RT-qPCR Analysis of WUS Expression

Three biological replicates of Arabidopsis thaliana Col-0 and Lp2-2 root segments were sampled after 3 days on SIM following protocol b (~100 mg fresh weight/sample). RNA extraction was done with the Qiagen RNeasy Plant Mini Kit and cDNA synthesis was done with the Promega GoScript^TM system using random hexamer primers. Yield and purity were assessed by nanodrop. WUS expression was normalized to UBC9 and TIP41L levels and a sample maximization strategy was applied for the plate layout (assays were spread over three runs, including two technical replicates, no-RT controls, and triplicate no-template controls). RT-qPCR was set-up with the GoTaq® master mix using primers described in Supplementary Table 1 and run on a Stratagene Mx3005P cycler. Results were analysed according to the ∆∆Ct method using the R package pcr^{98 (link)} (Supplementary Code).