An efficient extraction method for short single-stranded DNA from agarose gels in aptamer screening

With the rapid advancements in aptamer screening, the efficient extraction of short single-stranded DNA (ssDNA) from agarose gel has become a new requirement. However, the currently available products are primarily designed for double-stranded DNA (dsDNA) and exhibit limited efficacy when applied to the extraction of short ssDNA. In this study, we successfully developed a novel method based on amino-modified silica-coated magnetic particles (ASMPs) for the extraction of short ssDNA from agarose gel. The gel slices containing short ssDNA were subjected to centrifugation in a spin column/centrifugation tube assembly with silica wool, followed by the adsorption using ASMPs. Subsequently, reagents containing phosphate groups were employed to desorb ssDNA from the surface of ASMPs. Through optimization of each step, we realized remarkable efficiency in the extraction of short ssDNA. To assess the efficacy of our method, we utilized it in aptamer screening. The results demonstrated that our method outperformed three commercially available DNA gel extraction products (Q-kit, S-kit, and V-kit). The relative recovery rates of all methods were as follows: M-dNTP (100.00 %) > M-BB (63.38 %) > Q-kit (46.64 %) > S-kit (15.98 %) > V-kit (0.38 %). The results strongly suggest that the developed method holds promise for short ssDNA extraction from agarose gel.