title: Detection and quantification of methylation in DNA using solid-state nanopores.
abstract: Epigenetic modifications in eukaryotic genomes occur primarily in the form of 5-methylcytosine (5 mC). These modifications are heavily involved in transcriptional repression, gene regulation, development and the progression of diseases including cancer. We report a new single-molecule assay for the detection of DNA methylation using solid-state nanopores. Methylation is detected by selectively labeling methylation sites with MBD1 (MBD-1x) proteins, the complex inducing a 3 fold increase in ionic blockage current relative to unmethylated DNA. Furthermore, the discrimination of methylated and unmethylated DNA is demonstrated in the presence of only a single bound protein, thereby giving a resolution of a single methylated CpG dinucleotide. The extent of methylation of a target molecule could also be coarsely quantified using this novel approach. This nanopore-based methylation sensitive assay circumvents the need for bisulfite conversion, fluorescent labeling, and PCR and could therefore prove very useful in studying the role of epigenetics in human disease.
authors.affiliation: Department of Bioengineering, University of Illinois at Urbana-Champaign, Urbana, IL, USA.
authors.forename: Jiwook
authors.initials: J
authors.lastname: Shim
authors.forename: Gwendolyn I
authors.initials: GI
authors.lastname: Humphreys
authors.forename: Bala Murali
authors.initials: BM
authors.lastname: Venkatesan
authors.forename: Jan Marie
authors.initials: JM
authors.lastname: Munz
authors.forename: Xueqing
authors.initials: X
authors.lastname: Zou
authors.forename: Chaitanya
authors.initials: C
authors.lastname: Sathe
authors.forename: Klaus
authors.initials: K
authors.lastname: Schulten
authors.forename: Farhad
authors.initials: F
authors.lastname: Kosari
authors.forename: Ann M
authors.initials: AM
authors.lastname: Nardulli
authors.forename: George
authors.initials: G
authors.lastname: Vasmatzis
authors.forename: Rashid
authors.initials: R
authors.lastname: Bashir
mesh_terms: D020121:5' Untranslated Regions; D018899:CpG Islands; D004247:DNA / Q000032:analysis*; D019175:DNA Methylation; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D005456:Fluorescent Dyes / Q000737:chemistry; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D058608:Nanopores; D016133:Polymerase Chain Reaction; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
chemical_list: D020121:5' Untranslated Regions; D004268:DNA-Binding Proteins; C088334:Distal-less homeobox proteins; D005456:Fluorescent Dyes; D018398:Homeodomain Proteins; C066141:MBD1 protein, human; D014157:Transcription Factors; D004247:DNA