At least seven human genetic disorders involve mutant trinucleotide repeats (CCG and CTG). These include fragile X syndrome, mutation resulting in a closely related fragile site, myotonic dystrophy, spino-bulbar muscular atrophy, Huntington's disease, spino-cerebellar ataxia type 1, and dentatorubral-pallidoluysian atrophy. The number of triplet repeats increases dramatically from normal individuals to carriers to afflicted individuals. This increase in copy number of the unstable sequence with subsequent generations accounts for the non-Mendelian genetic property termed """"""""anticipation"""""""". This laboratory has investigated non-B DNA structures for almost three decades. Several types of preliminary investigations demonstrate that these and related simple repeating sequences adopt unusual DNA structures. We believe that these non-B structures and these capacities to undergo slippage during replication account for the """"""""mutagenic mutations"""""""" found by human geneticists. We shall perform biochemical, physical, and genetic studies on all ten repeating triplet sequences in recombinant plasmids, DNA polymers, and oligomers. Our conclusions that CTG repeats form non-B structures are derived from 2D gel electrophoresis experiments, chemical and enzymatic probe analyses, EM determinations related to nucleosome positioning, and DNA polymerase pause sites in the CTG triplet repeats. The role of methylation of C on DNA conformational features will be explored. Expansion and deletion of CTG sequences has been found in procaryotic cells; this system may serve as an important model for understanding expansion which is related tot he age of onset and severity of human genetic diseases. Also, the role of triplet repeat sequences in genetic recombination in eucaryotic chromosomes will be investigated.
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