A facile and pragmatic electrochemical biosensing strategy for ultrasensitive detection of DNA in real sample based on defective T junction induced transcription amplification

A novel and pragmatic electrochemical sensing strategy was developed for ultrasensitive and specific detection of nucleic acids by combining with defective T junction induced transcription amplification (DTITA). The homogeneous recognition and specific binding of target DNA with a pair of designed probes formed a defective T junction, further triggered primer extension reaction and in vitro transcription amplification to produce numerous single-stranded RNA. These RNA products of DTITA could hybridized with the biotinylated detection probes and immobilized capture probes for enzyme-amplified electrochemical detection on the surface of the biosensor. The proposed isothermal DTITA strategy displayed remarkable signal amplification performance and reproducibility. The electrochemical DNA biosensor showed very high sensitivity for target DNA with a low detection limit of 0.4 fM (240 molecules of the synthetic DNA), and can directly detect target pathogenic gene of Group B Streptococci (GBS) from as low as 400 copies of genomic DNA. Moreover, the established biosensor was successfully verified for directly identifying GBS in clinical samples. This proposed strategy presented a simple and pragmatic platform toward ultrasensitive and handy nucleic acids detection, and would become a potential tool for general application in point-of-care setting.