Biological And Molecular Investigation Of The Halophyte Suaeda Glauca:Micropropagation,Salinity Tolerance,Photoperiodism,and Stress-Associated Proteins

Crops are a vital source of sustenance for human beings;however,in recent years,they have faced severe threats due to the impact of human activities,natural disasters,and climate change,leading to a continuous reduction in arable land and crop yields.Of particular concern is the exacerbation of soil salinization,which has become one of the primary factors jeopardizing food security.China,as a country facing relatively severe salinity and alkalinity issues,is dealing with soil salinization,while Afghanistan faces even more severe soil salinity problems.Most crops are"glycophytes,"struggling to grow in high-salinity soils.In contrast,the Chenopodiaceae family,particularly the salt-tolerant plant S.glauca,can withstand salt concentrations as high as 600 m M Na Cl,thanks to its unique abilities in salt uptake and storage.A profound exploration of the salt tolerance mechanisms in S.glauca holds the potential to provide essential guidance for crop genetic improvement and serve as a valuable resource of salt tolerance genes for crop breeding.Considering the applicant’s home country’s situation and the research focus of the laboratory where they work,the applicant is dedicated to in-depth research on salt-tolerant S.glauca.The applicant has pursued four main research directions:1.An efficient regeneration system is a necessary prerequisite for conducting functional genomics research.However,for a long time,an efficient regeneration system for S.glauca,a salt-tolerant plant,remained elusive.The primary task of the applicant was to overcome this challenge.Through exploration and optimization of the S.glauca regeneration system,the applicant successfully established a regeneration system based on an organogenesis program primarily using the plant hormones 6-BA and IBA.Additionally,considering the current difficulty in genetic transformation of S.glauca,the applicant developed a comprehensive protocol for the isolation and extraction of S.glauca protoplasts,along with a polyethylene glycol-mediated genetic transformation system.These established techniques lay a crucial technological foundation for developing S.glauca as a model salt-tolerant plant.2.S.glauca,as a highly salt-tolerant plant,exhibits significant adaptability in its growth,development,and physiology when exposed to high salt concentrations.To gain a deeper understanding of its salt tolerance mechanisms,the applicant conducted research focused on salt stress.The findings are as follows:（1）Under moderate to low salt concentrations（Na Cl）,S.glauca’s growth and accumulation of photosynthetic pigments were promoted,even though high salt concentrations inhibited branching and chlorophyll content,still promoting biomass accumulation.（2）Low salt stress enhanced the values of Fo and Fm,and salt treatment further increased these values at low salt concentrations,with no significant changes in the maximum quantum efficiency of PSII（Fv/Fm）and quantum yield of PSII（φPSII）.（3）Low salt concentrations led to an increase in the photochemical quenching（q P）and a decrease in non-photochemical quenching（NPQ）,which were reversed under moderate to high salt concentrations.These results indicate that even under high salt concentrations（400 m M Na Cl）,S.glauca still exhibits notable salt tolerance.The growth parameters,photosynthetic pigments,and chlorophyll fluorescence data for PSII under high salt stress can be considered fundamental information for cultivating S.glauca in saline-alkali soils.3.Photosynthesis is one of the most crucial life processes in plants.However,to date,there has been insufficient molecular biology evidence regarding the photosynthetic type of S.glauca.To bridge this knowledge gap,the applicant adopted a sampling strategy,collecting samples every two hours throughout a 24-hour period,to conduct comprehensive research on S.glauca.RNA-seq technology was employed to analyze gene expression changes at the genome level.Through this research,the applicant identified rhythmic genes in the S.glauca genome and further determined the photosynthetic type of S.glauca based on the expression patterns of these genes.The research findings are as follows:（1）Successful identification of1676 rhythmic genes in S.glauca,with in-depth analysis of their expression and regulation.（2）Clear identification of the core circadian clock genes in S.glauca,along with the revelation of their regulatory pathways.（3）Analysis of genes involved in light absorption,light reactions,and dark reactions in S.glauca,followed by periodic expression analysis,ultimately determining S.glauca’s photosynthetic type as a C₄plant.These research results provide a solid foundation for gaining a deeper understanding of how this regulatory system influences the agronomic traits of S.glauca and other C₄plants.4.An important task of the research team where the applicant is affiliated is the identification and isolation of salt-tolerant gene resources in S.glauca.In this field,the applicant conducted genomic identification,analysis,and functional validation of genes related to Stress-Associated Proteins（SAPs）in S.glauca.The primary research progress includes:（1）Identification of 15 SAP genes in S.glauca and a detailed analysis of their gene structures,protein motifs,and expression characteristics;（2）Sequence alignment,conservation domains,and genomic structure analysis revealed the high conservation of S.glauca SAPs,which encode A20/AN1 zinc finger proteins;（3）Transcriptome heat map analysis showed significant upregulation of most S.glauca SAP genes under salt stress;（4）Genes that were highly expressed under salt stress conditions were cloned and transferred to the model plant Arabidopsis thaliana,where functional analysis of three candidate genes was performed.The results indicated that transgenic plant lines Sg SAP2 and Sg SAP4 exhibited high expression and enhanced tolerance to salt stress.This research serves as a demonstration and reference for the exploration of salt-tolerant genes in S.glauca through gene family and gene expression analysis.In summary,the research conducted by the doctoral degree applicant deepens our understanding of the salt-tolerant plant S.glauca.It elucidates S.glauca’s response mechanisms to salt stress,reveals its photosynthetic type at the molecular level,and successfully uncovers three potential salt-tolerant candidate genes.This study establishes a solid foundation for developing S.glauca as a model plant for salt-affected soils.Furthermore,the knowledge and techniques possessed by the applicant in this research will soon contribute to the advancement of agriculture in Afghanistan.