Extraction of RNA for detection of dengue and zika virus using RT-PCR, as what I know, RNA degrade easily. Is it necessary for us to extract from fresh sample/sample that kept in -80*C?
While it is almost always better to keep sample at ultra cold temperatures, we tried testing pools of 25 mosquitoes (containing either negative mosquitoes or a single mosquito infected with West Nile virus) that had been held for up to 2 weeks at ambient temperature. I was certain that all of the RNA would long since have been destroyed. However, quantitative RT-PCR results from pools held at room temperature for two weeks were indistinguishable from those tested immediately or those held at -70C. See Turell et al. 2002. Effect of Holding Conditions on the Detection of West Nile Viral RNA by Reverse Transcriptase-Polymerase Chain Reaction from Mosquito (Diptera: Culicidae) Pools. J. Med. Entomol. 39:1-3. Several studies have come out since then with similar findings for dengue, chikungunya (an alphavirus) and Venezuelan equine encephalitis (another alphavirus) viruses.
Thank you so much for the answer, really appreciate it ;)
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