DOI: 10.5683/sp3/ftacof
title: Replication Data for: Improving abiotic stress tolerance in alfalfa through the down-regulation and genome editing-mediated knockout of multiple genes

description: This study sought to provide insight into the mechanisms behind resilience to waterlogging and drought stresses in alfalfa through the assessment of genotypes in which two transcription factors, WUSCHEL-RELATED HOMEOBOX 13-2 (MsWOX13-2) and TELOMERASE ACTIVATOR 1/REGULATOR OF SYMBIOSOME DIFFERENTIATION (MsTAC1/RSD), had been down-regulated, respectively.
In the case of alfalfa MsWOX13-2, the RNAi-mediated down-regulation of MsWOX13-2 in alfalfa had no effects on growth or morphological characteristics under control (well-watered) conditions. However, under waterlogged conditions, MsWOX13-2 RNAi plants exhibited enhanced performance, as evidenced by a reduced impact of stress on morphology and greater survivability compared to empty vector genotypes. In addition, MsWOX13-2 RNAi genotypes exhibited an apparent reduction in leaf chlorosis under waterlogging, which correlated with higher chlorophyll retention and maximum quantum efficiency of photosystem II (Fv/Fm), compared to empty vector genotypes. RNA-Seq analysis confirmed the presence of differentially expressed genes (DEGs) related to photosynthesis, antioxidant activities, anaerobic respiration, cell wall modulation, phytohormone-related pathways, and transcription factors. On the other hand, MsTAC1/RSD RNAi genotypes exhibited improvements in their resilience to water-deficit compared to empty vector control genotypes as evidenced by higher plant height and dry root biomass following drought recovery. This tolerance to drought could be attributed, at least in part, to a reduction in stomatal density, and a consequent decrease in leaf water loss, along with improved baseline total antioxidant capacity and higher superoxide dismutase (SOD) and catalase (CAT) activity under drought stress. The transcript profiling via RNA-Seq demonstrated changes in the expression of genes related to photosynthesis, antioxidant activities, cell wall modulation, phytohormones, and transcription factors, which could have contributed to the observed improvements in MsTAC1/RSD RNAi genotypes under drought stress. Taken together, these results indicate that MsWOX13-2 and MsTAC1/RSD function as negative regulators of waterlogging and drought stress response in alfalfa, respectively. 

The files listed here contains the differentially expressed genes obtained via RNA-Sequencing conducted on alfalfa genotypes under waterlogging and drought stress conditions. The description of files is as follows: 

Supplemental File 1: List of DEGs between N4.4.2 wild-type and MsWOX13-2 RNAi genotypes grown under normally-watered and waterlogged conditions, and between normally-watered and waterlogged conditions in MsWOX13-2 RNAi and wild-type genotypes. 
Supplemental File 2: Information regarding DEGs between N4.4.2 wild-type and MsWOX13-2 RNAi genotypes under waterlogging stress falling into a selection of MapMan categories with putative functions in abiotic stress response. 
Supplemental File : List of DEGs between N4.4.2 wild-type and MsTAC1/RSD RNAi genotypes grown under well-watered and drought conditions, and between well-watered and drought conditions in MsTAC1/RSD RNAi and wild-type genotypes. 
Supplemental File 4: Information regarding DEGs between N4.4.2 wild-type and MsTAC1/RSD RNAi genotypes under well-watered, and drought stress conditions falling into a selection of MapMan categories with putative functions in abiotic stress response.