Nuclease escape elements protect messenger RNA against cleavage by multiple viral endonucleases

Mandy Muller; Britt A. Glaunsinger

The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. . https://doi.org/10.1101/155457 doi: bioRxiv preprint two fractions. Beads containing the fraction used for western blotting were resuspended in 489 30µL lysis buffer. Beads containing the fraction used for RNA extraction were resuspended 490 in Proteinase K buffer (NaCl 100mM, Tris pH 7.4 10mM, EDTA 1mM, SDS 0.5%) containing 491 1µL of PK (Proteinase K). Samples were incubated overnight at 65°C to reverse 492 crosslinking. Samples to be analyzed by western blot were then supplemented with 10µL of 493 4X loading buffer before resolution by SDS-PAGE. RNA samples were resuspend in Trizol 494 and were processed as described above. The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. transfected with the indicated GFP reporters mutated within G-SRE at the residues marked 801 by a * in (B) along with a control empty vector (mock) or a plasmid expressing SOX. After 802 24 h, total RNA was harvested and subjected to RT-qPCR to measure GFP mRNA levels. 803 control non targeting siRNAs. 48h later, cells were transfected with the GFP-GADD45B-820