基于聚集诱导发射的无模板 DNA 聚合扩增检测 DNA 甲基转移酶活性。
Detection of DNA methyltransferase activity using template-free DNA polymerization amplification based on aggregation-induced emission.
机构信息
Key Laboratory of Sensor Analysis of Tumor Marker, Ministry of Education, Shangdong Key Laboratory of Biochemical Analysis, Key Laboratory of Analytical Chemistry for Life Science in Universities of Shandong, College of Chemistry and Molecular Engineering, Qingdao University of Science and Technology, Qingdao, 266042, PR China.
Key Laboratory of Sensor Analysis of Tumor Marker, Ministry of Education, Shangdong Key Laboratory of Biochemical Analysis, Key Laboratory of Analytical Chemistry for Life Science in Universities of Shandong, College of Chemistry and Molecular Engineering, Qingdao University of Science and Technology, Qingdao, 266042, PR China..
出版信息
Anal Biochem. 2020 Feb 1;590:113532. doi: 10.1016/j.ab.2019.113532. Epub 2019 Dec 7.
A sensitive and selective fluorescence assay for DNA methyltransferase (MTase) activity detection was designed based on aggregation-induced emission (AIE) and target initiated template-free DNA polymerization. Quaternized tetraphenylethene salt was synthesized as the AIE probe, which binds to single-stranded DNA by electrostatic interaction. A hairpin probe was designed with a specific sequence for DNA MTase. In the presence of DNA MTase, the methylation reaction initiated DNA polymerization with terminal deoxynucleotidyl transferase (TdT), which activated the fluorescence intensity through AIE. The designed DNA sensor displayed a linear response to concentrations of DNA adenine methyltransferase (Dam) MTase from 0.5 U·mL to 100 U mL, with a limit of detection of 0.16 U mL. The assay was also effective for detection of DNA MTase activity in human serum and for showing the inhibitory effect of 5-fluorouracil on Dam MTase.
基于聚集诱导发光（AIE）和靶引发无模板 DNA 聚合，设计了一种用于 DNA 甲基转移酶（MTase）活性检测的灵敏和选择性荧光分析方法。合成了季铵化四苯乙烯盐作为 AIE 探针，通过静电相互作用与单链 DNA 结合。设计了一个具有特定序列的发夹探针用于 DNA MTase。在 DNA MTase 的存在下，甲基化反应通过末端脱氧核苷酸转移酶（TdT）引发 DNA 聚合，通过 AIE 激活荧光强度。所设计的 DNA 传感器对 DNA 腺嘌呤甲基转移酶（Dam）MTase 的浓度表现出线性响应，从 0.5 U·mL 到 100 U·mL，检测限为 0.16 U·mL。该测定法还可有效用于检测人血清中的 DNA MTase 活性，并显示 5-氟尿嘧啶对 Dam MTase 的抑制作用。
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