Development and evaluation of probe based real time loop mediated isothermal amplification for Salmonella: A new tool for DNA quantification

被引:39
作者
Mashooq, Mohmad [1 ]
Kumar, Deepak [2 ]
Niranjan, Ankush Kiran [1 ]
Agarwal, Rajesh Kumar [1 ]
Rathore, Rajesh [3 ]
机构
[1] Indian Vet Res Inst, Div Bacteriol & Mycol, Izatnagar 243122, Uttar Pradesh, India
[2] Indian Vet Res Inst, Div Vet Biotechnol, Izatnagar 243122, Uttar Pradesh, India
[3] Indian Vet Res Inst, CADRAD, Izatnagar 243122, Uttar Pradesh, India
关键词
RT LAMP; Taqman based qPCR; Salmonella; DNA; Enterobacteriaceae;
D O I
10.1016/j.mimet.2016.04.014
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
A one step, single tube, accelerated probe based real time loop mediated isothermal amplification (RT LAMP) assay was developed for detecting the invasion gene (InvA) of Salmonella. The probe based RT LAMP is a novel method of gene amplification that amplifies nucleic acid with high specificity and rapidity under isothermal conditions with a set of six primers. The whole procedure is very simple and rapid, and amplification can be obtained in 20 min. Detection of gene amplification was accomplished by amplification curve, turbidity and addition of DNA binding dye at the end of the reaction results in colour difference and can be visualized under normal day light and in UV. The sensitivity of developed assay was found 10 fold higher than taqman based qPCR. The specificity of the RT LAMP assay was validated by the absence of any cross reaction with other members of enterobacteriaceae family and other gram negative bacteria. These results indicate that the probe based RT LAMP assay is extremely rapid, cost effective, highly specific and sensitivity and has potential usefulness for rapid Salmonella surveillance. Crown Copyright (C) 2016 Published by Elsevier B.V. All rights reserved.
引用
收藏
页码:24 / 29
页数:6
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