Differential binding cell-SELEX method to identify cell-specific aptamers using high-throughput sequencing

Abstract

Aptamers have evolved as a viable alternative to antibodies in recent years. High throughput sequencing (HTS) has revolutionized the aptamer research by increasing the number of reads from few using Sanger sequencing to millions of reads using HTS approach. Despite the availability and advantages of HTS compared to Sanger sequencing there are only 50 aptamer HTS sequencing samples available on public databases. HTS data for aptamer research are mostly used to compare sequence enrichment between subsequent selection cycles. This approach does not take full advantage of HTS because enrichment of sequences during selection can be due to inefficient negative selection when using live cells. Here we present differential binding cell-SELEX (systematic evolution of ligands by exponential enrichment) workflow that adapts FASTAptamer toolbox and bioinformatics tool edgeR that is mainly used for functional genomics to achieve more informative metrics about the selection process. We propose fast and practical high throughput aptamer identification method to be used with cell-SELEX technique to increase successful aptamer selection rate against live cells. The feasibility of our approach is demonstrated by performing aptamer selection against clear cell renal cell carcinoma (ccRCC) RCC-MF cell line using RC-124 cell line from healthy kidney tissue for negative selection.