Supplementary material for : Development of an RNA interference (RNAi) gene knockdown protocol in the anaerobic gut fungus Pecoramyces ruminantium strain C1A
Figure S1. Uptake of fluorescently (Cy3) tagged siRNA by C1A spores. (A) The ldhD-specific siRNA was added to the flooding solution 75 minutes after the onset of flooding followed by incubation for 15 more minutes at 39ºC. Samples (a few microliters) were taken at regular intervals for visualization. The same field is shown for DAPI-, and Cy3-labeled germinating spores (Note that the spores were concurrently stained with DAPI and fluorescing green indicating the uptake of the Cy3-labeled siRNA) (bar=20 μm). (B) Effect of the siRNA treatment on fungal growth rate. siRNA-treated spores were collected and used to inoculate fresh RFC medium. Control cultures were started at the same time using siRNA-untreated spores. Headspace pressure was measured daily and used to calculate fungal biomass as described previously (1). Error bars are standard deviations from at least three replicate cultures for each condition.
Table S1. Transcripts with a significant (False Discovery Rate (FDR) < 0.1) fold change in the ldhD-siRNA-treated cultures.