生物技术通报2025，Vol.41Issue(6)：297-306,10.DOI:10.13560/j.cnki.biotech.bull.1985.2024-1143
家榆UpNOA1的克隆及其互作蛋白的筛选
Cloning of UpNOA1 Gene from Ulmus pumila L.and Screening of Its Interacting Proteins
摘要
Abstract
[Objective]Nitric oxide-related protein NOA1 is associated to the levels of NO in plants,which may be involved in various physiological activities of plants.The UpNOA1 gene was cloned,its tissue-specific expression in elm(Ulmus pumila L.)was analyzed,and the proteins interacting with UpNOA1 was screened,which may provide the basis for studying the function of UpNOA1 in elm.[Method]Ulmus pumila L.was used as the experimental material,and the NO-related UpNOA1 gene was cloned.The domain architecture and phylogenetic relationships were analyzed using bioinformatics approaches.Tissue-specific expression of UpNOA1 was assessed via RT-qPCR.A vector was constructed for prokaryotic expression analysis,and the resulting UpNOA1 protein was purified.Finally,the UpNOA1 interaction proteins were screened and analyzed by His-PULL DOWN.[Result]The UpNOA1 gene possesses a 1 698 bp open reading frame,encoding 565 amino acids and containing a conserved GTP/Mg2+binding sites.Subcellular localization prediction using Plant-mPLoc indicated that the UpNOA1 protein is targeted to chloroplasts.Phylogenetic analysis revealed that the UpNOA1 protein shows the highest sequence homology to UaNOA1 protein from American elm(Ulmus americana)and clusters with it in the phylogenetic tree.RT-qPCR analysis revealed that the expression of UpNOA1 gene is the highest in the leaves,followed by the seeds.A total of 131 proteins were identified as UpNOA1-interacting proteins via His-PULL DOWN assay.KEGG and GO enrichment analyses indicated a predominance of ribosomal proteins is associated with genetic information processing among the interacting proteins.Following annotation and comparison,a list of 25 candidate proteins potentially involved in stress resistance and GTP binding was compiled.[Conclusion]UpNOA1 gene is cloned.The highest expression is found in leaves,followed by elm seeds.A total of 131 UpNOA1 interacting proteins are identified,of which 25 are associated with stress resistance and GTP binding.关键词
家榆/NOA1/组织特异性表达/PULL DOWN/互作蛋白Key words
Ulmus pumila L./NOA1/tissue specific expression/PULL DOWN/interaction protein引用本文复制引用
刘晓天,何毓琦,刘畅,薛华..家榆UpNOA1的克隆及其互作蛋白的筛选[J].生物技术通报,2025,41(6):297-306,10.基金项目
北京林业大学"5·5工程"科研创新团队项目(BLRC2023C06),国家自然科学基金面上项目(31971646) （BLRC2023C06）
