A. thaliana WT and ICT1 mutant treated vs. non treated with I3C

We work with Arabidopsis thaliana. One of the things we focus on is a secondary metabolite named Indol 3 carbinol (I3C). This molecule is synthesized following insect attack (wounding). We have noticed that addition of I3C to the growth medium causes inhibition of plant/bacterial/fungal growth. We have discovered that I3C also affects different pathways in the plant. By screening overexpression library we have found Arabidopsis mutants that overexpress S30 gene (ribosomal subunit of 40s) are tolerant to the addition of I3C (their growth is less affected by the addition of I3C). To understand better I3C mode of action we have planned the following transcriptome experiment: 1. To see what are the pathways activated following wounding, we took the wt Arabidopsis thaliana (Col-0) and kept one group of plants untreated (control) and the other was wounded (8 hours). 2. To see what are the pathways activated following incubation with 2 concentrations of I3C, we took the wt Arabidopsis thaliana (Col-0) and kept one group of plants untreated (control) and the other was incubated for 2 hours with 200µM (physiological concentration) and 400µM I3C. 3. To see if there are any differences in gene expression between the wt and the mutants, we took wt and 2 I3C tolerant mutants. Neither of them was treated. 4. To see if in the mutant other pathways are activated following incubation with I3C, both mutants were incubated in I3C. In all cases we have used the roots only for RNA extraction. Overall design: The samples: • Colombia 0– wt (used as reference) • psx08098 (3) homozygous line (FOX overexpression library) • 2(5) homozygous line (overexpressing transgene made in the lab) Treatments: • no treatment (DMSO) • I3C 200uM • I3C 400uM • Wounding Triplicate of each sample were used Colombia no treatment (DMSO) X3 Colombia I3C 200uM X3 Colombia I3C 400uM X3 Colombia Wounding X3 Psx08098 (3) no treatment (DMSO) X3 Psx08098 (3) I3C 200uM X3 2(5) no treatment (DMSO) X3 2 (5) I3C 200uM X3 2 (5) I3C 400uM X3