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A novel fluorescence method for activity assay and drug screening of T4 PNK coupling rGO with ligase reaction

Abstract

T4 polynucleotide kinase (PNK) is a primary member of 5’-kinase family that can transfer the γ-phosphate residue of ATP to the 5’-hydroxyl group of oligonucleotides. In this article, using the differential quenching ability of reduced graphene oxide (rGO) to the fluorophore labeled DNA probe, we proposed a novel method for detecting T4 PNK activity assisted with ligase reaction. Under the optimized conditions, the detection limit of T4 PNK is estimated to be 0.0002 U/μL in the linear region of 0.001 U/μL~0.1 U/μL. Additionally, the developed method was used to screen regulators of T4 PNK from natural compounds. The compound f isolated from the root of Kadsura coccinea (Lem.) A.C. Smith was found to stimulate T4 PNK activity in a concentration-dependent manner in vitro. Finally, the method was used to monitor the relation of T4 PNK activity with pelvic inflammatory disease (PID). The results demonstrated that the development of this disease could inhibit T4 PNK activity in some extent. In summary, these above data indicate that the method not only provides a universal platform for monitoring T4 PNK activity but also shows great potential to be used in drug screening and clinic diagnosis.

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Publication details

The article was received on 06 Nov 2018, accepted on 04 Dec 2018 and first published on 05 Dec 2018


Article type: Paper
DOI: 10.1039/C8AN02147C
Citation: Analyst, 2018, Accepted Manuscript
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    A novel fluorescence method for activity assay and drug screening of T4 PNK coupling rGO with ligase reaction

    H. Zhou, C. Tong, W. Zou, Y. Yang, Y. Liu, B. Li, Y. Qin, W. Dang, B. Liu and W. Wang, Analyst, 2018, Accepted Manuscript , DOI: 10.1039/C8AN02147C

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