Journal
SENSORS AND ACTUATORS B-CHEMICAL
Volume 380, Issue -, Pages -Publisher
ELSEVIER SCIENCE SA
DOI: 10.1016/j.snb.2023.133400
Keywords
Self activation; Symbiotic multi -DNA machine; Terminal deoxynucleotidyl transferase; Isothermal exponential amplification; Biomarker analysis
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In this study, we developed a versatile DNA machine (SM-DNA machine) for ultrasensitive analysis of terminal deoxynucleotidyl transferase (TdT) activity. The machine is composed of a single poly-thymine (Poly-T) tailed functional hairpin probe (T-HP), dependent on the template-free elongation property of TdT. By introducing external TdT, the T-HP is elongated with a long poly-adenine chain, triggering intermolecular interactions between different T-HPs and switching the SM-DNA machine to an ON state. This design enables multiple machine-like movements and exponentially amplifies the fluorescence signal, achieving a limit of detection as low as 0.01 U/mL. The TdT-based strand elongation ensures excellent specificity of the machine to discriminate TdT from other enzymes. This DNA machine shows great potential for TdT biomarker biosensing and can be expanded as a universal platform for various sensing applications and disease diagnosis.
DNA machines with great design flexibility and precise programmability have exhibited promising potential for biomedical analysis. In this contribution, we engineered a symbiotic multi-DNA machine (SM-DNA machine) for ultrasensitively analysis of terminal deoxynucleotidyl transferase (TdT) activity. The machine, made from only one poly-thymine (Poly-T) tailed functional hairpin probe (T-HP) to simplify the machine composition, is dependent on the template-free elongation property of TdT. This design ensures that in the off state there are no physical links between intermolecular T-HPs. Introduction of external TdT would elongate the T-HP with a long poly-adenine chain to trigger the intermolecular interactions between different T-HPs, which tuned the SM-DNA machine to an ON state responsible for the creation of multiple cyclic signal amplification processes of strand replication, cleavage, and displacement. Such multiple machine-like movements result in an exponentially amplified fluorescence signal to ensure the system has a limit of detection as low as 0.01 U/mL. The TdT based strand elongation ensures the machine has an excellent specificity to discriminate TdT from other enzymes. We expect this DNA machine with an excellent assay performance for TdT biomarker biosensing could be readily expanded as a universal platform for various sensing applications and disease diagnosis.
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