A novel restriction endonuclease GlaI for rapid and highly sensitive detection of DNA methylation coupled with isothermal exponential amplification reaction

Abstract

An elegant GlaI–EXPAR strategy is proposed which allows accurate detection of site-specific DNA methylations with ultrahigh sensitivity and specificity.

A single quantum dot-based nanosensor with multilayer of multiple acceptors for ultrasensitive detection of human alkyladenine DNA glycosylase

“…We designed an AP probe with an AP site (green + purple color, Scheme 1) at the 23rd base from the 5′ end. The AP probe is paired with the trigger (pink color, Scheme 1) generated by the cleavage of the hairpin probe for the initiation of isothermal strand displacement amplification (SDA)41,42 in the presence of APE1. In addition, we designed a capture probe (yellow color, Scheme 1) which can hybridize with the primer (green color, Scheme 1) for the initiation of DNA polymerase-assisted amplification.…”

Construction of a self-directed replication system for label-free and real-time sensing of repair glycosylases with zero background

“…Recently, a series of enzyme-based exponential amplication approaches have been developed for sensitive quantication of low-abundance targets, including polymerase chain reaction (PCR), 25,26 branched rolling circle amplication (RCA), 27 exponential isothermal amplication reaction (EXPAR), [28][29][30] and ligase chain reaction (LCR). 30,31 PCR is a standard enzymatic amplication technique based on thermal cycle-mediated DNA amplication with the involvement of long assay time and precise thermal cycling.…”

“…30,31 PCR is a standard enzymatic amplication technique based on thermal cycle-mediated DNA amplication with the involvement of long assay time and precise thermal cycling. 25,26 Branched RCA 27 and EXPAR [28][29][30] are isothermal enzymatic amplication techniques with the requirement of multiple tool enzymes (i.e., polymerases and nickases), complex operation steps, and high experimental cost. In addition, PCR, 25,26 branched RCA 27 and EXPAR [28][29][30] are two or one primer-dependent amplication techniques with the disadvantage of cross-contamination from nonspecic ampli-cation.…”

“…25,26 Branched RCA 27 and EXPAR [28][29][30] are isothermal enzymatic amplication techniques with the requirement of multiple tool enzymes (i.e., polymerases and nickases), complex operation steps, and high experimental cost. In addition, PCR, 25,26 branched RCA 27 and EXPAR [28][29][30] are two or one primer-dependent amplication techniques with the disadvantage of cross-contamination from nonspecic ampli-cation. LCR is based on thermostable DNA ligase-mediated repetitive cycles of ligation of adjacent hybridized DNA probes, 30,31 but the analysis of LCR products is always challenged by electrophoresis separation.…”

Cytosine-5 methylation-directed construction of a Au nanoparticle-based nanosensor for simultaneous detection of multiple DNA methyltransferases at the single-molecule level

“…Unlike MSREs that identify and cleave unmethylated DNAs, GlaI is a newly discovered methyl-directed DNA restriction endonuclease with good specicity toward 5-mC and high activity toward various catalytic substrates, and it can recognize and cleave the site-specic methylated cytosines, with unmethylated cytosines remaining intact. 31,32 Generally, MSRE-based DNA MTase assays are based on detection of uncleaved methylated DNAs with the assumption that unmethylated DNAs are completely cleaved. 33 In fact, methylated DNAs actually account for a very small percentage of whole genomic DNA, 34 and consequently even a trivial portion of incomplete cleaved unmethylated DNAs may cause signicant interference.…”