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African Journal of Biotechnology

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Comparison of various commercial products for phenol-guanidine-based classical swine fever virus RNA extraction

MA Hofmann, M Mader

Abstract


TRIzol®, TRI Reagent®, and RNAzol® are widely used commercial reagents for the extraction of cellular or viral RNA. Several other brand name products, some of which are advertised for the processing of specific sample types such as blood, are also available. Here, we compare the efficiency of these products for classical swine fever virus RNA extraction from cell culture supernatant, serum, and tonsil tissue, assessed by quantitative RT-PCR. Furthermore, the detection of a synthetic RNA transcript used as an internal positive control for extraction and RT-qCR was compared as well. Most tested products showed a similar extraction efficiency, and none of the products recommended for specific sample types performed better than the all-purpose reagents. We also show that the homogenization method for tissue samples has a significant impact on the detection efficiency of the RNA after extraction from the homogenized tissue. Homogenization of 100 mg tissue in 5 ml cell culture medium and using an UltraTurrax® tissue grinder yielded the best results, whereas TissueLyser®-mediated homogenization in 1 ml cell culture medium or direct homogenization in RNA extraction medium proved to be less efficient.

Keywords: RNA extraction, homogenization, comparison, TRIzol, TRI reagent, reverse transcription-quantitative PCR (RT-qPCR)

African Journal of Biotechnology, Vol 13(42) 4100-4104



http://dx.doi.org/10.5897/AJB2014.13894
AJOL African Journals Online