Confirmation of homozygosity for a single nucleotide substitution mutation in a Cockayne syndrome patient using monoallelic mutation analysis in somatic cell hybrids
✍ Scribed by Lisa D. McDaniel; Randy Legerski; Alan R. Lehmann; Errol C. Friedberg; Roger A. Schultz
- Publisher
- John Wiley and Sons
- Year
- 1997
- Tongue
- English
- Weight
- 175 KB
- Volume
- 10
- Category
- Article
- ISSN
- 1059-7794
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✦ Synopsis
The identification of individuals homozygous for a specific mutation offers advantages for the elucidation of molecular mechanisms of hereditary disease states. Cockayne syndrome is a rare autosomal recessive disorder, the molecular basis of which is complicated by significant genetic and clinical heterogeneity. The genes associated with both genetic complementation groups, CSA and CS-B, have been identified. We have previously identified a number of CSA mutations, including a single base substitution that introduces a stop codon ( 322 Tyr®Stop) mutation in the C-terminal region for at least one allele of the CSA gene in a severely affected patient. We now present data confirming the existence of homozygosity in this patient using a strategy with general applicability. Somatic cell hybrids were established by fusing patient cells with mouse A9 cells. Screening with chromosome 5 specific polymorphic markers facilitated identification of hybrid clones bearing only one of the distinct CSA alleles. Sequencing of a portion of the human CSA gene in a subset of these hybrids permitted monoallelic mutation analysis and confirmed the presence of the 322 Tyr®Stop mutation in both alleles.