Figure 3. Co-targeting of the 3′ NCR for multiple tick-specific miRNAs does not prevent the emergence of escape mutants in ISE6 cells.

(A) Schematic representation of recombinant LGTV genomes used in this study. Top: diagram indicating sites for miRNA target insertions in the predicted secondary structure of the 3′ NCR of LGTV. Bottom: recovered viruses with specific composition of the miRNA target(s) inserted into the 3′ NCR. (B) Growth kinetics of wt-EcoR* and miRNA targeted LGT viruses in ISE6 cells infected at an MOI of 0.01. (C) The titer (±standard deviation) of miRNA targeted viruses after 2^nd passage in ISE6 cells. To initiate the 2^nd cycle of replication, ISE6 cells were infected at an MOI of 1 and the mean virus titer in the cell supernatants was determined in two replicates in Vero cells.