Multiple GO-SELEX for efficient screening of flexible aptamers

Abstract

We describe a simple, high-speed, high-throughput aptamer screening for a group of small molecules using graphene oxide (simple Multi-GO-SELEX) without immobilizing targets. The affinities of ten different ssDNA aptamers successfully obtained for three pesticides were in the range of 10-100 nM. Besides a specific aptamer for each target, we found a couple of flexible multi-target aptamers, which can bind with 2 or 3 different molecules. These flexible aptamers developed for binding with a mixture of targets are not only significant for the rapid screening of a group of small molecules but also offer great promise for aptamer-based biosensor applications.

Controlling uncertainty in aptamer selection

“…In light of these results, we revisit the topic of optimizing target concentration as discussed in previous works [14][15][16], and show that the assumed initial KD distribution strongly influences protocol optimizations. We also provide additional insights regarding non-covalent ligand immobilization to support more recent efforts to develop robust protocols for small molecule SELEX [26][27][28]. Integrating these ideas, we show that simultaneously lowering the target concentration and the substrate binding dissociation constant over the SELEX cycles can lead to improved selection outcomes for a wide range of initial conditions.…”

“…Altering the length of this fixed sequence is a means to tune KS. Moreover, different immobilization techniques, such as the use of graphene oxide [27,28], will lead to variations of KS within a given pool, but we do not consider such cases here and consider KS to be constant throughout a single cycle of SELEX. Combining ligand-target and ligand-substrate binding, the full system of steady-state equilibrium binding conditions can be described by the set of equations:…”

A Bottom-Up Approach for Developing Aptasensors for Abused Drugs: Biosensors in Forensics

“…However, the chemical coupling of the small-molecule target through its functional group may severely affect aptamer-target interactions [39]. Thus, the Capture-SELEX [40][41][42] and Graphene oxide-assisted SELEX (GO-SELEX) [43,44] methods have been developed to eliminate the target immobilization. Both methods have been successfully used to develop aptamers for a number of small molecules [45][46][47][48].…”

Aptamer-Based Biosensing with a Cationic AIEgen

“…Aptamers are single-stranded DNA or RNA oligonucleotides selected from large oligonucleotide sequence libraries using an in vitro technique known as 'systematic evolution of ligands by exponential enrichment' (SELEX). [1,2] As each aptamer assumes distinct secondary and tertiary conformations, they possess high binding affinity and specificity to a wide variety of ligands, including metal ions, [3][4][5] small molecules, [6][7][8] peptides, [9,10] proteins, [11][12][13] and even microorganisms. [14][15][16] Compared with monoclonal antibodies, aptamers offer similar binding profiles with many additional advantages such as inexpensive synthesis, better chemical and thermal stability, reusability, improved tissue penetration, and low immunogenicity.…”