Figure 3: ClkSV40 flies express a stable mRNA that generates ectopic clock cells
From: Clk post-transcriptional control denoises circadian transcription both temporally and spatially
(a) AGO1 binding to endogenous genomic Clk mRNA or ClkV5 from ClkWT or ClkSV40 transgenic fly heads (measured by RT-PCR, ratio of IP vs input is shown, one representative experiment out of three is presented). (b) RT-PCR results showing the ratio of Clk mRNA bound versus unbound- to oligo-dT beads in ClkWT and ClkSV40 fly heads (*P<0.036, student t-test; mean±s.e. n=6 for each genotype, average of five independent insertions of these transgenes were plotted). (c,d) Immunofluorescence (IF) analysis of (c) ClkSV40 or (d) ClkWT Drosophila brains using an anti-V5 antibody (lower panel represents magnification of rectangle area). (e) Expression levels of CLKV5, total CLK and tubulin proteins determined by western blot analysis of head lysates from ClkWT or ClkSV40 flies (five independent insertions for each transgene). The shown experiment is one out of four representative experiments. (f,g) Immunofluorescence analysis of (f) ClkSV40 or (g) ClkWT Drosophila brains using an anti-VRI antibody. (h,i) ClkV5 and endogenous Clk mRNAs levels from ClkSV40 and ClkWT fly heads determined by RT-PCR from (h) total RNA and (i) polyA+-selected RNA (for each time point, the average of five independent insertions for each transgene is plotted). See also Supplementary Movies 1 and 2.
