Transcriptional profiling of two contrasting genotypes uncovers molecular mechanisms underlying salt tolerance in alfalfa

Authors: KAUNDAL, RAKESH - Utah State University; DUHAN, NAVEEN - Utah State University; ACHARYA, BISWA - University Of California; Pudussery, Manju; Ferreira, Jorge; Suarez, Donald; Sandhu, Devinder

Submitted to: Scientific Reports
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 2/12/2021
Publication Date: 3/4/2021
Citation: Kaundal, R., Duhan, N., Acharya, B.R., Pudussery, M.V., Ferreira, J.F., Suarez, D.L., Sandhu, D. 2021. Transcriptional profiling of two contrasting genotypes uncovers molecular mechanisms underlying salt tolerance in alfalfa. Scientific Reports. 11. Article 5210. https://doi.org/10.1038/s41598-021-84461-w.
DOI: https://doi.org/10.1038/s41598-021-84461-w

Interpretive Summary: Salinity is one of the major stresses encountered by plants in nature that reduces crop productivity worldwide. Currently, there is little understanding about how plants, including alfalfa change their physiological status to cope with soil salinity. Thus, there is a need for a comprehensive study of alfalfa response to salinity to help in identifying and generating new salt-tolerant lines that can be used in breeding programs. Alfalfa is the most widely cultivated perennial forage legume in the world due to its high protein content and palatability to livestock. Better understanding of the mechanisms of salt-tolerance in alfalfa can be generated by employing new cutting-edge technologies. In this study, we explored the differential gene expression between a salt-tolerant and a salt-sensitive alfalfa genotype at maturity stage by (gene expression (transcriptional) profiling of root and leaf tissues of plants kept under long-term irrigation with either high- or low-salinity water treatments. RNA of 24 alfalfa samples was sequenced and predicted functions of alfalfa genes were determined. Several putative genes that regulate salinity tolerance in alfalfa were identified, which included genes involved in important pathways such as hormonal signaling, calcium signaling, redox signaling, and transcriptional regulation. Suitable candidate genes can be further explored for their role in salinity-tolerance of alfalfa and used for the breeding of new salt-tolerant varieties. Hence, our results will help alfalfa breeders and geneticists to develop new salt-tolerant alfalfa varieties, which in turn would allow farmers to increase crop yield in marginal lands, and to use cheaper recycled waters.

Technical Abstract: Alfalfa is an important forage crop that is moderately tolerant to salinity; however, little is known about its salt-tolerance mechanisms. We studied root and leaf transcriptomes of a salt-tolerant (G03) and a salt-sensitive (G09) genotype, irrigated with waters of low and high salinities. RNA sequencing led to 1.73 billion high-quality reads that were assembled into 418,480 unigenes; 35% of which were assigned to 57 Gene Ontology annotations. The unigenes were assigned to pathway databases for understanding high-level functions. The comparison of two genotypes suggested that the low salt tolerance index for transpiration rate and stomatal conductance of G03 compared to G09 may be due to its reduced salt uptake under salinity. The differences in shoot biomass between the salt-tolerant and salt-sensitive lines were explained by their differential expressions of genes regulating shoot number. Differentially expressed genes involved in hormone-, calcium-, and redox-signaling, showed treatment- and genotype-specific differences and led to the identification of various candidate genes involved in salinity stress, which can be investigated further to improve salinity tolerance in alfalfa. Validation of RNA-seq results using qRT-PCR displayed a high level of consistency between the two experiments. This study provides valuable insight into the molecular mechanisms regulating salt tolerance in alfalfa.