Fluorescent conjugated polyelectrolyte as an indicator for convenient detection of DNA methylation.
A convenient, sensitive, and label-free method to determine the DNA methylation status of CpG sites of plasmid and human colon cancer cell has been developed. The system relies on highly selective single base extension reaction and significant optical amplification of cationic conjugated polyelectrolytes (CCP-1). The higher fluorescence resonance energy transfer efficiency between CCP-1 and fluorescein-labeled dGTP (dGTP-Fl) is correlated to the incorporation of dGTP-Fl into the probe DNA by single base extension reaction when the target/probe pair is complementary at the methylation site. As low as 1% methylation status can be determined by this new assay method. Because of the optical amplification property of CCP-1, the method exhibited high sensitivity with a concentration of analyte DNA at the picomolar level. The CCP-1 can form a complex with negatively charged DNA through electrostatic interactions, avoiding labeling the DNA target and probe by covalent linking. The isolation steps employed in other typical assays were avoided to simplify operations and increase repeatability. These features make the system promising for future use for early cancer diagnosis.
Feng
Fude
F
Beijing National Laboratory for Molecular Sciences, Key Laboratory of Organic Solids, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100080, PR China.
Wang
Hongzhong
H
Han
Lingli
L
Wang
Shu
S
D015374:Biosensing Techniques / Q000379:methods*; D045744:Cell Line, Tumor / Q000473:pathology; D003110:Colonic Neoplasms / Q000473:pathology*; D018899:CpG Islands / Q000235:genetics / Q000502:physiology; D019175:DNA Methylation; D003848:Deoxyguanine Nucleotides / Q000737:chemistry; D004573:Electrolytes / Q000737:chemistry*; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods; D005456:Fluorescent Dyes / Q000737:chemistry*; D006801:Humans; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D012680:Sensitivity and Specificity; D013194:Staining and Labeling; D055672:Static Electricity
D003848:Deoxyguanine Nucleotides; D004573:Electrolytes; D005456:Fluorescent Dyes; C029603:deoxyguanosine triphosphate