通讯机构：
[Lu, Fangguo] U;[Hu, J; Lu, FG] H;Hunan Univ Chinese Med, Dept Microbiol, Sch Med, Changsha 410208, Hunan, Peoples R China.;Univ Innovat Team Hunan Prov, Key Discipline Pathogen Biol, Changsha 410208, Hunan, Peoples R China.
关键词：
*16S rRNA;*Carboxyfluorescein;*DNA probe;*DNA/RNA hybrids;*Drug-resistant bacteria;*Enzymatic reaction;*Fluorescence resonance energy transfer;*Fluorometry;*Quenching
摘要：
The authors describe a method for the fluorometric determination of methicillin-resistant Staphylococcus aureus (MRSA) by exploiting target-triggered chain reactions and deoxyribonuclease I (DNase I)-aided target recycling. It is making use of a carboxy-fluorescein (FAM)-labeled single-stranded probe containing two sections. One is complementary to the 5' terminus of the target, while the 3' terminus of the other target is adsorbed on the surface of graphene oxide (GO) via pi-stacking interactions without the target (16S rRNA). This adsorption results in quenching of the fluorescence of the label and protects it from being cleaved by DNase I. However, upon addition of the target, DNA/RNA hybrids are repelled by GO. This leads to fluorescence recovery as measured at excitation/emission wavelengths of 480/514 nm due to a chain reaction that is triggered by the target. The signal is strongly amplified by using DNase I-mediated target recycling. The 16S rRNA of MRSA can be detected by this method in the 1 to 30 nM concentration range, and the detection limit is 0.02 nM. The method was applied to analyze bacterial samples, and the detection limit is as low as 30 CFU . mL(-1). The assay is highly sensitive and selective and in our percpetion has a large potential in diagnosis of drug-resistant bacteria. Graphical abstract Schematic of the graphene oxide-based fluorescent bioassay for Methicillin-resistant Staphylococcus aureus detection by using target-triggered chain reaction and deoxyribonuclease I-aided signal amplification.
作者机构：
[陈珂珂; 唐亮; 贺气志; 罗怀青; 徐倩] School of Basic Medical Science, Changsha Medical University, Changsha, 410219, China;[宁毅] The Medicine School of Hunan University of Chinese Medicine, Changsha, 410208, China
通讯机构：
The Medicine School of Hunan University of Chinese Medicine, Changsha, China
摘要：
The authors describe an aptamer based fluorometric assay for the determination of ATP. It is based on deoxyribonuclease I-aided target recycling and signal amplification. The DNA probe consists of two regions (sequences) that represent the capture probe and the signal probe, respectively. In the absence of ATP, the probe is adsorbed by the surface of graphene oxide (GO) via π-stacking interactions. This results in quenching of the fluorescent label (carboxyfluorescein) and protects it from being cleaved by DNase. Upon adding ATP, the probe will be repelled by GO because ATP binds to the aptamer. This triggers an increase in fluorescence as measured at excitation/emission wavelengths of 480/514 nm. The detection limit is as low as 0.2 nM, and the calibration plot is linear in the 10 to 400 nM ATP concentration range. The assay is specific and sensitive, and in our perception has a large potential in terms of detecting other species including pathogenic microorganisms, small molecules, metal ions, and proteins. Graphical abstractSchematic of the fluorescent strategy for adenosine triphosphate assay by using aptamer-based target recognition and deoxyribonuclease I-aided signal amplification.
摘要：
The authors describe a low-cost and sensitive method for the fluorometric determination of Salmonella enteritidis (S. enteritidis). It is based on the use of nanographite (NG) as a quencher of fluorescence, and on cyclic signal amplification by using deoxyribonuclease I (DNase I). The probe containing a capture probe and two short sequences (probe 1 and probe 2) labeled with carboxyfluorescein (FAM) are used as the signal probe that is adsorbed on the surface of NG via p-stacking interactions. Adsorption results in quenching of the fluorescence of FAM. If S. enteridis is introduced, fluorescence is restored due to the displacement of the probe from the surface of the NG due competitive binding. The signal is considerably amplified by applying DNase I-mediated target recycling. The assay has a linear response that covers the 1 to 50 nM concentration range, with a 0.5 nM lower limit of detection (LOD). A milk sample spiked with S. enteritidis was analyzed and the detection limit is as low as 50 colony-forming units (CFU) per mL. Accordingly, this biosensor is highly sensitive and selective but also cost-effective. Conceivably, the detection scheme may be extended to the detection of other biomolecules. Graphical abstractSchematic of the fluorescent strategy for Salmonella enteritidis assay by using nanographite as a quencher and deoxyribonuclease I-aided signal amplification
作者机构：
[陈珂珂; 贺气志; 唐亮] Department of Applied Biology, School of Basic Medical Sciences, Changsha Medical University, Changsha, 410219, China;[宁毅; 陈伶利] Department of Pathogenic Biology, Medicine School, Hunan University of Chinese Medicine, Changsha, 410208, China
通讯机构：
[Chen, L.] D;Department of Pathogenic Biology, Medicine School, Hunan University of Chinese Medicine, Changsha, China
