For the purpose of DNA replication, the HpaII tiny fragment Enrichment by Ligation-mediated PCR Assay (HELP Assay)[1] is one of several techniques used for determining whether DNA has been methylated. The technique can be adapted to examine DNA methylation within and around individual genes, or it can be expanded to examine methylation in an entire genome.
The technique relies upon the properties of two restriction enzymes: HpaII and MspI. The HELP assay[1] compares representations generated by HpaII and by MspI digestion of the genome followed by ligation-mediated PCR. HpaII only digests 5'-CCGG-3' sites when the cytosine in the central CG dinucleotide is unmethylated, the HpaII representation is enriched for the hypomethylated fraction of the genome. The MspI representation is a control for copy number changes and PCR amplification difficulties.
It was recently shown that cytosine methylation patterns tend to be concordant over short (~1 kb) regions.[2] The patterns represented by the HpaII sites therefore tend to be representative of other CG dinucleotides locally.
The analysis of HELP data involves quality analysis and normalization. An analytical pipeline written in the R programming language was recently published to allow HELP data processing.[3]
Referencesedit
^ abKhulan B, Thompson RF, Ye K, et al. (August 2006). "Comparative isoschizomer profiling of cytosine methylation: The HELP assay". Genome Research. 16 (8): 1046–55. doi:10.1101/gr.5273806. PMC1524864. PMID 16809668.
^Eckhardt F, Lewin J, Cortese R, et al. (December 2006). "DNA methylation profiling of human chromosomes 6, 20 and 22". Nature Genetics. 38 (12): 1378–85. doi:10.1038/ng1909. PMC3082778. PMID 17072317.
^Thompson RF, Reimers M, Khulan B, et al. (May 2008). "An analytical pipeline for genomic representations used for cytosine methylation studies". Bioinformatics. 24 (9): 1161–7. doi:10.1093/bioinformatics/btn096. PMC5061929. PMID 18353789.
External linksedit
April 14, 2024
help, assay, purpose, replication, hpaii, tiny, fragment, enrichment, ligation, mediated, assay, help, assay, several, techniques, used, determining, whether, been, methylated, technique, adapted, examine, methylation, within, around, individual, genes, expand. For the purpose of DNA replication the HpaII tiny fragment Enrichment by Ligation mediated PCR Assay HELP Assay 1 is one of several techniques used for determining whether DNA has been methylated The technique can be adapted to examine DNA methylation within and around individual genes or it can be expanded to examine methylation in an entire genome The technique relies upon the properties of two restriction enzymes HpaII and MspI The HELP assay 1 compares representations generated by HpaII and by MspI digestion of the genome followed by ligation mediated PCR HpaII only digests 5 CCGG 3 sites when the cytosine in the central CG dinucleotide is unmethylated the HpaII representation is enriched for the hypomethylated fraction of the genome The MspI representation is a control for copy number changes and PCR amplification difficulties It was recently shown that cytosine methylation patterns tend to be concordant over short 1 kb regions 2 The patterns represented by the HpaII sites therefore tend to be representative of other CG dinucleotides locally The analysis of HELP data involves quality analysis and normalization An analytical pipeline written in the R programming language was recently published to allow HELP data processing 3 References edit a b Khulan B Thompson RF Ye K et al August 2006 Comparative isoschizomer profiling of cytosine methylation The HELP assay Genome Research 16 8 1046 55 doi 10 1101 gr 5273806 PMC 1524864 PMID 16809668 Eckhardt F Lewin J Cortese R et al December 2006 DNA methylation profiling of human chromosomes 6 20 and 22 Nature Genetics 38 12 1378 85 doi 10 1038 ng1909 PMC 3082778 PMID 17072317 Thompson RF Reimers M Khulan B et al May 2008 An analytical pipeline for genomic representations used for cytosine methylation studies Bioinformatics 24 9 1161 7 doi 10 1093 bioinformatics btn096 PMC 5061929 PMID 18353789 External links editThe protocol for the HELP assay Retrieved from https en wikipedia org w index php title HELP assay amp oldid 1123481862, wikipedia, wiki, book, books, library,
