Converting pyrophosphate generated during loop mediated isothermal amplification to ATP: Application to electrochemical detection of Nosema bombycis genomic DNA PTP1.

Traditionally, genomic DNA detection is relay on a rigorous DNA amplification process, which always accompanied with complicated gel electrophoresis or expensive fluorescence detection methods. In this work, we have translated genomic DNA detection into adenosine triphosphate (ATP) test based on a split aptamer-based electrochemical sandwich assay. The key characteristic of our method are list as follows: first, nucleic acid amplification of the target gene was performed by the use of a loop mediated isothermal amplification (LAMP) process. The pyrophosphate (PPi), which released as the byproduct during the LAMP reaction, were further converted into ATP in the presence of adenosine 5'-phosphosulfate (APS) and ATP sulfurylase. Thereafter, the converted ATP was detected by constructing an electrochemical sandwich aptasensor. With such design, the conversion from the difficult detecting target (genomic DNA) into a convenient measured object (ATP) has been achieved. This proposed strategy was highly sensitive for Nosema bombycis genomic DNA PTP1 detection with a detection limit as low as 0.47 fg/μL and a linear range from 0.001pg/μL to 50ng/μL. And we supposed that this novel target conversion electroanalytical strategy established a universal approach for quantitative analysis of any other kinds of nucleic acid in assistance of nucleic acid polymerization reaction.