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文章摘要
食源性致病菌的核酸可视微阵列检测技术研究
Research on Nucleic Acid Visual Microarray Detection of Food-borne Pathogenic Bacteria
投稿时间:2019-06-14  修订日期:2019-08-06
DOI:
中文关键词: 食源性致病菌  基因组DNA  可视化微阵列  银染
英文关键词: food-borne pathogenic bacteria  genomic DNA  visual microarray  silver staining
基金项目:甘肃省科学院应用开发项目(2016JK-07);甘肃省科技计划资助(18JR3RA249 )。
作者单位E-mail
李云霞 甘肃省科学院传感技术研究所 864315576@qq.com 
马生龙 甘肃省科学院传感技术研究所  
聂莹莹 甘肃省科学院传感技术研究所  
马莉萍 甘肃省科学院传感技术研究所 279559916@qq.com 
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中文摘要:
      建立了一种快速检测三种食源性致病菌的核酸可视微阵列。分别设计与沙门氏菌InvA基因,金黄色葡萄球菌nuc基因,志贺氏菌ipaH基因两端互补的基片探针和检测探针,设计三种基因的特异靶序列,特异靶序列或基因组DNA一端与固定在玻片上的氨基化基片探针杂交,另一端与巯基化检测探针连接形成复合体。使胶体金与检测探针结合,通过银染放大信号,检测三种食源性致病菌DNA。结果表明:通过对检测探针分别与基因组DNA、PCR产物及特异靶序列的杂交银染结果比较,三种结果都显示阳性,表明该方法可直接利用基因组DNA实现对食源性微生物的快速、高效检测,对三种食源性致病菌DNA的检测下限为1 pmol/L,在1 mmol/L ~1 pmol/L浓度范围内得到肉眼可见的清晰结果。检测探针与基因组DNA的杂交实验结果表明,微阵列对三种食源性微生物的检测具有较好的特异性。说明该技术可快速、简便检测食源性微生物,市场前景广阔。
英文摘要:
      A visualized microarray for rapid detection of nucleic acid of three food-borne pathogenic bacteria is developed. Substrate probe and detection probe which complementary to both ends of the gene the Salmonella InvA gene, Staphylococcus aureus nuc gene and Shigella ipaH gene are designed, respectively. Specific target sequences of three genes are designed. One end of the specific target sequence or genomic DNA is hybridized with the amination probe fixed on the slide, and the other end is complementary to the sulfhydrylation probe to form a complex. The detection probe was Connected with colloidal gold, and the signal was amplified by silver staining for the detection of food-borne pathogenic bacteria. The results showed as follows: Through comparative analysis the results of hybridization silver staining with genomic DNA, PCR products and specific target sequences, all results are positive, which demonstrate that the method could directly use genomic DNA to detect food-borne microorganisms rapidly and efficiently. The detection limit of genomic DNA of three foodborne pathogenic bacteria is 1 pmol/L, and clear results could be obtained by naked eyes in the concentration range of 1 mmol/L ~1 pmol/L. The results of hybridization between probe and genomic DNA indicate that visual microarray had good specificity for the detection of microorganisms. In conclusion, this method can detect food-borne microorganisms quickly and easily, and it has broad market prospects.
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