Genetic Control of Lemna Growth Rate and Protein Content

Duckweed (Lemna spp.) has emerged as a promising sustainable protein source due to its rapid growth and adaptability to diverse environmental conditions. This thesis investigated the effects of nitrogen availability and temperature on duckweed growth, protein accumulation, and the underlying physiological and molecular mechanisms, aiming to optimize its use for human and animal consumption. The study began with the establishment of a Lemna collection comprising 50 clones sourced globally. A novel method to quantify total nitrogen and nitrate using FT-IR spectroscopy was developed and applied throughout the research. The first experimental phase (Chapter 3) examined how different nitrogen sources—nitrate, Ammonium-Nitrate, Urea-Nitrate, and nitrogen-free—affected growth rate, protein content, and nitrate accumulation. Ammonium treatment significantly reduced growth in some clones due to pH acidification, though clone 7796 maintained higher growth rates under ammonium and Urea-Nitrate treatments. This clone also exhibited the highest protein accumulation across all nitrogen treatments. Expression analysis of eight nitrogen assimilation genes (NR, NiR, GS1;1, GS1;2, GS2, CLCa, Fd-GOGAT, and NADH-GOGAT) revealed distinct regulation patterns depending on nitrogen source and clone, underscoring the importance of selecting appropriate nitrogen sources to optimize protein yield. The second experimental phase (Chapter 4) investigated heat stress tolerance in 42 Lemna clones (36 L. gibba and 6 L. minor) collected from diverse geographic regions. Physiological assessments at 20°C and 35°C identified three heat-tolerant clones (6861, 7763, and 7796) and one heat-sensitive clone (8703), with the widely used clone Manor serving as a control. Further testing across a broader temperature range (15°C–35°C) revealed that while all clones exhibited reduced growth at higher temperatures, protein content increased in heat-tolerant clones but declined in the heat-sensitive clone at 35°C. In the final experimental phase (Chapter 5), transcriptomic analysis provided insights into the molecular mechanisms underlying heat tolerance in the selected five clones. Differential gene expression analyses revealed upregulation of genes involved in photosynthesis (e.g., ATP synthase), zinc ion binding, and stress response pathways in heat-tolerant clones, while these genes were downregulated in the heat-sensitive clone and the control. KEGG and GO pathway enrichment analyses highlighted critical metabolic and regulatory pathways associated with heat resilience. Together, these findings demonstrate that nitrogen source selection influences duckweed growth and protein accumulation, with clone-specific responses to ammonium availability. Heat-tolerant clones maintain higher protein levels under elevated temperatures, and their transcriptomic profiles suggest a genetic basis for resilience to heat stress. These results provide valuable insights into optimizing duckweed cultivation under variable environmental conditions, supporting its potential as a sustainable protein source in the context of climate change.