Analysis of differentially expressed genes and related pathways in space-mutated alfalfa materials under drought stress

CHEN Zhilong; SHANG Jihong; SHA Xiaodi; MAO Yanni; ZENG Yanxia

doi:10.13557/j.cnki.issn1002-2813.2026.02.013

Abstract：This experiment aimed to analyze the differentially expressed genes(DEGs)and related pathways in space-mutated alfalfa materials under drought stress.Using both space-mutated and non-mutated materials as the research subjects,drought stress was simulated with polyethylene glycol 6000(PEG-6000),while distilled water treatment served as the control group.Four treatments were established:Non-mutated material+distilled water(CK1),non-mutated material+drought stress(T1),mutated material+distilled water(CK2),and mutated material+drought stress(T2).Each treatment had three replicates,with 20 seedlings per replicate.After treatment,transcriptome sequencing was conducted to analyze DEGs,and real-time quantitative PCR(RT-qPCR)was used for validation.The results showed that for the Juneng 7,DS310FY,and Adina,the total numbers of DEGs in the CK1-VS-T1,CK2-VS-T2,AND T1-VS-T2 comparison groups were 1 373,1 050,and 259,1 654,1 649,and 1 177,and 1 077,951,and 599,respectively.Compared to their respective controls,under drought stress,Juneng 7 had a higher number of up-regulated genes,while DS310FY had a higher number of down-regulated genes.For Adina,the non-mutated material showed slightly more up-regulated genes,whereas the mutated material showed slightly more down-regulated genes.Compared to the T1 treatment,the T2 treatment showed more down-regulated genes in Juneng 7 and Adina,but more up-regulated genes in DS310FY.Gene ontology(GO)enrichment analysis revealed that the DEGs were enriched in the biological processes of metabolic process,cellular process,and single-organism process,the molecular functions of binding and catalytic activity,and the cellular components of cell part,cell,and membrane.Kyoto encyclopedia of genes and genomes(KEGG)enrichment analysis showed that the DEGs were enriched in pathways such as plant hormone signal transduction,biosynthesis of secondary metabolites,and the mitogen-activated protein kinase(MAPK)signaling pathway-plant.Among the six genes verified by RT-qPCR,the expression trends of five genes were consistent with the transcriptome data.The study shows that after space mutation,Juneng 7 may enhance its drought tolerance by suppressing the up-regulation of certain genes,while DS310FY shows the opposite pattern.The reduced drought tolerance in Adina may be due to the down-regulation of related genes.Plant hormone signal transduction,biosynthesis of secondary metabolites,and the MAPK signaling pathway-plant are likely key pathways in their response to drought stress.