Detection of Methylated Cytosine on Single Strand DNA Based on Immune-Functionalized and Glutaraldehyde Modified Liquid Exfoliated Graphene Field Effect Transistor

摘要

Detection of the 5-methylcytosine (5mC) is one of the requirements of modern epigenetics research. The mainchallenges in this issue are how to distinguish the 5mC site from the unmethylated ones in the chain of nucleotides, anddetermine its amount. To streamline the tedious operations in traditional bisulfite conversion (BC) and polymerase chainreaction (PCR) based detection methods, lots of graphene derivatives and electrochemistry (EC) sensors have beenexploited. In this work, we would like to propose an electronic method for DNA methylation detection by using fivetesting single strand DNA (ssDNA) chains as proof-of-concepts, based on the glutaraldehyde modified liquid exfoliatedgraphene field effect transistor (LEG-FET). First of all, for the sake of identifying the 5mC site, the immunorecognitionstrategy is utilized and incorporated with LEG-FET working principle. That is, the methylation sites on testing ssDNAchains are first recognized by the fixed 5mC’s antibody (5mCab) molecules on the channel of LEG-FET, then they aretransduced to the varied current between the electrodes of drain and source (I_(DS)). It is found, the changing ratios of I_(DS)(△I_(DS)/I_(DS0)) are in negative relation with the amount of 5mC sites (N_(mC)) at each of the ssDNA concentrations (C_(ssDNA)).When CssDNA is varied from 1 to 10~6 pM, the slopes of the responding curves -△I_(DS)/I_(DS0) vs. N_(mC) are increased from 0.54to 3.70 %/N_(mC); meanwhile, at each of constant N_(mc), the slopes of -△I_(DS)/I_(DS0) vs. C_(ssDNA) are also examined to proof therepeatability in DNA methylation detection.