Selection of DNA Aptamers for Ovarian Cancer Biomarker CA125 Using One-Pot SELEX and High-Throughput Sequencing.
Selection of DNA Aptamers for Ovarian Cancer Biomarker CA125 Using One-Pot SELEX and High-Throughput Sequencing.
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DOI：
10.1155/2017/9879135
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发表时间：
2017
影响因子：
2.3
通讯作者：
Whelan RJ
中科院分区：
文献类型：
--
作者：
Scoville DJ;Uhm TK;Shallcross JA;Whelan RJ
CA125是一种粘蛋白糖蛋白，其血清浓度与女性患卵巢癌的风险相关，也表明确诊患者对治疗的反应。在患者样本中准确检测这种大而复杂的蛋白质具有重要的临床意义。我们认为，通过开发具有CA125亲和力的核酸适体，可能会实现强大的新诊断工具。在这里，我们报告了我们使用One-Pot SELEX分离具有CA125亲和力的单链DNA适体，然后对所选的寡核苷酸进行高通量测序。这种数据丰富的方法与生物信息学工具相结合，使整个选择过程得以表征。利用荧光各向异性和亲和探针毛细管电泳，对4个候选适配体的结合亲和性进行了评价。两个适配体，CA125_1和CA125_12，都没有引物，被发现与临床相关浓度的蛋白靶结合。Mg2+离子的存在对CA125_1和CA125_12的结合有不同的影响。综上所述，One-Pot SELEX被认为是一种很有前途的选择方法，可以获得临床重要蛋白质靶点的DNA适体。
CA125 is a mucin glycoprotein whose concentration in serum correlates with a woman's risk of developing ovarian cancer and also indicates response to therapy in diagnosed patients. Accurate detection of this large, complex protein in patient samples is of great clinical relevance. We suggest that powerful new diagnostic tools may be enabled by the development of nucleic acid aptamers with affinity for CA125. Here, we report on our use of One-Pot SELEX to isolate single-stranded DNA aptamers with affinity for CA125, followed by high-throughput sequencing of the selected oligonucleotides. This data-rich approach, combined with bioinformatics tools, enabled the entire selection process to be characterized. Using fluorescence anisotropy and affinity probe capillary electrophoresis, the binding affinities of four aptamer candidates were evaluated. Two aptamers, CA125_1 and CA125_12, both without primers, were found to bind to clinically relevant concentrations of the protein target. Binding was differently influenced by the presence of Mg2+ ions, being required for binding of CA125_1 and abrogating binding of CA125_12. In conclusion, One-Pot SELEX was found to be a promising selection method that yielded DNA aptamers to a clinically important protein target.