Figure 3. Expression of the mir-35 family is regulated by chronic hypoxia.

(A–B) qRT-PCR showing mir-35 family member expression levels in wild type embryos exposed to 21% O₂ (black bars) or 0.5% O₂ (orange bars) for 20 mins (A) or 4 hrs (B). The level of normoxic expression was set to 1 for each of the three repetitions. (C) Graphical representation of mir-35 family member abundance in normoxia. mir-38 and mir-41 are less abundant than the other family members, even from those miRNAs transcribed in the same cluster. Values on the graph are logarithmic functions with base 10 of the fold change value for each miRNA. mir-41 showed the lowest relative abundance and was arbitrarily set as the value 1. Primer efficiencies are 116%, 118%, 116%, 113%, 93%, 93%, 109% and 118% for each respective miRNA. (D) Intensity of GFP expression driven by the mir-35–41 promoter is unaffected by 4 hrs of hypoxic exposure. The transgene used is wwIs8[pmir-35–41::GFP + unc-119(+)]. (E–F) gfp transcription, driven by the mir-35–41 promoter in wild type embryos exposed to 21% O₂ (black bars) or 0.5% O₂ (orange bars) for 20 mins (E) or 4 hrs (F). Data are presented as means of at least 3 independent repetitions and error bars represent ± SD. Students t-test was used to assess for statistical significance. *p ≤ 0.05, **≤0.01, ***p ≤ 0.001, ****≤ 0.0001, n.s. = not significant.