Human Molecular Genetics, 2002, Vol. 11, No. 7 791-798
© 2002 Oxford University Press
Age and insertion site dependence of repeat number instability of a human DM1 transgene in individual mouse sperm
Department of Experimental Radiation Oncology and 1Department of Molecular Genetics, The University of Texas M. D. Anderson Cancer Center, 2Department of Environmental Sciences, The University of Texas School of Public Health, Houston, TX 77030, USA and 3Institute of Biomedical and Life Sciences, University of Glasgow, Anderson College, Glasgow G11 6NU, UK
Precise measurement of germline repeat number mutations is important for understanding the molecular etiology of expanded trinucleotide repeat diseases. We used single genome-equivalent PCR of sperm DNA to measure the mutation frequencies in two lines of Dmt transgenic mice containing an expanded CTG·CAG tract on an identical genetic background. Single genome-equivalent PCR indicated that apparent mutational spectra derived in other investigations from PCR of bulk sperm DNA were largely the consequence of PCR stutter and not mutations. Here we show that sperm from 8-week-old Dmt-D mice had a significantly higher mutation frequency (change of >1 repeat) (14.2%) than those of Dmt-E mice of the same age (5.5%), in agreement with pedigree analysis. Furthermore, the mutation frequency in sperm of Dmt-D mice increased significantly with age (28.0% at 17 weeks). The age dependence of the degree of expansion implies that mutations accumulate with time in spermatogenic stem cells. Similar rates of expansion per spermatogenic cycle in man would yield the large expansions observed in human diseases such as myotonic dystrophy type 1. Pedigree data showed a significant age-dependent bias toward repeat contraction in female transmissions and a trend towards expansion with age in male transmissions. Thus, direct single genome-equivalent PCR of the sperm DNA of an individual male appears to predict the distribution of mutant allele sizes that might be inherited by its offspring. In further contrast to a recent report, the sex of the offspring had no detectable effect on the direction of the mutational length change.
+ To whom correspondence should be addressed at present address: CIIT Centers for Health Research, 6 Davis Drive, Research Triangle Park, NC 27709, USA. Tel: +1 919 558 1264; Fax: +1 919 558 1300; Email: yzhang@ciit.org
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