Targeted Selection of Aptamer Complementary Elements toward Rapid Development of Aptamer Transducers

Biosensing using aptamers has been a recent interestfor theirversatility in detecting many different analytes across a wide rangeof applications, including medical and environmental applications.In our last work, we introduced a customizable aptamer transducer(AT) that could successfully feed-forward many different output domainsto target a variety of reporters and amplification reaction networks.In this paper, we explore the kinetic behavior and performance ofnovel ATs by modifying the aptamer complementary element (ACE) chosenbased on a technique for exploring the ligand-binding landscape ofduplexed aptamers. Using published data, we selected and constructedseveral modified ATs that contain ACEs with varying length, positionof the start sites, and position of single mismatches, whose kineticresponses were tracked with a simple fluorescence reporter. A kineticmodel for ATs was derived and used to extract the strand-displacementreaction constant k (1) and the effectiveaptamer dissociation constant K (d,eff),allowing us to calculate a relative performance metric, k (1)/K (d,eff). Comparing our resultswith the predictions based on the literature data, we provide usefulinsight into the dynamics of the adenosine AT's duplexed aptamerdomain and suggest a high-throughput approach for future ATs to bedeveloped with improved sensitivity. The performance of our ATs showeda moderate correlation to those predicted by the ACE scan method.Here, we find that predicted performance based on our ACE selectionmethod was moderately correlated to our AT's performance.

Automated summary

Biosensing using aptamers has gained attention due to their versatility in detecting different analytes in various fields. This study explored the kinetic behavior and performance of modified aptamer transducers (ATs) by altering the aptamer complementary elements (ACEs). The results showed that the ATs' performance correlated moderately with the predictions based on the ACE selection method, providing valuable insights into the dynamics of the duplexed aptamer domain and suggesting a high-throughput approach for developing ATs with improved sensitivity.