Showing papers by "Jay M. Short published in 1993"

Transgenic systems for in vivo mutation analysis.

Abstract: Transgenic mice carrying shuttle vectors containing the lacI gene as the target permit the in vivo measurement of mutations in multiple tissues and have been used to test the mutagenic effects of several compounds. Tissue-specific and time-dependent responses have been observed, and the spectrum of mutations determined by sequencing allows analysis of the role of expression time in mutagenesis. The results obtained from sequencing analysis have demonstrated spectra paralleling those observed in alternative in vivo assays. In addition to color screening, modifications to this system have permitted direct selection for mutations in the lacI target by a variety of methods. Transgenic rats containing the same lambda/lacI shuttle vector have been developed for inter-species comparison of mutagenesis testing results, which may offer a better understanding of the specific mechanisms involved in mutagenesis at the molecular level in vivo.

Induction of hepatic mutations in lacI transgenic mice

Abstract: Transgenic B6C3F1 and C57BL/6 mice containing a lambda shuttle vector that carries a lacI target and an alpha lacZ reporter gene have been constructed for use in in vivo mutagenesis assays. After chemical treatment of mice carrying the lacI target gene, genomic DNA is isolated and the shuttle vector is recovered by exposing the DNA to lambda phage packaging extracts in vitro. Mutations in the lacI target gene that inactivate the repressor gene allow expression of the alpha lacZ reporter gene, resulting in blue mutant plaques. We have examined the ability of two genotoxic agents, dimethylnitrosamine (DMN) and methylmethane sulfonate (MMS), to induce mutations in these transgenic mice. Both compounds induce a variety of DNA adducts in mouse liver; DMN is a hepatocarcinogen that induces significant hepatic cell proliferation, but MMS is not hepatocarcinogenic and does not induce hepatic cell proliferation. The effects of animal age, differences in strain and dosing regimen, and length of expression time were evaluated. Mice were treated for 5, 14 or 21 days and were sacrificed 1, 8 or 22 days after the final dose to evaluate the effects of increased expression time on mutant frequency in liver. In 3 week old mice, DMN (2 mg/kg/day) produced 10- to 20-fold elevations in mutant frequency that increased with expression time and the number of treatments. In contrast, MMS (20 mg/kg/day) failed to increase the mutant frequency. DMN failed to induce mutations in 6 week old mice at 2 mg/kg/day, but 4 mg/kg/day yielded significant elevations in hepatic mutations. Sequencing results indicate that treatment of mice with DMN produced predominantly C:G-->T:A transitions.(ABSTRACT TRUNCATED AT 250 WORDS)

Mutagenesis testing using transgenic non-human animals carrying test DNA sequences

Abstract: An assay for monitoring and assessing the mutagenic potential of agents which involves creating transgenic non-human animals carrying a test DNA sequence or sequences that can be quickly recovered and examined for mutations following exposure to one or more suspected mutagenic agents.

Nucleic acid construct encoding a nuclear transport peptide operatively linked to an inducible promoter

Abstract: A system for regulating expression of eukaryotic genes in cells is described. The system contains two recombinant DNA molecules, a first molecule that encodes a nucleus-targeted inducible repressor polypeptide, and a second molecule that encodes an operator-regulated reporter polypeptide. Transgenic animals containing the system, and methods for using the system are also described.