Dnmt1 activity detection fluorescent sensor based on G-quadruplex

• Main Authors: WANG YING, LIU SHANSHAN, BIAN ZHAOYANG, FAN ZIYAN, LIU YANG, YANG FEI, DENG HUIMIN, TANG GANGLING
• Format: Patent
• Language: Chinese; English
• Published: 03.01.2020
• Subjects: BEER; BIOCHEMISTRY; CHEMISTRY; COMPOSITIONS OR TEST PAPERS THEREFOR; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL ORENZYMOLOGICAL PROCESSES; ENZYMOLOGY; INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIRCHEMICAL OR PHYSICAL PROPERTIES; MEASURING; MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEICACIDS OR MICROORGANISMS; METALLURGY; MICROBIOLOGY; MUTATION OR GENETIC ENGINEERING; PHYSICS; PROCESSES OF PREPARING SUCH COMPOSITIONS; SPIRITS; TESTING; VINEGAR; WINE

The invention discloses a Dnmt1 activity detection fluorescent sensor based on a G-quadruplex and belongs to the field of biochemical analysis. Firstly, Hairpin DNA which contains a G-quadruplex sequence and subjected to hemi-methylation modification is used as a substrate, and Dnmt1 catalyzes the substrate to have a methylation reaction at 37 DEG C in the presence of S-adenosyl-L-methionine (SAM); then, restriction endonuclease BssHII sensitive to methylation is added, a shear reaction is conducted at 60 DEG C, and the completely methylated substrate cannot be recognized by the BssHII and thus cannot be sheared while the incompletely methylated substrate can be sheared to release a G-quadruplex fragment; and finally, NMM is added to a system and can produce an intensive florescence signalafter binding with the G-quadruplex, the methylation level of substrate DNA can be judged according to intensity of the fluorescence signal, so that the activity of Dnmt1 in the methylation reactioncan be further reflected.