Fig 1. BAC16 K8.1pr-mIFP2 infected iSLK cells enables fluorescent readout of late gene activity.

a) The late gene reporter is expressed 48–96 hours post-reactivation. iSLK cells infected with either the wildtype BAC16 or the late-gene reporter modified BAC16 were reactivated, and reporter activity was monitored every day for 96 hours. b) Sensitivity of late gene reporter to inhibitors of viral DNA replication phosphonoacetic acid (PAA) and cidofovir (CDV). Reporter activity was monitored by flow cytometry 72 hours after reactivation. Inhibitors of viral DNA replication restricted reporter activity. c) Virion production of late gene reporter virus after transfer of supernatant to uninfected HEK293T cells. 72 hours post-reactivation, viral supernatant was filtered and transferred to naïve HEK293T cells; infection of HEK293T cells was monitored by expression of the BAC16-encoded, constitutive EGFP. Error bars are standard error from three technical replicates. d) Virion production after supernatant transfer to uninfected HEK293T cells. SAFE denotes sgRNAs targeting the host in regions without expected function [47]. Non-coding viral 1 and 2 indicates sgRNAs targeting two viral regions free of ORFs. Each bar represents an independent sgRNA, and error bars represent standard error from three independent reactivations.