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[Detection of Prader-Willi syndrome by methylation-specific PCR]
1Human Reproductive Engineering Laboratory, Hunan Medical University, Changsha, Hunan, 410078 P.R. China. threlhmu@public.cs.hn.cn
Objective:
Using a faster and more efficient method to diagnose Prader-Willi syndrome(PWS).
Methods:
Differential DNA methylation at several sites in the PWS critical region where the maternal homologue is unmethylated and transcriptionally active. Methylation-specific PCR(MSPCR) is based on sodium bisulfite treatment of DNA, which converts unmethylated but not methylated cytosine residues to uracil, and PCR primers specific for the maternal and the paternal allele. Methylation analysis by Southern blot is made to validate the MSPCR result.
Results:
Bisulfite-modified DNA from PWS patients amplified only with methylated allele-specific primer pair showed only maternal 174bp PCR product. MSPCR and PW71B methylation studies showed an abnormal pattern consistent with the clinical diagnosis of PWS. Untreated DNA does not produce a PCR product.
Conclusion:
MSPCR can be used to detect all presently testable causes of PWS (deletion, uniparental disomy, and imprinting mutation) in an efficient first step for stepwise diagnostic testing, compared with either Southern blot analysis which is more timeconsuming, or fluorescence in situ hybridization(FISH) which can not detect uniparental disomy or imprinting mutation.