Position: Home > Articles > Detection of Cronobacter sakazakii in Infant Formula Powder by Helicase-Dependent Isothermal DNA Amplification Assay
FOOD SCIENCE
2014,35
(4)
155-158
婴儿配方乳粉中阪崎克罗诺杆菌解旋酶恒温基因扩增检测方法的建立
作 者：
周巍;张薇;刘亮;刘东;李永波;田浩;张岩;张志胜
单 位：
河北农业大学食品科技学院;河北省食品质量监督检验研究院
关键词：
解旋酶恒温基因扩增法;婴儿配方乳粉;阪崎克罗诺杆菌;检测
摘 要：
目的:建立一种检测婴儿配方乳粉中阪崎克罗诺杆菌的解旋酶恒温基因扩增方法。方法:根据阪崎克罗诺杆菌ITS基因设计特异性引物,优化解旋酶恒温基因扩增法反应条件UvrD helicase、T4 gp32的浓度,人工添加阪崎克罗诺杆菌确定检出限,多种致病菌在建立的解旋酶恒温基因扩增体系中扩增验证特异性,电泳检测扩增产物。结果:解旋酶恒温基因扩增法检测婴儿配方乳粉中阪崎克罗诺杆菌得到与设计序列长度一致的100 bp基因片段,检出限为10 CFU/g,优化反应条件UvrD helicase、T4 gp32的终浓度分别为0.1、5.0μg。结论:解旋酶恒温基因扩增法用于检测婴儿配方乳粉中阪崎克罗诺杆菌的特异性强、灵敏度高、耗时短,为婴儿配方乳粉中阪崎克罗诺杆菌的快速检测提供了新的方法。
译 名：
Detection of Cronobacter sakazakii in Infant Formula Powder by Helicase-Dependent Isothermal DNA Amplification Assay
作 者：
ZHOU Wei;ZHANG Wei;LIU Liang;LIU Dong;LI Yong-bo;TIAN Hao;ZHANG Yan;ZHANG Zhi-sheng;College of Food Science and Technology, Agricultural University of Hebei;Hebei Institute of Food Quality Supervision Inspection and Research;
关键词：
helicase-dependent isothermal DNA amplification(HDA);;infant formula powder;;Cronobacter sakazakii;;detection
摘 要：
Objective: The aim of this study was to establish a helicase-dependent isothermal DNA amplification(HAD) method for rapid and accurate detection of Cronobacter sakazakii in infant formula powder. Methods: A pair of oligonucleotide primers exclusively targeting the internal transcribed spacer(ITS) gene of Cronobacter sakazakii was designed, and the final concentration of UvrD helicase and T4 gp32 were optimized. Cronobacter sakazakii in infant formula powder was then directly detected by this method and amplification products were separated and detected by agarose gel electrophoresis. Results: An amplicon of 100 bp in length was obtained, which was the same as the designed gene fragment by HDA. The sensitivity of HDA was 101 CFU/g, the optimized concentration of UvrD helicase was 0.1 μg, and T4 gp32 was 5.0 μg. Conclusion: The helicase-dependent isothermal DNA amplification assay provides a new, specific, sensitive and fast for the detection of Cronobacter sakazakii in infant formula powder.
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