A feasible approach to find mutants in your favourite pathway

The concentration of IAA is determined by three interconnected factors: (1) synthesis, (2) transport, and (3) degradation. Despite years of research by the scientific community on IAA and its action, many questions remain regarding its metabolism. In this study, we designed a direct genetic approach to generate, identify, and characterize new factors involved in this process.

To achieve this, we mutagenized with ethyl methanesulfonate (EMS) two transgenic lines carrying reporter transgenes for auxin signaling. In the M2 generation, a screen was carried out for mutant lines whose auxin signaling or plant morphology differed from the corresponding wild-type strain. As a result, 325 candidate lines were identified and subjected to an auxin profiling analysis using liquid chromatography coupled with mass spectrometry.

We then performed multivariable analysis with all the metabolomics data to select the most promising mutants. In this work, three mutants were selected, and after identification through mapping-by-sequencing, we demonstrated that they carried mutations in genes whose roles in auxin metabolism had been previously published.

In conclusion, we have demonstrated that this is a viable approach for identifying mutants in complex metabolic pathways.