Aptamers是通过Exponential Enrichment（SELEX）系统性进化配体生成的单链（ss）核酸分子的一类，涉及耗时繁琐的选择、扩增和富集步骤的迭代。为了弥补传统SELEX的缺点，我们设计了一种基于计算机的方法，以实现aptamer选择的成本效益和容易性。在这里，我们报告了使用具有对AR的自然亲和力的雄激素反应元素（ARE）对DNA aptamers进行分离的结果。准备了一个ARE序列的虚拟库，并经过严格的选择标准，以生成具有稳定的发夹构象和高GC含量的序列池。所选的单链ARE的3D结构经过刚性对接和分子动力学（MD）模拟来检查它们作为AR结合物的潜力。通过直接酶标记aptasorbent分析（ELASA），进一步验证了预测序列，包括在复杂生物环境下测量它们的结合亲和力、特异性和目标鉴别特性。选择了一个简短的15个核苷酸（nts）ssDNA aptamer，称为ARapt1，其估计Kd值为5.5±3 nm，根据计算机模拟和体外评估结果的一致性，被选定为最突出的AR aptamer。ARapt1的高目标结合亲和力和选择性表明其作为与前列腺癌和相关疾病相关的诊断应用的多功能工具的潜在用途。 ©2023 Wiley-VCH GmbH。

Aptamers are a class of single-stranded (ss) nucleic acid molecules generated through Systematic Evolution of Ligands by Exponential Enrichment (SELEX) that involves iterations of time-consuming and tedious selection, amplification, and enrichment steps. To compensate for the drawbacks of conventional SELEX, we have devised an in-silico methodology that facilitates a cost-effective and facile manner of aptamer selection. Here, we report the isolation of DNA aptamers against androgen receptors (ARs) using androgen response elements (ARE) that possess natural affinity toward AR. A virtual library of ARE sequences was prepared and subjected to a stringent selection criterion to generate a sequence pool having stable hairpin conformations and high GC content. The 3D-structures of the selected ss AREs were modeled and screened through rigid docking and molecular dynamic (MD) simulation to examine their potency as potential AR binders. The predicted sequences were further validated using direct enzyme-linked aptasorbent assay (ELASA), which includes the measurement of their binding affinity, specificity, and target discrimination properties under complex biological enviroments. A short, 15 nucleotides (nts), ssDNA aptamer, termed ARapt1 with the estimated Kd value of 5.5 ± 3 nm, was chosen as the most prominent aptamer against AR based on the coherence of both the in-silico and in-vitro evaluation results. The high target-binding affinity and selectivity of ARapt1 signify its potential use as a versatile tool in diagnostic applications relevant to prostate cancer and related diseases.© 2023 Wiley-VCH GmbH.