chemicals and has been focused on seven specific loci (Russell 1951) that generated visible phenotypes when mutated. In the mid-1970s, Dr. Russell discovered that the chemical N-ethyl-N-nitrosourea (ENU) is a supermutagen for mouse spermatogonial stem cells (Russell and others 1979), inducing primarily point mutations (single base pair substitutions) (Russell and Montgomery 1982) and thus a variety of types of mutations including nulls and hypomorphic alleles (Bedell and others 1996; Ji and others 1999; Marker and others 1997). In 1986, one of us (E.M.R) launched a pilot ENU-mutagenesis experiment (Rinchik and Carpenter 1999) focused at the albino (c; now called Tyr [tyrosinase]) locus in mouse chromosome (Chr) 7. This region is covered by an extensive series of radiation-induced deletion mutations resulting from the Russell specific locus tests. The mutagenesis strategy for the Tyr region, adapted from a similar approach used by Drosophila geneticists, was to mate ENU-mutagenized males (BALB/cR1) that are homozygous for the c coat-color marker to wild-type females (Rinchik and Carpenter 1999). The F1 mice bearing the BALB/c Chr 7 carrying newly induced point mutations, some of which will be closely linked to c, were then mated to carriers of a large “selector” deletion at c. Of the progeny from this second mating, 25% should be albino and may also express an additional new mutant phenotype if an ENU mutation is so closely linked to c that it maps within the limits of the large c deletion. A set of simple complementation crosses to smaller c deletions localized new mutant phenotypes to intervals suitable for a positional-cloning approach (Rinchik and Carpenter 1993, 1999; Rinchik and others 1993). A similar experiment for the deletion complex surrounding the pink-eyed dilution (p) locus, also on Chr 7, is currently under way (Johnson and others 1995; Rinchik and others 1995; Rinchik, Carpenter, and Johnson, manuscript in preparation). Our successes in designing the genetics and logistics of large mutagenesis experiments have led to the establishment of our current program of inducing new mutations in the proximal two thirds of Chr 7 (which includes the p and c regions), the central one half of Chr 10, the distal half of Chr 15, and a small segment of the X chromosome.
In all of these experiments, broad-based screening for the detection of new mutant phenotypes plays a new and very prominent role. The current program has benefited from our hands-on experience as we increase both the chromosomal region target sizes and our scope and capacity for examining mice for as many different kinds of abnormalities as possible. Our current experiments take advantage of deletion screens, as described above, and more powerful methods utilizing chromosomal inversions as tools to make newly mutagenized chromosomes homozygous —all without molecular genotyping.
We have learned three important lessons from pilot ENU experiments that have influenced the much broader mutagenesis and phenotype-screening pro