshire¹,

Epstein²,

Young³

et al. 1994

Mol. Cell. Biol.

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Nonhomologous recombination (NHR) is a major pathway for the repair of chromosomal double-strand breaks in the DNA of somatic cells. In this study, a comparison was made between the nonhomologous end joining of transfected adenovirus DNA fragments in vivo and the ability of purified human proteins to catalyze nonhomologous end joining in vitro. Adenovirus DNA fragments were shown to be efficiently joined in human cells regardless of the structure of the ends. Sequence analysis of these junctions revealed that the two participating ends frequently lost nucleotides from the 3' strands at the site of the joint. To examine the biochemical basis of the end joining, nuclear extracts were prepared from a wide variety of mammalian cell lines and tested for their ability to join test plasmid substrates. Efficient ligation of the linear substrate DNA was observed, the in vitro products being similar to the in vivo products with respect to the loss of 3' nucleotides at the junction. Substantial purification of the end-joining activity was carried out with the human immature T-cell-line HPB-ALL. The protein preparation was found to join all types of linear DNA substrates containing heterologous ends with closely equivalent efficiencies. The in vitro system for end joining does not appear to contain any of the three known DNA ligases, on the basis of a number of criteria, and has been termed the NHR ligase. The enriched activity resides in a high-molecular-weight recombination complex that appears to include and require the human homologous pairing protein HPP-1 as well as the NHR ligase. Characterization of the product molecules of the NHR ligase reaction suggests that they are linear oligomers of the monomer substrate joined nonrandomly head-to-head and/or tail-to-tail. The joined ends of the products were found to be modified by a 3' exonuclease prior to ligation, and no circular DNA molecules were detected. These types of products are similar to those required for the breakage-fusion-bridge cycle, a major NHR pathway for chromosome double-strand break repair.

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“…However, Southern blot analysis with a neo probe indicated that of 60 Agouti offspring none carried the transgene. It is possible that the transgene inactivates a gene that is imprinted or essential in haploid sperm cells; alternatively, the transgene may be unstable in the germ line (Wilkie et al 1991). In the ROSA~-geo25 strain, heterozygous animals were consistently 25-40% smaller than their wild-type littermates.…”

Section: Phenotypes Associated With Promoter Trap Eventsmentioning

confidence: 99%

Promoter traps in embryonic stem cells: a genetic screen to identify and mutate developmental genes in mice.

Friedrich¹,

Soriano²

1991

Genes Dev.

1,278

933

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A general strategy for selecting insertion mutations in mice has been devised. Constructs lacking a promoter and including a [3-galactosidase gene, or a reporter gene encoding a protein with both [3-galactosidase and neomycin phosphotransferase activity, were designed so that activation of the reporter gene depends on its insertion within an active transcription unit. Such insertion events create a mutation in the tagged gene and allow its expression to be followed by [3-galactosidase activity. Introduction of promoter trap constructs into embryonic stem (ES) cells by electroporation or retroviral infection has led to the derivation of transgenic lines that show a variety of 13-galactosidase expression patterns. Intercrossing of heterozygotes from 24 strains that express [3-galactosidase identified 9 strains in which homozygosity leads to an embryonic lethality. Because no overt phenotype was detected in the remaining strains, these results suggest that a substantial proportion of mammalian genes identified by this approach are not essential for development.

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Germ-line intrachromosomal recombination restores fertility in transgenic MyK-103 male mice.

Cited by 31 publications

References 34 publications

High-frequency germ line gene conversion in transgenic mice.

High-frequency germ line gene conversion in transgenic mice.

Nonhomologous recombination in human cells.

Promoter traps in embryonic stem cells: a genetic screen to identify and mutate developmental genes in mice.

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