## 3 Results and Discussion
### Effect of Selenocyanates on Arabidopsis Seeding Root Growth  
This assay was included in the study to unravel the effects of selenocyanates in a multicellular model organism, such as _Arabidopsis thaliana_ along with unicellular yeast. _A. thaliana_ mutants, deficient in glutathione, are available and commonly employed to evaluate the effects of compounds targeting the glutathione pool and metabolism. The inhibition of glutathione deficient mutants in this assay confirms the results of chemogenetic screening in a multicellular organism. The effects of selenocyanates on the growth of _Arabidopsis_ seedling roots were observed by employing wildtype, as well as mutants deficient in glutathione. employed. A threshold concentration of glutathione is an important prerequisite  
Figure 7: The streak test technique for comparing the inhibition of wt and different mutants by the compounds. This assay does not give such a clear result as the plate inhibition zone test (Figure 5), however wt and several mutants can be tested on a single Petri plate. (a) In the example shown, 20 \(\upmu\)L of 1 mM of test compound was pipetted into the central well. (b) Graphs showing the zones of inhibition of wt and mutants treated with compound 1 (Graph on the left side) and compound 2 (Graph on the right side) Values represent mean \(\pm\) S.D. \({}^{*}\)\(p<0.05\), \({}^{**}\)\(p<0.01\) and \({}^{***}\)\(p<0.001\). The growth inhibition zone of mutants was compared with that of wt after treatment with selenocyanates.  
for root growth [52]. Seeds were allowed to grow for three days before placing them on medium containing the corresponding concentration of the respective selenocyanate and after three days of continual exposure, root length was measured [30]. _Arabidopsis_ Col-0 wt and _gr1_ and _pad2_ mutants in the Col-0 background were employed in the study. The _gr1_ mutant line is a knockout mutant of glutathione reductase and cannot reduce GSSG back to GSH. So, it has a higher proportion of GSSG in the glutathione pool [53]. The _pad2_ line is mutated in the glutamate cysteine ligase gene and contains only approximately 20% of the GSH level as compared to wt [54]. The mutants _pad2_ and _gr1_, compromised respectively in GSH synthesis and GSSG reduction, are more sensitive than the Col-0 wildtype upon treatment with selenocyanates. Thus, the root growth was impaired in these mutants with reference to wt (Figure 8). The enhanced sensitivity of mutants, deficient in genes coding for glutathione synthesis compared to the Col-0 wt, confirms the results of the chemogenetic screening with yeast mutants and again highlights the central role of cellular GSH in the resistance of cells to selenocyanates.  
Figure 8: Effect of selenocyanates on _Arabidopsis_ root growth. (**a**) Shows the phenotypes of Col-0 (wt), _pad2_ and _gr1_ seedlings observed after three days of continual exposure to 20 \(\upmu\)M of selenocyanates in the growth medium. Scale bar = 1 mm. (**b**) Shows the effect of continual exposure to 5, 10 and 20 \(\upmu\)M of selenocyanates on root growth of Col-0 (wt), _pad2_ and _gr1_ seedlings. The box plots show the range of individual measurements, the mean \(\pm\) standard deviation, and the median value indicated as a horizontal line. C1—Compound **1** and C2—Compound **2**. Values represent mean \(\pm\) S.D. **\(p<0.01\) and ***\(p<0.001\).