Seo Seung Beom, Lee Jina, Kim Eunjung, Lim Jaewoo, Jang Soojin, Son Seong Uk, Jeong Yeonwoo, Kang Taejeoon, Jung Juyeon, Lee Kyoung G, Lee Sung-Woon, Kim Kyujung, Lim Eun-Kyung
Bionanotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, 34141, Republic of Korea; Department of Cogno-Mechatronics Engineering, Pusan National University, Pusan, 46241, Republic of Korea.
Bionanotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, 34141, Republic of Korea; Department of Nanobiotechnology, KRIBB School of Biotechnology, UST, Daejeon, 34113, Republic of Korea.
Talanta. 2024 Aug 1;275:126073. doi: 10.1016/j.talanta.2024.126073. Epub 2024 Apr 13.
Methicillin-resistant Staphylococcus aureus (MRSA) has a high incidence in infectious hospitals and communities, highlighting the need for early on-site detection due to its resistance to methicillin antibiotics. The present study introduces a highly sensitive detection system for mecA, a crucial methicillin marker, utilizing an RCA-based isothermal exponential amplification reaction. The G-quadruplex-based isothermal exponential amplification reaction (GQ-EXPAR) method designs probes to establish G-quadruplex secondary structures incorporating thioflavin T for fluorescence. The system, unlike conventional genetic detection methods, works with portable isothermal PCR devices (isoQuark), facilitating on-site detection. A detection limit of 0.1 fmol was demonstrated using synthetic DNA, and effective detection was proven using thermal lysis. The study also validated the detection of targets swabbed from surfaces within bacterial 3D nanostructures using the GQ-EXPAR method. After applying complementary sequences to the padlock probe for the target, the GQ-EXPAR method can be used on various targets. The developed method could facilitate rapid and accurate diagnostics within MRSA strains.
耐甲氧西林金黄色葡萄球菌（MRSA）在传染病医院和社区中的发病率很高，由于其对甲氧西林类抗生素具有抗性，因此凸显了早期现场检测的必要性。本研究介绍了一种用于检测关键甲氧西林标志物mecA的高灵敏度检测系统，该系统利用基于滚环扩增（RCA）的等温指数扩增反应。基于G-四链体的等温指数扩增反应（GQ-EXPAR）方法设计探针，以建立包含硫黄素T用于荧光的G-四链体二级结构。与传统的基因检测方法不同，该系统可与便携式等温PCR设备（isoQuark）配合使用，便于现场检测。使用合成DNA证明检测限为0.1 fmol，并通过热裂解证明了有效检测。该研究还验证了使用GQ-EXPAR方法检测从细菌三维纳米结构表面擦拭的靶标。在将互补序列应用于靶标的锁式探针后，GQ-EXPAR方法可用于各种靶标。所开发的方法有助于对MRSA菌株进行快速准确的诊断。
