Meeting ID: 281 491 465 746  Passcode: YNdNny
Event Type:
MSE Grad Presentation
Talk Title:
Comparing competitive and Evolutionary Screening Approaches for a SARS-CoV-2 Receptor binding Domain Aptamer
​​​​​​​In-person in MoSE 3201A and via Teams Meeting

Committee Members: 

Dr. Valeria Milam, Advisor, MSE

Dr. Vladimir Tsukruk, MSE

Dr. Meisha Shofner, MSE

Dr. Loren Williams, SoCB

Dr. Philip Santangelo, BME


Comparing competitive and Evolutionary Screening Approaches for a SARS-CoV-2 Receptor binding Domain Aptamer

ABSTRACT: Aptamers are single-stranded oligonucleotide sequences that self-fold into 3D structures capable of binding to a variety of biological and non-biological targets with high affinity and specificity. As oligonucleotide analogs to antibodies, the nucleic acid chemistry of aptamers affords them advantages over antibodies in reproducibility, cost, stability, ease of chemical modification and facile integration into modern assays and diagnostic systems. Aptamers are typically discovered through a directed evolution platform called SELEX (Systematic Evolution of Ligands by EXponential enrichment) whereby a random library of oligonucleotides is subjected to successive cycles of target incubation, separation of bound species, and enzymatic amplification of sequence winners from a given selection cycle. In recent decades, this evolutionary aptamer screening platform has been used to produce aptamers with promise as affinity reagents. However, SELEX is a laborious process that typically takes weeks to perform and, though popular in the aptamer literature, SELEX is still considered a “black box” screening approach since the criteria for selecting winners is unknown and complicated by separation inefficiencies and enzymatic amplification artifacts.

To address these screening and analysis limitations, the Milam group has developed a new competition-based platform called CompELS (Competition Enhanced Ligand Selection) that can be performed in a single day while circumventing selection biases stemming from library enrichment during enzymatic amplification. I have recently prepared groups of random DNA libraries embedded with unique sequence-specific group identifiers or “barcodes”. These barcoded libraries will provide us insight into the selection process by quantifying the emergence of competitive binders round by round. These barcoded libraries were used during CompELS screening against the receptor binding domain (RBD) of the SARS-CoV-2 spike protein (BA.2/Omicron variant). Optimization efforts are underway to sequence the results using high throughput next generation sequencing. Once a pool of winners (i.e., first generation aptamer candidates that bind to RBD) and losers (i.e., sequences in the library that do not bind to RBD) are identified, the binding performance of select aptamers will be characterized. Future work will quantitatively compare our CompELS workflow to SELEX as an aptamer discovery method to highlight differences and any advantages of our newer screening method.