Published online 19 April 2004
Nucleic Acids Research, 2004, Vol. 32, No. 7 e63
© 2004 Oxford University Press
A novel technique based on a PNA hybridization probe and FRET principle for quantification of mutant genotype in fibrous dysplasia/McCune–Albright syndrome
1 Craniofacial and Skeletal Diseases Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Department of Health and Human Services, Bethesda, MD, USA, 2 Dipartimento di Medicina Sperimentale, Università dell’Aquila, L’Aquila, Italy, 3 Parco Scientifico Biomedico San Raffaele, Rome, Italy, 4 Dipartimento di Medicina Sperimentale e Patologia, Università La Sapienza, Rome, Italy and 5 Center for Biologics Evaluation and Research, Food and Drug Administration, Department of Health and Human Services, Bethesda, MD, USA
*To whom correspondence should be addressed at 30 Convent Drive, Building 30, Room 223, MSC 4320, Craniofacial and Skeletal Diseases Branch, National Institute of Dental and Craniofacial Research, NIH, DHHS, Bethesda, MD 20892-4320, USA. Tel: +1 301 402 2684; Fax: +1 301 402 0824; Email: akaradag{at}dir.nidcr.nih.gov
Received December 15, 2003; Revised March 1, 2004; Accepted March 20, 2004
Somatic mutations are present in various proportions in numerous developmental pathologies. Somatic activating missense mutations of the GNAS gene encoding the Gs
protein have previously been shown to be the cause of fibrous dysplasia of bone (FD)/McCune-Albright syndrome (MAS). Because in MAS patients, tissues as diverse as melanocytes, gonads and bone are affected, it is generally accepted that the GNAS mutation in this disease must have occurred early in development. Interestingly, it has been shown that the development of an active FD lesion may require both normal and mutant cells. Studies of the somatic mosaic states of FD/MAS and many other somatic diseases need an accurate method to determine the ratio of mutant to normal cells in a given tissue. A new method for quantification of the mutant:normal ratio of cells using a PNA hybridization probe-based FRET technique was developed. This novel technique, with a linear sensitivity of 2.5% mutant alleles, was used to detect the percentage mutant cells in a number of tissue and cell culture samples derived from FD/MAS lesions and could easily be adapted for the quantification of mutations in a large spectrum of diseases including cancer.
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