Mapping large spontaneous deletion endpoints in the human HPRT gene

In an attempt to understand the nature, frequency, and molecular origin of spontaneous mutations in human cells, we have analyzed 85 independent, spontaneous HPRT- human B-lymphoblast clones with particular emphasis on the determination and characterization of large structural alterations (i.e., deletions, insertions, duplications, etc.). Southern blot analysis using a full-length HPRT cDNA probe revealed that 39% (33/85) of these spontaneous mutants contained alterations affecting different regions of the gene. 12% (10/85) were total gene deletions, 25% (21/85) involved alterations with one or both endpoints intragenic to HPRT, and 2% (2/85) showed wild-type banding patterns with an additional hybridizing band. To further address the positional behavior of these alterations, the endpoints of the large deletions were mapped to specific exon/intron regions by hybridization of Southern blots with a series of HPRT exon-specific probes. This analysis revealed a disproportionate number of endpoints within the 3' portion of the gene. These findings are discussed in relation to the positional specificity of large alterations in human cells and the use of such an analysis for assessing the molecular mechanism(s) responsible for their production.