Screening of a Sialyllactose-Specific Aptamer and Engineering a Pair of Recognition Elements with Unique Fluorescent Characteristics for Sensitive Detection of Sialyllactose

Abstract

A single-stranded DNA (ssDNA) aptamer specific for 6′-sialyllactose (6′-SL) was screened through magnetic separation-based SELEX and post-SELEX truncation and used to construct unique aptamer bio-dots for sensitive detection of 6′-SL. Eighteen rounds of screening were conducted during the SELEX process. The ssDNA aptamer Apt9 (K d = 152.3 nM) with a length of 79 nucleotides (nt) was demonstrated as the optimal aptamer candidate after affinity and specificity evaluation. Then, Apt9 was truncated and optimized according to secondary structure and molecular docking. A 35 nt truncated aptamer Apt9-1 (K d = 91.75 nM) with higher affinity than Apt9 was finally obtained. Furthermore, Apt9-1 was used to synthesize bio-dots as a new recognition element of 6′-SL, and the aminobenzene boric acid functionalized carbon dots were employed as the other recognition element. With the respective fluorescent characteristics, the two quantum dots (QDs) were made a pair to construct a 6′-SL fluorescent biosensor. The linear detection range of the biosensor is 10 μM to 5 mM, and the detection limit is 0.9 μM. With the advantages of time-saving, high efficiency, and simplicity in the actual sample detection, the screened aptamer and dual-QD-based biosensor have broad application prospects in 6′-SL detection.

Cell-Surface Glycan Labeling and Sensing

“…The K d for the aptamer with the highest affinity to sialyllactose (SL-11) was 4.9 µM. Zhou's group screened an ssDNA aptamer for sialyllactose with an affinity of 152.3 nM using magnetic separation-based SELEX and post-SELEX truncation methods [48].…”

Single-Round Circular Aptamer Discovery Using Bioinspired Magnetosome-Like Magnetic Chain Cross-Linked Graphene Oxide

“…Traditionally, aptamers are selected from the random-sequence oligonucleotide libraries via systematic evolution of ligands by exponential enrichment (SELEX) [ 20 – 22 ]. A typical SELEX procedure requires iterated rounds of binding–separation–amplification, until reaching the desired binder (target-binding oligonucleotides)-to-nonbinder (nonbinding oligonucleotides) ratio [ 23 , 24 ]. However, this multiround selection procedure is inherently prone to failure because of ineffective separation, polymerase chain reaction (PCR) amplification bias, and even conformational changes of the targets or aptamer candidates being selected [ 25 , 26 ].…”