Establishment Real- time PCR for Detection Noroviruses in Stool Specimens
Veterinary Faculty, Microbiology Department, ALEPPO University, ALfurat University, SYRIA
Molecular Pathogens, 2012, Vol. 3, No. 2 doi: 10.5376/mp.2012.03.0002
Received: 14 May, 2012 Accepted: 21 May, 2012 Published: 23 May, 2012
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Preferred citation for this article:
Hasan, 2012, Establishment Real-time PCR for Detection Noroviruses in Stool Specimens, Molecular Pathogens, Vol.3, No.2 6-11 (doi: 10.5376/mp.2012.03.0002)
Noroviruses are usually detected in stool specimens. Such samples commonly contain components reported to be (RT-) PCR inhibitors as well as the main inhibitory substance is the reverse transcriptase enzyme itself. This study provides a complex overview of the influence of RT systems, on the efficiency of PCR to detect Norovirus in stool specimens. The efficiency of three commonly used reverse transcriptases, namely Moloney Murine Leukemia Virus (M-MuLV), Avian Myeloblastoma Leukemia Virus (AMV) and Improm II reverse transcriptase (RT), have been compared to RT-PCR. The study used total RNA extracted from the same stool sample, no bias could therefore be attributed to sample preparation or host individuality. All amplification reactions were run using starting material from the stock, 1/10, 1/100 and 1/1000 cDNA dilutes.
In the results, Improm II yielded very weak bands, in the presence of the stock cDNA and completely failed to amplify DNA in the presence of the 1/10, 1/100, 1/1000 cDNA dilute. Whereas AMV yielded bands in the presence of the stock and 1/10 cDNA dilute and M-MuLV yielded bands in the presence of the stock and all cDNA dilutes.
Noroviruses; Reverse transcriptase; PCR; Stool