I am working on an EMCV like virus with about 8kbp genome. The RNA is usually prepared from digested plasmids by in-vitro transcription, followed by DNase I digestion, followed by P/C and Ethanol cleanup and finally electroporated into Hela cells. However, due to the large fragile nature of this genome, it is hard to get the complete sequence in the cells without it falling appart, and often it has a low read, even if it showed good bands on a gel run on the same day.
So I was thinking and wanted to try skipping the purification procedures all together and just adding EDTA to bind the magnesium and then electroporating. Do you think it would be a good idea? Any other tips and tricks for this problem?
Thanks!
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