Home > Nanotechnology Columns > Magda Carvalho PhD, JD > Biosensors: DNA Methylation
There is an urgent need of new technologies in early cancer detection. DNA methylation in the promoter region of tumor suppressor genes warrants investigation for its likely role on tumors. A patent application describes methods that use nano transistor surfaces to detect DNA methylation in biological samples (label free). The method is simple and ultra-sensitive (no false positives).
August 20th, 2009
Biosensors: DNA Methylation
DNA methylation is a chemical modification of the DNA (epigenetic alteration). It involves the addition of a methyl group to the DNA which can be subsequently removed without changing the original nucleotide. Addition of a methyl group to the number 5 carbon of the cytosine pyrimidine ring is present in every vertebrate — in this case it reduces or silences gene expression. In adult somatic tissues, DNA methylation typically occurs in a CpG dinucleotide context (60-90% of all CpGs); while in embryonic stem cells there is a lack of methylation at these CpG dinucleotides.
DNA methylation is part of and essential for the normal development and essential functions. It is associated with a number of key processes including imprinting, X-chromosome inactivation, suppression of repetitive elements, long term memory storage in humans, and suppression of tumors.
"CpG islands" are regions of high unmethylated CpGs which are present in the 5' regulatory regions (promoters) of several housekeeping genes including genes that suppress formation of tumors. However, in several forms of cancer, the "CpG islands" present in the promoter region of tumor suppressor genes acquire abnormal hypermethylation, thus silencing the gene. DNA methylation effects the transcription of genes in two ways. First, the methylation of DNA may itself physically impede the binding of regulatory proteins to the gene and secondly, methylated DNA may be bound by proteins that normally bind to methylated nucleotides known as methyl-CpG-binding domain proteins. Since in cancer cells, the CpG islands in the promoter are most likely to be methylated, the detection of methylation in such regions is one vital assay in early cancer diagnosis.
Patent Application US 20090068649: Methods of DNA Methylation Detection.
The invention provides for methods of generating and detecting specific electronic signals that report the methylation status of targeted DNA molecules in biological samples. The detection sensitivity falls at the atom mole level. The novelty relies on the fact that the invention distinguishes methylated from unmethylated DNA using charge detectors bypassing the fact that both methylated and unmethylated DNA carry the same amount of electrical charge.
The invention describes two methods for detection of methylated DNA molecules in a nano transistor —direct and indirect (RNA signal). In the direct detection, the methylated target DNA from a biological sample is identified with a monoclonal anti-methylcytosine antibody immobilized on a sensing surface of a nano transistor; the methylated DNA is detected by changing electrical properties of the sensing surface thus generating a signal. In the indirect detection, anti-5-methylcytosine antibodies are used to identify the target DNA to be subsequently used as template for production of RNA in in vitro transcription; these RNA molecules are captured on the sensing surface and change the electrical properties of the nano transistor thereby generating detectable electronic signals.
In summary, this invention discloses a novel and advanced technology that focus on the methylation status of the promoter region of a gene. This method avoids all the problems in the current DNA methylation detection processes, does not require PCR amplification and provides for a high sensitivity. This strategy is an advancement in cancer early diagnosis.
Maki WC, Mishra NN, Cameron EG, Filanoski B, Rastogi SK, Maki GK. Nanowire-transistor based ultra-sensitive DNA methylation detection. Biosensors & Bioelectronics. 2008 Jan 18;23(6):780-7. Epub 2007 Aug 30.
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