• 文献标题:   Fluorometric determination of the activity of uracil-DNA glycosylase by using graphene oxide and exonuclease I assisted signal amplification
  • 文献类型:   Article
  • 作  者:   CHEN MJ, LI WK, MA CB, WU KF, HE HL, WANG KM
  • 作者关键词:   uracildna glycosylase, dna repair, fluorescence, activity, inhibitor
  • 出版物名称:   MICROCHIMICA ACTA
  • ISSN:   0026-3672 EI 1436-5073
  • 通讯作者地址:   Cent South Univ
  • 被引频次:   14
  • DOI:   10.1007/s00604-019-3247-6
  • 出版年:   2019

▎ 摘  要

The base-excision repair enzyme uracil-DNA glycosylase (UDG) plays a crucial role in the maintenance of genome integrity. The authors describe a fluorometric method for the detection of the activity of UDG. It is making use of (a) a 3'-FAM-labeled hairpin DNA probe with two uracil deoxyribonucleotides in the self-complementary duplex region of its hairpin structure, (b) exonuclease I (Exo I) that catalyzes the release of FAM from the UDG-induced stretched ssDNA probe, and (c) graphene oxide that quenches the greenFAM fluorescence of the intact hairpin DNA probe in the absence of UDG. If Exo I causes the release of FAM from the hairpin DNA probe, the fluorescence peakingat 517nm is turned off in the absence of UDG but turned on in its presence. The resulting assay has a wide linear range (0.008 to 1UmL(-1)) and a detection limit as low as 0.005UmL(-1). It has good specificity for UDG over potentially interfering enzymes and gave satisfactory results when applied to biological samples. Conceivably, the method may be used in a wide range of applications such as in diagnosis, drug screening, and in studying the repair of DNA lesions.