FIGURE 2: RNA interference on VTC4 in L. guyanensis. (A) Schematic map of SSU rRNA locus in Leishmania and targeting of VTC4. Regions are the SSU rRNA (grey box), VTC4 region (black arrow), puromycin-resistance gene ORF (PAC, white arrow) and the stem loop stuffer fragment (striped box). (B) Relative VTC4 mRNA levels were assessed by qRT-PCR with VTC4 and KMP11 gene primers. Primer sequences are presented in Table S2. (C) Relative polyP was quantified by digesting the extracted polyP and measuring the released Pi colorimetrically. A and B; 21, 22, 23 and 24; 83 and 85 represented recombinant Leishmania clones selected by antibiotic resistances, for more detail see Fig. S2. Results of a pool of 3 independent experiments were expressed as mean ± SD. Statistical significance was assessed by Student’s t-test (B and C); * p < 0.05, *** p < 0.001, ns: non-significant.