MULTIPLEX POLYMERASE CHAIN REACTION-BASED DELETION ANALYSIS OF SPONTANEOUS, GAMMA-RAY-INDUCED AND ALPHA-INDUCED HPRT MUTANTS OF CHO-K1 CELLS

Independent Chinese hamster ovary (CHO)-K1 cell mutants at the hypoxan thine-guanine phosphoribosyltransferase (hprt) locus were isolated fro m untreated, Co-60 gamma ray-and Bi-212 alpha-exposed cells and the ge netic changes underlying the mutation determined by multiplex polymera se chain reaction (PCR)-based exon deletion analysis. In the 71 sponta neous mutants analyzed, 77.5% of the clones showed no change in exon n umber or size, 15.5% showed a loss of a single exon, 4.2% showed a los s of 2-8 exons, and 2.8% showed loss of all nine hprt exons (total gen e deletion). Exposure to 6 Gy of gamma rays, which reduced survival le vels to 10%, produced a significantly different deletion spectrum that was shifted toward deletions with 45% of the 20 mutants analyzed show ing a loss of a single exon and 30% showing a loss of all nine exons. Exposure to 2 Gy alpha radiation from Bi-212, a Rn-220 daughter, a dos e which also reduced survival levels to about 10%, resulted in a delet ion spectrum similar to the gamma-ray spectrum in that more than 75% o f the 49 mutants analyzed were deletions. The alpha spectrum, however, was significantly different from both the spontaneous and gamma spect ra with 55.1% of the alpha mutants showing a loss of all nine exons, 1 0.2% showing loss of a single exon, and 14.3% showing loss of 2-8 exon s. Thus, alpha-radiation appears to produce larger intragenic deletion s than gamma radiation. The results suggest that intragenic deletion s ize should be considered when low- and high linear energy transfer (LE T) mutation spectra are compared.