Loop-Mediated Isothermal Amplification assay for rapid detection of G1 human Rotavirus
Subject Areas : International Journal of Bio-Inorganic Hybrid Nanomaterials
Keywords: Nested PCR, LAMP, Isothermal, Rotavirus, VP7 gene,
Abstract :
We developed a Loop-Mediated Isothermal Amplification (LAMP) assay for rapid detection of the most prevalent Rotavirus genotype (G1) responsible for the hospitalization of children worldwide, and compared the sensitivity of LAMP with nested-PCR. A total of 365 stool samples from young children were analyzed by using 6 set of primers targeting conserved sequences of VP7 gene, within 90 min, under isothermal conditions at 62°C, by only a regular laboratory heat block. Then, the LAMP products were analyzed by using gel electrophoresis and the naked-eye after adding SYBR Green I. A ladder pattern on gel electrophoresis was observed specifically only for G1 Rotaviruses and not for other viruses. This LAMP reaction had the same sensitivity as a nested-PCR assay, the detection limit for the both systems were found to be 10 copies ml-1 of G1 Rotavirus RNA. The LAMP assay reported here is faster than nested PCR, cost-effective, and easy to perform and will be valuable tool for rapid and reliable clinical diagnosis of Rotavirus in developing countries.