Association Mapping,Transcriptome Analysis And Hormonal Regulation Of Salt Tolerance In Tall Fescue And Bermudagrass

Soil salinity is a serious abiotic factor that constrains plant growth and limits crop productivity in many areas of the globe.Adaptation of plants to environmental stress is a slow process,and since studying soil salinity in tofo is a eomplex process,there i5 a need to focus on the environmental responses in planta.Tall fescue(Festuca arundinacea)and bermudagrass(Cynodon dactylon)are the most important cool and warm season turf grasses,respectively which have promising potential to be used as forage saline soil.In addition,their turf can provide improved land utilization and reduce soil erosion which contributes significantly towards environmental protection,recreation,and aesthetics.However,increased soil salinization constrains their cultivation as well as their productivity.This is critical because the evaluation of different genotypes for the selection and breeding is not yet fully explored.Therefore,in order to facilitate the identification,selection,and breeding,we first evaluated a diverse tall fescue population from different origins across the globe using 99 SSR markers as well as five functional phenotypic traits i.e.,leaf water content,turf quality,evapotranspiration rate,relative growth rate and chlorophyll content.There was significant accession-treatment effect across the population despite similar salt concentration level.The five functional traits showed high diversity and significant mutual correlations in response to salt stress in tall fescue.Two population groups were detected in the panel.Association mapping between the SSRs and the functional physiological traits identified a total of 1024 allele markers most of which had significant associations with the five traits.Some allele markers were associated with more than one trait.Then we ranked the plants based on tolerance level and were able to obtain the most tolerant and the most sensitive accessions based on performance assessment.In principle,association mapping studies not only provide selection and breeding ground for genotypes with superior salt tolerance but also lay a platform for subsequent molecular studies.The absence of tall fescue full genome sequence makes a transcriptome study very resourceful in providing insights into potential molecular responses to salinity stress.Here,we used the identified salt-tolerant tall fescue accession’Puregold’to conduct a comprehensive transcriptome analysis.After assembly,unigenes and their various characteristics were obtained.The unigenes were subsequently aligned to seven databases for functional annotations,which grouped them into various categories including those that may play important roles in salt tolerance.In addition,coding DNA sequences were detected as well as SSRs and SNPs that were distributed among the unigenes.Also predicted were key Transcription Factors,most of which belong to salt-tolerant families.Phytohormones act as crucial molecules that may promote plant growth and crop productivity under stressful environmental conditions.After characterizing different tall fescue accession under salt stress,we also investigated the role of a novel hormone paclobutrazol on tall fescue salt tolerance.We observed that paclobutrazol triggers the upregulation of salt tolerant genes,promotes salt tolerance via improving the leaf water content,osmotic adjustment,leaf water content as well as biomass and photosynthetic capacity.In general,the hormone ameliorated the negative effects of salt stress in tall fescue.We then reviewed and proposed a summarized model for salt tolerance in tall fescue based on our results as well as previous literature.Finally,as the most important warm-season turfgrass,we also analyzed the role of microRNA on bermudagrass salt tolerance.MicroRNAs not only play an important role in various biological processes but also enhance plant resistance to environmental stress.Here,quantification of functional physiological traits revealed that bermudagrass cultivar’43’(’C43’)is more salt-tolerant than cultivar ’ 198’(’C198’)as observed by higher K+/Na+ ratio,higher membrane integrity,as well as higher water use efficiency under salt stress.To understand the post-transcriptional salt stress response,RNA sequencing was used to construct four bermudagrass small RNA libraries i.e.,C43_salt,C198_salt,C43_control,and C198_control.Totally,146 miRNAs were expressed which belonged to 17 conserved families,12 non-conserved,and novel sequences.Also,536 miRNAs were differentially expressed across the four libraries and salt stress-induced downregulation of more miRNAs in ’C43’ than ’C198’.The differentially expressed miRNAs targeted putatively 1891 genes out of which 584 were annotated to various gene ontology terms.Genes annotated to metabolic process,regulation of transcription,DNA template,signal transduction,lipid metabolism,cell cycle,oxidation-reduction process,and protein binding were highly enriched.Also enriched was transcription factor complex under which key salt-responsive and growth-related transcription factors and their cognate miRNAs were downregulated in C43 vs C198 salt regime.The expression patterns of miRNAs and their targets were validated by real-time qPCR which was consistent with sequencing results.Our results provide new insights into the potential salt response regulatory role of miRNAs in the root apex of bermudagrass where cell division and specialization occurs.