Figure 3

Four categories of electropherogram profiles generated by high-throughput TGCE mutation screening of DNA and cDNA samples in the CMMB. The 22 ENU-induced mutations identified in this study (see Table 1) by TGCE analysis of heteroduplexed PCR products produced electropherogram patterns that can be divided into four general categories based on the number and shape of the peaks. Shown are representative electropherograms from each category, which include: (A) three peaks (CMMB #1128, 484-bp amplicon, Scnm1); (B) two peaks of approximately equal intensity (CMMB #3789, 484-bp amplicon, Scnm1); (C) two peaks with lower intensity of the heteroduplex peak (CMMB #1954, 336-bp amplicon, Usp29): and (D) a shoulder on the main peak (CMMB #1176, 335-bp amplicon, Antxr1). In each of the four panels the electropherogram on the top is from the indicated CMMB sample with a heterozygous ENU mutation and the electropherogram on the bottom is from an untreated C57BL/6J control sample. All patterns shown were obtained from first-pass TGCE screening of unpurified PCR products using a single 50–60°C gradient. Heteroduplex molecules (arrows) denature faster than the homoduplexes as temperature increases during electrophoresis. The distribution of electropherogram patterns observed for the 22 mutations can be categorized as follows: (A) 2, (B) 4, (C) 12, and (D) 4 mutations. Thus, 82% of the profiles exhibited two or more peaks and the remaining 18% were shoulders.