title: Direct ultrasensitive electrochemical biosensing of pathogenic DNA using homogeneous target-initiated transcription amplification.
abstract: Sensitive and specific methodologies for detection of pathogenic gene at the point-of-care are still urgent demands in rapid diagnosis of infectious diseases. This work develops a simple and pragmatic electrochemical biosensing strategy for ultrasensitive and specific detection of pathogenic nucleic acids directly by integrating homogeneous target-initiated transcription amplification (HTITA) with interfacial sensing process in single analysis system. The homogeneous recognition and specific binding of target DNA with the designed hairpin probe triggered circular primer extension reaction to form DNA double-strands which contained T7 RNA polymerase promoter and served as templates for in vitro transcription amplification. The HTITA protocol resulted in numerous single-stranded RNA products which could synchronously hybridized with the detection probes and immobilized capture probes for enzyme-amplified electrochemical detection on the biosensor surface. The proposed electrochemical biosensing strategy showed very high sensitivity and selectivity for target DNA with a dynamic response range from 1 fM to 100 pM. Using salmonella as a model, the established strategy was successfully applied to directly detect invA gene from genomic DNA extract. This proposed strategy presented a simple, pragmatic platform toward ultrasensitive nucleic acids detection and would become a versatile and powerful tool for point-of-care pathogen identification.
authors.affiliation: The center for Clinical Molecular Medical detection, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016, PR China.|Key Laboratory of Laboratory Medical Diagnostics (Ministry of Education of China), Department of Laboratory Medicine, Chongqing Medical University, Chongqing 400016, PR China.
authors.forename: Yurong
authors.initials: Y
authors.lastname: Yan
authors.affiliation: Key Laboratory of Laboratory Medical Diagnostics (Ministry of Education of China), Department of Laboratory Medicine, Chongqing Medical University, Chongqing 400016, PR China.
authors.forename: Shijia
authors.initials: S
authors.lastname: Ding
authors.affiliation: Key Laboratory of Laboratory Medical Diagnostics (Ministry of Education of China), Department of Laboratory Medicine, Chongqing Medical University, Chongqing 400016, PR China.
authors.forename: Dan
authors.initials: D
authors.lastname: Zhao
authors.affiliation: Key Laboratory of Laboratory Medical Diagnostics (Ministry of Education of China), Department of Laboratory Medicine, Chongqing Medical University, Chongqing 400016, PR China.
authors.forename: Rui
authors.initials: R
authors.lastname: Yuan
authors.affiliation: The center for Clinical Molecular Medical detection, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016, PR China.
authors.forename: Yuhong
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: The center for Clinical Molecular Medical detection, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016, PR China.
authors.forename: Wei
authors.initials: W
authors.lastname: Cheng
mesh_terms: D015374:Biosensing Techniques; D004247:DNA; D058266:Dielectric Spectroscopy; D055664:Electrochemical Techniques; D021141:Nucleic Acid Amplification Techniques; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014158:Transcription, Genetic
chemical_list: D004247:DNA