Transcriptome profiling of Arabidopsis roots upon lateral root induction upon shy2-2 or slr-1 expression

To identify genes involved in the early phases of lateral root initiation, we profiled the transcriptomes of plants synchronously induced for lateral root initiation after 0, 1, 2, 4 and 6h of auxin treatment in conditions where IAA14 or IAA3-dependent auxin signaling is blocked. For this we used seedlings expressing non-degradable versions of the AUX/IAAs IAA14 (slr-1) or IAA3 (shy2-2) fused to the glucocorticoid receptor domain (slr-1:GR or shy2-2:GR) under the control of the pericycle and founder cell specific GATA23 promoter. Treatment with dexamethasone induces, specifically in pericycle cells, the nuclear translocation of the non-degradable AUX/IAA that acts as a dominant repressor of auxin signaling resulting in a complete block of lateral root formation Overall design: Samples were prepared for harvesting following a modified version of the lateral root induction system protocol (Himanen, K. et al. The Plant Cell 14, 2339–2351 (2002)). Seeds of the pGATA23::shy2-2:GR and pGATA23::slr-1:GR lines were germinated on Nitex membranes (Sefar) on 0.5X MS medium and transferred 4d after germination under 16h light/8h dark conditions to 10µM NPA for 22h and shifted to plates containing 10µM NPA and 10µM DEX or DMSO control for 2h. Seedlings were then transferred to plates containing 10µM IAA, and 10µM DEX and DMSO control. Root tissue was harvested at 0, 2, 4 and 6 h after transfer onto auxin plates. All sampling points were performed in triplicate. For each sample, app. 200 segments of the lower two-thirds of the seedling roots were pooled.