International Mammalian Genome Society

The 14th International Mouse Genome Conference (2000)

F7. A Noble Strategy for Mutagenesis in the Mouse: Heavy Ion Beams and Fetal Phenotype Screening -----III. Expression Profiling in the Mutant Tissues

Chikako Yoshida-Noro1, Hiroki Nagase2, Atsushi Yoshiki1 and Moriaki Kusakabe1
1Experimental Animal Research Division, and
2Molecular Oncology, RIKEN Tsukuba Institute, Tsukuba, 305-0074, Japan

Mouse mutant archive is a useful resource for the functional genomics in the post-genomic era. We propose here noble systematic approaches for the production, screening, preservation and analysis of mouse mutants as follows;

1. Heavy Ion Beam Mutagenesis
2. Fetal Phenotype Screening
3. Analysis for Expression Profiles in the Mutant Tissues
4. Preservation and Functional Analysis using EG cells

In this paper, we describe the analysis of expression profiles in the mutant tissues.

The heavy ion beam was irradiated to the mouse testis, and the G1 mice were obtained by mating or by in vitro fertilization to wild-type mice. The G1 males were mated to wild-type mice, generating G2 progeny. The G2 males were backcrossed to the G1 females, and the G3 fetuses were screened for the phenotype abnormalities at 10-13 day of gestation. RNA from the small fetal tissue with abnormal morphology was extracted and cDNA subtraction library was made using normal fetal tissue cDNA as a driver. The genes expressed specifically in the wild type (or mutant) tissue were isolated and analyzed by using high-throughput system. These genes can be a useful marker for the mapping to specify the mutation site in the genome and also useful for the analysis for the function of the gene cascade which makes mutant phenotypes. 4-D (spatio-temporal) gene expression profile is also examined by in situ hybridization. By combining these data with genome analysis such as SSLP, RLGS or RDA, we study the mechanisms of which the mutated gene and resulted changes of gene cascade generate the mutant phenotype. We applied these approaches to an E13 fetus with abnormal head morphology and analyzed its gene expression profiles. We got several genes expressed in the wild type head but not in the mutant head. Using these cDNAs as probes, we investigate the causality of genetic defect and developmental abnormality.

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