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. 2014 Sep 8;9(9):e106477. doi: 10.1371/journal.pone.0106477

Figure 1. Antisense sRNAs to EhAgo2-2 are functional but do not silence gene.

Figure 1

(A) High resolution Northern blot of the trigger-EhAgo2-2 cell line probed for AS sRNAs to EhAgo2-2, EHI_188130 (loading control), and EHI_197520 (trigger) at 24 µg/ml drug and at 1 and 4 weeks without drug. Abundant AS sRNAs to EhAgo2-2 diminish over time with plasmid removal. (B) Northern blot for EhAgo2-2 transcripts in untransfected parasites and in the trigger-EhAgo2-2 cell line at 24 µg/ml drug and at 6 weeks post drug removal. (C) Luciferase assays in the trigger-EhAgo2-2 cell line transiently transfected with a luciferase expression plasmid (CS-luc), a promoter-less luciferase plasmid (pKTluc), or with the Ago2-2 trigger-luciferase plasmid. Parasites were maintained at 6 µg/ml drug or were removed from drug for 8 weeks. Luciferase expression was inhibited in the presence of an Ago2-2 trigger and restored when trigger-EhAgo2-2 plasmid was lost. Luciferase values were normalized to the CS-Renilla-luc expression. Experiments were performed in triplicates; average and standard errors are shown.