Molecular diagnosis of Beckwith-Wiedemann syndrome using quantitative methylation-sensitive polymerase chain reaction.
OBJECTIVE
Beckwith-Wiedemann Syndrome is caused by defects in imprinted gene expression at 11p15. Currently, quantitative Southern analysis using DNA methylation-sensitive restriction enzymes is used in molecular diagnosis of this syndrome.


METHODS
We describe a rapid and highly quantitative test for assessing DNA methylation at 11p15 using sodium bisulfite treatment of genomic DNA coupled with quantitative TaqMan methylation-sensitive polymerase chain reaction.


RESULTS
TaqMan MSP can assess DNA methylation at both differentially methylated region (DMR)1 and DMR2 at 11p15. In addition, by using TaqMan MSP we were able to determine the parent of origin of a duplication of 11p15 by quantification of both DMR1 and DMR2 DNA methylation.


CONCLUSIONS
TaqMan MSP method is a robust and rapid method for detecting changes in DNA methylation that compares favorably to the current standard of Southern blot for DNA methylation analysis. Assessment of DMR1 and DMR2 provides the most comprehensive assay for methylation defects in Beckwith Wiedemann Syndrome, accounting for more than 70% of the cases. The advantages of TaqMan MSP are that it requires less DNA and that it is rapid, less labor-intensive, and amenable to high-throughput analysis. Moreover, this approach can be modified to assess DNA methylation changes anywhere in the genome.
Coffee
Bradford
B
Department of Human Genetics, Emory University School of Medicine, Atlanta, Georgia 30322, USA.
Muralidharan
Kasinathan
K
Highsmith
William E
WE
Lapunzina
Pablo
P
Warren
Stephen T
ST
D001506:Beckwith-Wiedemann Syndrome / Q000175:diagnosis*; D002880:Chromosomes, Human, Pair 11 / Q000737:chemistry; D004247:DNA / Q000032:analysis; D019175:DNA Methylation; D005260:Female; D020440:Gene Duplication; D006801:Humans; D008297:Male; D016133:Polymerase Chain Reaction / Q000379:methods*
D004247:DNA