Materials & Manufacturing Directorate, Soft Matter Materials Branch , Air Force Research Laboratory , Wright-Patterson AFB , Ohio 45433 , United States.
ACS Comb Sci. 2018 Oct 8;20(10):585-593. doi: 10.1021/acscombsci.8b00048. Epub 2018 Sep 28.
Competition-enhanced ligand screening (CompELS) was employed to rapidly screen through large DNA libraries to identify single-stranded, oligonucleotide-based ligands called aptamers that bind to a nonbiological target. This previously unreported aptamer screening approach involves the repeated introduction of unenriched random sequence populations during the biopanning process, but avoids iterative elution and polymerase chain reaction (PCR) amplification steps inherent to traditional SELEX (systematic evolution of ligands by exponential enrichment) screening. In this study, 25 aptamers were identified against a gold surface via CompELS and evaluated to identify patterns in primary structures and predicted secondary structures. Following a final one-round competition experiment with the 25 identified aptamers, one particular aptamer sequence (1N) emerged as the most competitive adsorbate species for the gold substrate. Binding analysis indicated at least an order of magnitude difference in the binding affinity of 1N ( K = 5.6 × 10 M) compared to five other high affinity aptamer candidates ( K = 10-10 M) from identical secondary structure families. Collectively, these studies introduce a rapid, reliable screening and ranking platform along with a classification scheme well-suited for identifying and characterizing aptamers for nonbiological as well as biological targets.
竞争增强配体筛选（CompELS）被用于快速筛选大型 DNA 文库，以鉴定与非生物靶标结合的单链寡核苷酸配体，即适体。这种先前未报道的适体筛选方法涉及在生物淘选过程中反复引入未富集的随机序列群体，但避免了传统 SELEX（通过指数富集的配体系统进化）筛选中固有的迭代洗脱和聚合酶链反应（PCR）扩增步骤。在这项研究中，通过 CompELS 鉴定了针对金表面的 25 个适体，并对其进行了评估，以确定一级结构和预测二级结构中的模式。在对 25 个鉴定出的适体进行最终一轮竞争实验后，一种特定的适体序列（1N）成为金底物最具竞争力的吸附物种。结合分析表明，与来自相同二级结构家族的五个其他高亲和力适体候选物（ K = 10-10 M）相比，1N 的结合亲和力至少相差一个数量级（ K = 5.6×10 M）。总之，这些研究介绍了一种快速、可靠的筛选和排序平台，以及一种分类方案，非常适合鉴定和表征非生物和生物靶标适体。
