FIG. 1.

Phenotype rescue by BAC transgene (Tg). (A) BAC transgenic founder females (generated in B6 background) were outcrossed to C3H.B6-chaos1/chaos1 males (N10F1) in which the chaos1 allele had been introduced into C3H genome by backcrossing nine generations. This was necessary to identify chromosome 16 that carries the chaos1 allele. Resulting F₁ males carrying the BAC transgene were mated with C3H.B6-chaos1/chaos1 females. Among their offspring, chaos1/chaos1 mice were identified as those homozygous for the C3H alleles of the two polymorphic microsatellite markers D16Mit131 (proximal) and D16Mit106 (distal). chaos1/chaos1 mice (+/−Tg) were phenotyped by the micronucleus assay. (B) Spontaneous micronucleus frequencies were measured in CD71-negative normochromatic erythrocytes (NCE; lower quadrants of the plots). Micronucleated erythrocytes (MN-NCE) are in the population positive to propidium iodide (lower right quadrant). Anti-CD71 antibody was used to separate reticulocytes (younger erythrocytes) containing significant amounts of RNA, which potentially interferes with accurate enumeration of micronuclei in NCE. The transgene carriers show a normal range of micronucleus frequencies as comparable to those seen in wild-type mice, whereas chaos1/chaos1 mice exhibited significantly higher micronucleus frequencies, indicating complete phenotype correction. At least 10,000 erythrocytes were collected.