No abstract

Irrigation of crops accounts for a significant portion of fresh water consumption. In order to utilize this resource more efficiently, it is necessary to engineer crops that can more efficiently use water. Water use efficiency, defined as the ratio of plant growth to water used, is a complex property of plants affected by many different factors. Despite this complexity, genetic variability has been able to be identified in a number of different crops. The C4 model species Setaria viridis remains under-studied in this regard and consequently we sought to identify promising genetic loci contributing to variation in water use efficiency. In order to accomplish this goal we leveraged the high-throughput phenotyping platform at the Donald Danforth Plant Science center to grow S. viridis in well-watered and water-limited conditions. This automated system enables strict control of watering regimes as well as measures of plant traits extracted from photographs using computer vision. Combining these two sets of data allows for direct measurement of whole-plant water-use efficiency on a daily basis which was used as a response variable in a genome wide association study. Significant associations were found for water-use efficiency and related traits. These loci were then prioritized further by pooling information across each day of an experiment and across multiple experiments to zero in on the most likely locations of genes responsible for driving water-use efficiency in S. viridis.

Untargeted metabolomics enables direct quantification of metabolites without apriori knowledge of their identity. Liquid chromatography mass spectrometry (LC-MS), a popular method to implement untargeted metabolomics, identifies metabolites via combined mass/charge (m/z) and retention time as mass features. Improvements in the sensitivity of mass spectrometers has increased the complexity of data produced, leading to computational obstacles. One outstanding challenge is calling metabolite mass feature peaks rapidly and accurately in large LC-MS datasets (dozens to thousands of samples) in the presence of measurement and other noise. While existing algorithms are useful, they have limitations that become pronounced at scale and lead to false positive metabolite predictions as well as signal dropouts. To overcome some of these shortcomings, biochemists have developed hybrid computational and carbon labeling techniques, such as credentialing. Credentialing can validate metabolite signals, but is laborious and its applicability is limited. We have developed a suite of three computational tools to overcome the challenges of unreliable algorithms and inefficient validation protocols: isolock, autoCredential and anovAlign. Isolock uses isopairs, or metabolite-istopologue pairs, to calculate and correct for mass drift noise across LC-MS runs. AutoCredential leverages statistical features of LC-MS data to amplify naturally present 13C isotopologues and validate metabolites through isopairs. This obviates the need to artificially introduce carbon labeling. anovAlign, an anova-derived algorithm, is used to align retention time windows across samples to accurately delineate retention time windows for mass features. Using a large published clinical dataset as well as a plant dataset with biological replicates across time, genotype and treatment, we demonstrate that this suite of tools is more sensitive and reproducible than both an open source metabolomics pipelines, XCMS, and the commercial software progenesis QI. This software suite opens a new era for enhanced accuracy and increased throughput for untargeted metabolomics.