The data reveal numerous, Toxoplasma-specific changes to host miRNA levels and some of these changes are at least partly mediated at the transcriptional level. The Ginkgolide-C microarray profiling data showed that the miR-17 family and the miRNA families that are (-)-Licarin-B co-transcribed with it were generally more abundant in RNA samples from Toxoplasmainfected cells and that the abundance of these miRNAs marginally increased at 6-hours post-infection and showed greater increases at 12-hours and 24-hours post-infection. MiR-21 is a highly abundant unrelated miRNA in HFFs that is derived from an independent intergenic locus and is included in the microarray heat-map and subsequent analyses for comparison. To validate the microarray data and to specifically measure the effects of Toxoplasma infection on miRNAs encoded by miR-17,92, miR-106a,363 and miR-106b,25, we took two orthogonal approaches. First, we performed high-resolution northern blot analysis for miR-17. Consistent with the microarray data, the northern blot results showed that, collectively, the level of mature miR-17 family members increased,2.7-fold in Toxoplasmainfected HFFs relative to uninfected HFFs. The data also showed that the pre-miR-17 signal was a small fraction of the hybridization signal observed for mature miR-17, suggesting that the contribution of pre-miR-17 to the observed miRNA microarray hybridization intensity was negligible, an interpretation that is consistent with previous data. The second approach used to follow up on the microarray results was primer-extension analysis with primers specific for each miR family. The raw data shown in Figure 2A confirmed that the levels of mature miR-17 family members increased as a function of time in RNA samples derived from Toxoplasma-infected HFFs. Primer-extensions performed with probes specific for 5S rRNA and miR-21 gave very similar patterns to each other with no significant effect of Toxoplasma infection. MiR-18, miR19 and miR-25 family members also showed an increase in abundance in RNA samples derived form Toxoplasma-infected HFFs.