Improved forage quality and biomass yield of alfalfa (Medicago sativa L.) by Arabidopsis QQS orphan gene

Improving the forage quality of alfalfa in terms of digestibility and crude content is essential for any alfalfa quality breeding programs. Arabidopsis thaliana orphan gene QQS (Qua-Quine Starch) has been shown to improve protein content and alter carbohydrate composition in different food crops. However, there are significant differences in agronomic traits and nutritional conditions between alfalfa and other food crops. To explore the biological function and molecular mechanisms of QQS in alfalfa, we generated QQS transgenic plants and their segregated population (T1 generation), and evaluated their performance under normal- and nitrogen-deficient conditions. Our findings indicate that QQS can significantly enhance the total nitrogen and crude protein content of alfalfa and increase nodule weight under low-nitrogen conditions. Furthermore, QQS transgenic lines also showed reduced levels of neutral detergent fiber (NDF) and lignin, improving forage digestibility. By RNA sequencing and RT-qPCR analysis, we found that QQS affected the expression of genes involved in carbon and nitrogen metabolism, lignin biosynthesis and amino acid biosynthesis and degradation pathways in alfalfa. In addition, QQS also improved alfalfa biomass yield by increasing branch number and plant height in both greenhouse and field conditions. Our results demonstrate that QQS as a useful molecular tool can improve alfalfa biomass yield and overall forage quality and could have significant implications for the alfalfa breeding industry in satisfying the constant demands for high-quality and high-yielding forage.