title: Development and evaluation of probe based real time loop mediated isothermal amplification for Salmonella: A new tool for DNA quantification.
abstract: 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 20min. 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.
authors.affiliation: Division of Bacteriology & Mycology, Indian Veterinary Research Institute, Izatnagar 243122, UP, India.
authors.forename: Mohmad
authors.initials: M
authors.lastname: Mashooq
authors.affiliation: Division of Veterinary Biotechnology, Indian Veterinary Research Institute, Izatnagar 243122, UP, India.
authors.forename: Deepak
authors.initials: D
authors.lastname: Kumar
authors.affiliation: Division of Bacteriology & Mycology, Indian Veterinary Research Institute, Izatnagar 243122, UP, India.
authors.forename: Ankush Kiran
authors.initials: AK
authors.lastname: Niranjan
authors.affiliation: Division of Bacteriology & Mycology, Indian Veterinary Research Institute, Izatnagar 243122, UP, India.
authors.forename: Rajesh Kumar
authors.initials: RK
authors.lastname: Agarwal
authors.affiliation: CADRAD, Indian Veterinary Research Institute, Izatnagar 243122, UP, India. Electronic address: rathorerajesh@rediffmail.com.
authors.forename: Rajesh
authors.initials: R
authors.lastname: Rathore
mesh_terms: D017931:DNA Primers; D015342:DNA Probes; D004269:DNA, Bacterial / Q000032:analysis*; D004755:Enterobacteriaceae / Q000235:genetics; D021141:Nucleic Acid Amplification Techniques / Q000191:economics / Q000379:methods*; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D012475:Salmonella / Q000235:genetics* / Q000302:isolation & purification; D012680:Sensitivity and Specificity
chemical_list: D017931:DNA Primers; D015342:DNA Probes; D004269:DNA, Bacterial
keywords: DNA; Enterobacteriaceae; RT LAMP; Salmonella; Taqman based qPCR