HELP assay

The HELP Assay is "Hpa"II-tiny fragment Enrichment by Ligation-mediated PCR.

To study cytosine methylation throughout the genome, techniques usually involve the use of restriction enzymes or proteins with affinity for 5-methylcytosine.

The HELP assay [Genome Res. 2006 Aug;16(8):1046-55. [http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=16809668&dopt=Citation Pubmed ID: 16809668] ] compares representations generated by "Hpa"II and by "Msp"I digestion of the genome followed by ligation-mediated PCR. As "Hpa"II only digests 5'-CCGG-3' sites when the cytosine in the central CG dinucleotide is unmethylated, the "Hpa"II representation is enriched for the hypomethylated fraction of the genome. The "Msp"I representation is a control for copy number changes and PCR amplification difficulties.

It was recently shown [ Nat Genet. 2006 Dec;38(12):1378-85. [http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=17072317&dopt=Citation Pubmed ID: 17072317] ] that cytosine methylation patterns tend to be concordant over short (~1 kb) regions. The patterns represented by the "Hpa"II sites therefore tend to be representative of other CG dinucleotides locally.

The protocol for the HELP assay is available from [http://greallylab.aecom.yu.edu/HELP/help.html] .

The analysis of HELP data involves quality analysis and normalization. An analytical pipeline written in the R programming language was recently published [Bioinformatics. 2008 May 1;24(9):1161-7. [http://www.ncbi.nlm.nih.gov/pubmed/18353789?dopt=Citation Pubmed ID: 18353789] ] to allow HELP data processing.

References