title: HELP (HpaII tiny fragment enrichment by ligation-mediated PCR) assay for DNA methylation profiling of primary normal and malignant B lymphocytes.
abstract: The role of cytosine methylation in the regulation of gene expression during normal development and malignant transformation is currently under intense investigation. An ever increasing body of evidence demonstrates that carcinogenesis is associated with aberrant DNA methylation of the promoters of tumor suppressor genes (Chin Med J (Engl) 111:1028-1030, 1998; Leukemia 17:2533-2535, 2003), hypomethylation of oncogenes (Toxicol Appl Pharmacol 206:288-298, 2005; Toxicology 50:231-245, 1988), and concurrent loss of methylation in the intergenic areas and gene bodies, which may lead to genomic instability and chromosomal fragility (Cytogenet Cell Genet 89:121-128, 2000). Single locus methylation assays have focused largely on specific known tumor suppressor genes or oncogenes (Chin Med J (Engl) 111:1028-1030, 1998; Cancer Res 57:594-599, 1997; Hum Genet 94:491-496, 1994; Mol Cell Biol 14:4225-4232, 1994; Gastroenterology 116:394-400, 1999). Such approaches, while being useful, have clear limitations. With the advent of genome-wide microarray-based techniques, it has become possible to perform genome-wide exploratory studies to better understand genomic patterning of DNA methylation and also to discover new potential disease-specific epigenetic lesions (J Cell Biochem 88:138-143, 2003; Genome Res 16:1075-1083, 2006). In order to capture this type of information from primary human tissues, we have adopted and optimized the HELP assay (HpaII tiny fragment Enrichment by Ligation-mediated PCR) to compare and contrast the abundance of cytosine methylation of genomic regions that are relatively enriched for CpG dinucleotides. While we have mainly used a custom NimbleGen-Roche high-density oligonucleotide microarray containing 25,626 HpaII amplifiable fragments, many other microarray platforms or high throughput sequencing strategies can be used with HELP.
authors.affiliation: Division of Immunopathology, Department of Pathology, Weill Medical College, Cornell University, New York, NY, USA. ris9004@med.cornell.edu
authors.forename: Rita
authors.initials: R
authors.lastname: Shaknovich
authors.forename: Maria E
authors.initials: ME
authors.lastname: Figueroa
authors.forename: Ari
authors.initials: A
authors.lastname: Melnick
mesh_terms: D000818:Animals; D001402:B-Lymphocytes / Q000166:cytology* / Q000378:metabolism / Q000473:pathology*; D002417:Cattle; D002460:Cell Line; D002471:Cell Transformation, Neoplastic / Q000235:genetics* / Q000378:metabolism / Q000473:pathology*; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D019175:DNA Methylation; D019003:Deoxyribonuclease HpaII / Q000378:metabolism*; D005629:Frozen Sections; D006801:Humans; D009693:Nucleic Acid Hybridization; D016133:Polymerase Chain Reaction / Q000379:methods*
chemical_list: D004247:DNA; D019003:Deoxyribonuclease HpaII