De novo Propagation of Alfalfa Under Hydroponics

Abstract Background: The legume plant alfalfa (Medicago sativa L.) is a widely cultivated perennial forage due to its high protein levels, palatability, and strong adaptability to diverse soil types and agro-ecological zones. This forage plant is a self-incompatible, cross-pollinated autotetraploid with tetrasomic inheritance. Therefore, maintaining excellent traits through seed reproduction is challenging in alfalfa. However, the cutting propagation technology could enable consistent multiplication of quality plants that are genetically identical to the parent plant, for use in breeding and other research applications. Most of previous alfalfa omics researches used varieties as material on omics and gene mining experiment due to poor growth consistency of cuttings by existing cutting methods, which generate genetically un-identical cuttings and thus compromise on the reliability of the results. Therefore, this study aimed to develop a simple, cost-effective, reproducible, and efficient hydroponic cutting method for the preservation of alfalfa plants and molecular research applications such as genomic, transcriptomic, and proteomic analyses. Results: Alfalfa cultivar ‘Wudi’ grown under hydroponics for 30 days was used as source material for cuttings. The top, middle and bottom sections of its stem were used as cuttings. The rooting rate, root length, and stem height of the different stem sections were compared to determine the best segment for alfalfa propagation in four nutrient solutions (HM, HM+1/500H, HM+1/1000H and d HM+1/2000H). After 21 days of culture, the rooting rates of all the three stem types under four cutting nutrient solutions were above 78%, The rooting rate of the middle and bottom parts in HM +1/1000 H and HM +1/2000 H nutrient solutions reached more than 93% with higher health survey score (>4.70). Besides, root length and stem height in these two sections was exemplary. Conclusions: This study developed a de novo cutting propagation method that can be used to conserve and propagate germplasm in breeding programs and research. This article is the first report on the cutting propagation of alfalfa by hydroponics, which could supplement the existing cutting propagation methods.