草业学报2025，Vol.34Issue(5)：89-104,16.DOI:10.11686/cyxb2024247
紫花苜蓿低温诱导蛋白MsLTI65的分离及其对不同逆境的响应
Isolation of the low-temperature induced protein MsLTI65 from alfalfa and its response to different stresses
摘要
Abstract
Low temperature-inducible(LTI)proteins are a class of stress-responsive proteins in plants that play a crucial role in response to abiotic stress.To elucidate the structural characteristics of the MsLTI65 protein in alfalfa(Medicago sativa)and its response to various abiotic stresses,the cDNA sequence of the MsLTI65 gene was successfully cloned from alfalfa cultivar"Zhongmu No.1"using reverse-transcription polymerase chain reaction(RT-PCR)and 3′/5′ rapid-amplification of cDNA ends(RACE PCR)techniques.Bioinformatics tools were employed to analyze the gene sequence and structure,and a phylogenetic tree was constructed to explore the evolutionary relationships between MsLTI65 and LTI proteins from other plants.The expression pattern of MsLTI65 under different abiotic stress conditions was analyzed by quantitative real-time PCR(qRT-PCR)and Western blotting(WB).Sequence analysis indicated that the MsLTI65 gene had a coding region sequence length of 2016 bp,encoding a polypeptide of 671 amino acids with a molecular weight of 74 kDa and a theoretical isoelectric point of 4.55.The MsLTI65 protein showed high homology with the MtLTI65 protein from Medicago truncatula.The qRT-PCR results showed that MsLTI65 expression was induced and upregulated under cold,salt,drought,Cu2+,Zn2+,and abscisic acid(ABA)stresses.The antibody preparation results indicated that a polyclonal antibody against MsLTI65 was successfully generated.This polyclonal antibody exhibited high specificity and was able to recognize native LTI65 samples.Western-blot validation demonstrated that MsLTI65 protein expression is induced by drought,salt,cold and ABA stresses.The results of this study suggest that MsLTI65 functions as a positive regulatory factor in the signal transduction of various abiotic stresses,including cold,drought,Cu2+,Zn2+,salt,and ABA hormone signaling.关键词
紫花苜蓿/MsLTI65/基因克隆/表达分析/抗体/非生物胁迫Key words
Medicago sativa/MsLTI65/gene cloning/expression analysis/antibody/abiotic stresses引用本文复制引用
周昕越,王丽萍,蒋庆雪,马晓冉,仪登霞,王学敏..紫花苜蓿低温诱导蛋白MsLTI65的分离及其对不同逆境的响应[J].草业学报,2025,34(5):89-104,16.基金项目
财政部和农业农村部:国家现代农业产业技术体系(CARS-34),中国农业科学院科技创新工程项目(ASTIP-IAS10)和农业种质资源普查收集、保护鉴定服务(19240485)资助. （CARS-34）
