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.
Highlights
Alfalfa is an important forage crop that is moderately tolerant to salinity; little is known about its salt-tolerance mechanisms
In order to gain a general insight into gene expression profiles of two alfalfa genotypes (G03, salt-tolerant and G09, salt-sensitive) for roots (R) and leaves (L) under salt stress, 24 cDNA libraries were designed for RNA sequencing, representing control treatments (C) as C03L, C03R, C09L, and C09R and salt treatments (T) as T03L, T03R, T09L, and T09R in three replicates (Supplementary Table S1)
For the functional annotation of all unigenes, searches against seven public databases showed a significant similarity of unigenes with the sequences in these databases ranging from 65,987 (15.77% in KOG to 327,517 (78.26% in NT (NCBI non-redundant nucleotide sequences) (Supplementary Fig. S3)
Summary
Alfalfa is an important forage crop that is moderately tolerant to salinity; little is known about its salt-tolerance mechanisms. Some basic understanding has been established on various traits and processes involved in salinity tolerance (such as osmotic stress, ion exclusion, and tissue tolerance) in various crop plants[2,3]. Alfalfa is the most widely cultivated perennial forage legume in the world due to its high protein content and palatability to livestock It is cultivated on more than 23 million acres in the United States a lone[7]. Compared to other legume crops, alfalfa is moderately tolerant to salt s tress[2,5]; salinity stress is an important factor that is responsible for the lower yield of alfalfa. A better understanding of the mechanisms of salt-tolerance in alfalfa can be generated by employing cutting-edge technologies such as transcriptomics[13,14] and proteomics[15]
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