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靶向结直肠癌血清的单链DNA适配体的筛选及初步应用

Selection and preliminary application of a single stranded DNA aptamer targeting colorectal cancer serum.

作者信息

Li Kun, Qi Liqing, Gao LiMing, Shi Ming, Li Jian, Liu ZhiWei, Zhao Lu

机构信息

College of Environmental & Chemical Engineering, Yanshan University Qinhuangdao Hebei Province 066004 China

Applied Chemistry Key Laboratory of Hebei Province, Yanshan University Qinhuangdao Hebei Province 066004 China.

出版信息

RSC Adv. 2019 Nov 27;9(66):38867-38876. doi: 10.1039/c9ra04777h. eCollection 2019 Nov 25.

Abstract

Colorectal cancer is one of the common causes of malignant tumors in recent years, thus the discovery of potential compounds that detect the occurrence of colorectal cancer by efficient approaches is necessary. In this study, the method of systematic evolution of ligands by exponential enrichment (SELEX) was used for recognizing serum from colorectal cancer patients by a single-stranded DNA library of aptamers assisted by single-walled carbon nanotubes (SWCNTs) to remove single-stranded DNA with low affinity. Ten rounds of selection were applied using colorectal cancer serum as a target with the serum of healthy individuals as a control. As the result, we have successfully identified four candidate aptamers after high-throughput genome sequencing analysis, comparison analysis and secondary structure prediction. Among them, aptamer Seq-2 exhibited the highest affinity and the strongest selectivity with an equilibrium dissociation constant ( ) of 11.31 ± 3.25 nM and a difference value of 4.25 ± 0.38 between the colorectal cancer group and the healthy group. Moreover, with fifty negative control serum samples, the positive detection rate of fifty positive serum samples tested by aptamer Seq-2 was over 90%. In particular, aptamer Seq-2 can strongly bind the colorectal cancer serum, less strongly bind the non-colon cancer serum and hardly bind the healthy serum. Therefore, aptamer Seq-2 presents enormous potential for exploring as a tumor diagnostic kit and detecting unknown tumor markers in serum to reflect colorectal cancer.

摘要

结直肠癌是近年来恶性肿瘤的常见病因之一,因此有必要通过有效的方法发现能够检测结直肠癌发生的潜在化合物。在本研究中,指数富集配体系统进化(SELEX)方法用于通过单链DNA适配体文库识别结直肠癌患者的血清,该文库由单壁碳纳米管(SWCNT)辅助,以去除低亲和力的单链DNA。以结直肠癌血清为靶标,以健康个体血清为对照进行了十轮筛选。结果,通过高通量基因组测序分析、比较分析和二级结构预测,我们成功鉴定出四种候选适配体。其中,适配体Seq-2表现出最高的亲和力和最强的选择性,平衡解离常数( )为11.31±3.25 nM,结直肠癌组与健康组之间的差异值为4.25±0.38。此外,对于五十份阴性对照血清样本,用适配体Seq-2检测五十份阳性血清样本的阳性检出率超过90%。特别是,适配体Seq-2能与结直肠癌血清强烈结合,与非结肠癌血清结合较弱,与健康血清几乎不结合。因此,适配体Seq-2在开发肿瘤诊断试剂盒和检测血清中未知肿瘤标志物以反映结直肠癌方面具有巨大潜力。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/811a/9075956/2e6173aede6d/c9ra04777h-f1.jpg

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