Innate Immune Priming by cGAS as a Preparatory Countermeasure Against RNA Virus Infection

Michael T. Parker; Smita Gopinath; Corey E. Perez; Melissa M. Linehan; Jason M. Crawford; Akiko Iwasaki; Brett D. Lindenbach

with Protein A-magnetic beads. Washing was performed as follows: 2x with RIPA, 2x 475 with high salt RIPA (500 mM NaCl), 1x with IP-wash buffer (0.5 M LiCl, 1% NP-40, 476 1% deoxycholate, 100 mM Tris-HCl pH 8.0), and 2x with T 10 E 1 (10 mM Tris-HCl pH For RT-qPCR of mouse tissue-derived mRNAs, SYBR Green qPCR reactions 497 were run in triplicate with gene specific primers ( Table 1 ). The Ct values were 498 averaged, internally normalized against housekeeping gene HPRT, then normalized 499 to B6J control mice by using the ∆∆Ct method of comparison. Fold-expression was 500 estimated assuming one doubling per cycle (fold expression = 2 -∆∆Ct ). 501 . CC-BY-NC-ND 4.0 International license author/funder. It is made available under a The copyright holder for this preprint (which was not peer-reviewed) is the . https://doi.org/10.1101/434027 doi: bioRxiv preprint Site-directed mutagenesis of hcGAS was performed by using appropriate 512 primers (Table 1) and PfuTurbo (Agilent Technologies), as previously described 513