The AtSCL26 Transcription Factor controls cross-talk between GA-and Nitrogen control of root architecture in Arabidopsis thaliana Roots

We characterised mutants in the GRAS family transcription factor AtSCL26 in Arabidopsis thaliana using a combination of gene functional analysis, hormone treatments and expression profiling at the cell type level. This has enabled us to implicate AtSCL26 in the cell-specific control of nitrogen-giberellic acid response cross-talk to control root architecture. The whole experiment was carried out in triplicate with 24 microarrays in total (8 experiments). The response of cortical and epidermal cells, isolated using Fluorescence Activated Cell Sorting (FACS) was analysed after 2 hours of 5mM KNO3 treatment in Arabidopsis thaliana Col0 and atscl26 mutant roots; both genotypes carried GFP reporters marking either epidermal or cortical root cells. Gene expression was analysed using NimbleGen 12 x 135K microarrays, designed for the TAIR10 genome. Seedlings were grown on low nitrogen (0.1mM KNO3) agar plates for 9 days, then 5mM KNO3 was used as a nitrogen treatment for 2 hours compared to non- treated seedlings kept on low nitrogen plates for 2 hours. At the end of the treatment, roots were harvested, protoplasts generated and GFP-marked epidermal and cortical cells isolated according to Gifford et al 2008 PNAS. After normalisation two replicates from the complete set were removed since they had low QC values (atscl26 epidermal cells 2hr KCl control treated, biological rep3 and Col0 cortical cells 2hr KNO3 nitrogen treated, biological rep3).