Yunlei Zhou,Zhiqing Yang,Xue Li,Yue Wang,Huanshun Yin,Shiyun Ai
DNA hydroxymethylation (5-hydroxymethylcytosine, 5hmC) is a kind of new epigenetic modification, which plays key roles in nuclear reprogramming, regulates the gene activity, and initiates the DNA demethylation in mammals. For further understanding the functions of 5hmC and the correlation with tumour, it is essential to develop sensitive and selective methods for detecting and sequencing 5hmC. Herein, a kind of electrochemical biosensor was fabricated for 5hmC detection based on the glycosylation modification of 5hmC and enzymatic signal amplification. Under the catalytic effect of T4 β-glucosyltransferase, the 5hmC in target DNA was chemically modified with glucose. Then with the bridge connection of 1,4-phenyldiboronic acid, alkaline phosphatase was further captured on the electrode surface to catalyze the hydrolysis of p -nitrophenyl phosphate disodium salt to produce p -nitrophenol. Based on the relationship between the electrochemical oxidation signal of p -nitrophenol and the concentration of target DNA, the 5hmC level can be detected with high sensitivity and selectivity. The developed method also showed excellent reproducibility and stability.
Jia Ding,Lanlan Gao,Lulu Cao,Yunlei Zhou,Hao Wang,Jingrui Duan,Huanshun Yin,Jianchao Sun,Shiyun Ai
DNA hydroxymethylation is an important epigenetic modification, which plays crucial roles in many biological processes, such as gene expression and cell differentiation. Herein, a novel photoelectrochemical immunosensor was built for detecting 5-hydroxymethylcytosine (5hmC), where the perovskite Bi 4 TaO 8 Cl sensitized by 3,4,9,10-perylenetetracarboxylicacid (PTCA) was implemented as photoactive material, anti-5hmC antibody was served as target recognition reagent, ZrO 2 was used for connecting more modifications, and ferrocene was adopted as signal amplification unit. The prepared nanomaterials of Bi 4 TaO 8 Cl, PTCA and ZrO 2 were well characterized. The possible mechanism for the improved PEC response of Bi 4 TaO 8 Cl sensitized by PTCA was investigated carefully, indicated that the matched energy band facilitated the migration of photogenerated electron. In addition, the possible mechanism for signal amplification caused by ferrocene was investigated, showing that ferrocene acted as electron donor to accelerate electron transfer. Under the optimal condition, the biosensor manifested wide detection range from 0.02 to 100 nM with the low detection limit of 5.2 pM (S/N = 3). Moreover, the applicability of the biosensor was demonstrated by investigating the effect of heavy metal Cd 2+ and perfluorooctanoic acid on 5hmC content in the leaves and roots of wheat seedlings, as well as the effect of ALV-J on 5hmC content in chicken serum.
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