International Mammalian Genome Society

The 15th International Mouse Genome Conference (2001)

Table of Contents * Complex Genetics * Developmental Genetics * Gene Annotation * Gene Discovery * Genome Sequencing * Functional Genomics * Mutagenesis * Presentations * Verne Chapman Memorial Lecture

A GENOTYPE-BASED SCREEN IN MOUSE EMBRYONIC STEM CELLS FOR ENU-INDUCED MUTATIONS IN THE SMAD2 LOCUS

Jay Vivian
Department of Genetics
University of North Carolina
Campus Box 7264
Chapel Hill NC 27599 USA

Co-Authors: Chen Y, Magnuson T
Institutions: Department of Genetics, University of North Carolina

Developing a series of mutations of a gene of interest has largely been limited to invertebrate genetic model systems such as Drosophila and C. elegans. An efficient means of creating an allelic series in the mouse would greatly aid the analysis of complex gene function and in the development of mouse models for disease states. Our laboratory has recently described a technique to mutagenize mouse embryonic stem cells with ENU, while maintaining the capacity for germline contribution of the ES cells. We have extended this mutagenesis strategy to identify mutations in non-selectible genes, using a DHPLC-based mutation detection scheme, focusing on the mouse Smad2 locus. The SMAD proteins are a family of intracellular proteins important in TGF-beta signaling. Smad2 was chosen for this screen due to its diverse functions throughout embryonic and adult development. Several ES cell clones carrying mutations in the coding regions of Smad2 have been identified. Germline analysis of one of the Smad2 mutations shows it is a hypomorphic allele. Mice homozygous for the ENU-induced allele have a variety of embryonic defects, including defects in anterior neurectoderm development. Our work demonstrates that a large number of mutations in nonselectible genes can be readily identified by combining ENU mutagenesis in ES cells with genotype-based mutation detection schemes, and can be extended to any gene of interest.