1. A convenient electrogenerated chemiluminescence biosensing method for selective detection of 5-hydroxymethylcytosine in genomic DNA.
- Author
-
Wang, Huan, Liu, Meizhou, Bai, Wanqiao, Sun, Huiping, Li, Yan, and Deng, Haiqiang
- Subjects
- *
CHEMILUMINESCENCE , *BIOSENSORS , *QUANTITATIVE research , *THIOLATES , *DOUBLE-strand DNA breaks - Abstract
Highlights • A convenient ECL biosensing method for selective detection of 5-hmC was developed. • The method is based on thiolated anchor probe DNA to immobilize 5-hmC-dsDNA. • The method can be used for detection of 5-hmC in genomic DNA. Abstract Herein, a novel and convenient electrogenerated chemiluminescence (ECL) biosensing method for the quantitative detection of 5-hydroxymethylcytosine (5-hmC) in genomic DNA was developed by employing thiolated anchor probe DNA to immobilize 5-hydroxymethylcytosine double-stranded DNA (5-hmC-dsDNA) and a ruthenium(II) complex-tagged reporter probe as an ECL probe. Briefly, the 5-hmC-dsDNA was firstly oxidized to 5-formylcytosine dsDNA (5-fC-dsDNA) by KRuO 4 in high yields. Subsequently, the thiolated anchor probe DNA is employed to help immobilize the 5-fC-dsDNA onto the gold electrode surface via the Au-S bond. An ECL probe DNA, labeled with a signal transducer − Ru(bpy) 2 (cbpy)NHS, hybridized with the anchor probe DNA on the electrode, resulting in an intensive ECL emission. The obtained ECL intensity was linearly proportional to the content of 5-hmC in the range from 0.0090% to 0.5761%, with the limit of detection calculated to be 0.0036% (S/N = 3). Finally, we applied the developed 5-hmC detection strategy in various mice tissues. It demonstrates that the content of 5-hmC measured by this method in different mice tissues vary significantly: 5-hmC is highest in brain and lowest in spleen. The success in assaying the mice tissues samples signifies its great promise in a broad range of practical applications for the quantitative detection of 5-hmC in genomic DNA. [ABSTRACT FROM AUTHOR]
- Published
- 2019
- Full Text
- View/download PDF