福氏志贺菌核酸适配体的SELEX筛选及特异性研究
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浙江省科技计划项目(2017C32072)


Selection and Characterization of DNA Aptamers Specific for Shigella flexneri by SELEX
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    摘要:

    为探究核酸适配体技术在食源性致病菌检测和鉴定方面的应用,以福氏志贺菌为研究对象,采用全细胞SELEX技术筛选与其特异性结合的适配体。经多轮正筛和反筛,共克隆得到17条序列,4条高频适配体(F-1、F-3、F-9和F-16)对福氏志贺菌有较高的亲和力,其中F-1和F-9与福氏志贺菌的亲和力较高, 相应的亲和常数Kd值分别为(51.3±3.25) nmol/L和(42.6±7.11) nmol/L。软件模拟F1和F9的二级结构,其最小自由能分别为-7.39 kcal/mol和-6.98 kcal/mol。本文首次筛选出对福氏志贺菌具有较高亲和特异性的适配体,为后续利用适配体进行食源性致病菌快速检测奠定了基础。

    Abstract:

    To investigate the application of DNA aptamers on detection and identification of food-borne pathogens, the whole-bacteria SELEX method was used to select single-stranded(ss) DNA aptamers with high binding affinity and specificity to Shigella flexneri. After multiple rounds of selection and counter-selection, 17 supposed aptamers were obtained from the enriched pool. Among them, 4 high-frequency aptamers were found to have significant affinities and specificities to S. flexneri. Further analysis indicated that F-1 and F-9 displayed higher affinities, and their binding affinity to S. flexneri were (51.3 ± 3.25) nmol/L and (42.6 ± 7.11) nmol/L, respectively. According to the Mfold software, the minimum free energy (dG) of F-1 and F-9 were determined to be -7.39 and -6.98 kcal/mol, respectively. The present study demonstrated the feasibility to detect S. flexneri by aptamers, which would pave the way for its application in diagnosing food-borne pathogens.

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冯俊丽;田小兰;汪 艺.福氏志贺菌核酸适配体的SELEX筛选及特异性研究[J].中国食品学报,2020,20(6):256-262

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  • 在线发布日期: 2020-07-07
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