The investigation of variation and de novo mutation in the human genome

Hypervariable minisatellites form a subset of tandem repeat arrays which show high rates of germline instability. At the hypervariable minisatellite (MS31a, population studies indicated that there was no polarity of variability within the tandem repeat array. This contrasts with previous analysis of pedigree mutant alleles which demonstrated that mutation is polarised towards the 5' end of the repeat array. Population studies on the flanking DNA also demonstrated high levels of recombination throughout the MS31a locus. It is suggested that the absence of any polarity of variability within the repeat array may be due to these elevated levels of recombination flanking the minisatellite. Further evidence to support this has arisen from studies to isolate and characterise crossover molecules from sperm DNA. This work identified a putative recombination hotspot in the 5' flanking DNA adjacent to the most unstable part of the tandem repeat array. This correlates well with similar studies on the human minisatellite MS32, indicating that there may be a conserved mechanism of mutation at these minisatellites.;The second half of the project investigates instability of A1u elements. Alus are dispersed repeats which have been associated with recombination breakpoints in a significant number of genetic diseases. Two A1u rich regions were examined; the first is located upstream of the human hypervariable minisatellite MS32. The second is a region of the C1 inhibitor gene that has previously been associated with A1u-mediated mutation. Population and evolutionary studies predicted that rearrangement at these loci was very rare. Concentrating on the detection of deletion events in sperm DNA, the most sensitive techniques available at this time were unable to recover any mutant alleles at either locus. The implications are that mutation detection techniques must be improved before the molecular mechanisms of low level mutation can be elucidated.