A tri-functional probe mediated exponential amplification strategy for highly sensitive detection of Dnmt1 and UDG activities at single-cell level.

A tri-functional probe mediated exponential amplification strategy for highly sensitive detection of Dnmt1 and UDG activities at single-cell level.
Anal Chim Acta. 2020 Mar 22;1103:164-173
Authors: Fan L, Peng Y, Ning B, Wei H, Gao Z, Bai J, Guo L
Abstract
Multiplex DNA methylation and glycosylation are ubiquitous in the human body to ensure the normal function and stability of the genome. The methyltransferases and glycosylases rely on varied enzymes with different action mechanism, which still remain challenges for multiple detection. Herein, we developed a tri-functional dsDNA probe mediated exponential amplification strategy for sensitive detection of human DNA (cytosine-5) methyltransferase 1 (Dnmt1) and uracil-DNA glycosylase (UDG) activities. The tri-functional dsDNA probe was rationally designed with M-DNA and U-DNA. M-DNA contains the 5'-GCmGCGC-3' site for Dnmt1 recognition. U-DNA possesses one uracil as the substrate of UDG and a primer sequence for initiating the amplification reaction. M-DNA was complementary to partial sequence of U-DNA. In the presence of Dnmt1 and UDG, BssHⅡ and Endo Ⅳ were used to nick the 5'-GCGCGC-3' and AP sites respectively, resulting in the release of single-stranded DNA sequence (primer sequence), respectively. After magnetic separation, the released primer sequence hybridizes with padlock DNA (P-DNA), initiating exponential rolling circle amplification to produce numerous G-quadruplexes fo...

Source: Analytica Chimica Acta - February 23, 2020 Category: Chemistry Authors: Fan L, Peng Y, Ning B, Wei H, Gao Z, Bai J, Guo L Tags: Anal Chim Acta Source Type: research