Evaluation Of Maize(Zea May L.) Cold Tolerance At Seedling Stage,and Functional And Evolutionary Analyses Of Crops COLD1 Proteins

As one of the staple crops,maize has the following characteristics: high yield,high nutritional value,and is widely used in food,feed,fuel and other industries.Maize originated from tropical and subtropical regions belonging to thermophilic plants,therefore,low temperature has always been a critical factor limiting its growth and distribution.As the primary corn producing area,low temperature has become one of the dominating limiting factors of corn production in northeast China.The seedling stage is a pivotal period for maize growth and development,and maize is very sensitive to low temperature at this stage.All in all,digging deeper excellent resistant materials with strong cold tolerance and cold-regulated genes is of particular importance.1.Evaluation of cold tolerance at seedling stage was conducted on 205 corn germplasm resources with rich blood relationship,and the physiological and morphological differences of 205 maize seedlings were identified before and after cold treatment with 4 ° C for 5 days.Depend on the aforesaid measured results,a comprehensive evaluation on the basis of factor analysis were carried out,and the results indicated that all materials were divided into 5 groups,among which maize inbred line By813 showed extreme cold tolerance and maize inbred line CML454 showed extreme cold sensitivity.2.The differences on phenotypes and cell structures of cold resistant maize By813 and cold sensitive maize CML454 were further confirmed before and after cold treatment,and the results revealed that the differences are obvious.Transcriptome analysis on these samples was conducted,and KEGG analysis of differentially expressed genes(DEGs)was carried out.Results showed that the flavonoids biosynthesis pathway and plant hormone signal transduction pathway might be the candidate targets.3.G protein signal pathway is a vital aspect of plant signal perception and transmission,but the roles of plant GPCRs in the response to low temperature are short of study.In this study,three novel GTG/COLD1 genes,designed as Zm COLD1-10 A,Zm COLD1-10 B and Zm COLD1-10 C,were obtained from maize,and detailed researches on gene function were explored.The detailed results are as follows:(1)Three Zm COLD1-10 proteins possess the basic characteristics of GTG/COLD1 proteins,including GPCR-like topological structure,hydrophilic loop,DUF3735 domain,GTP hydrolases activate domain,ATP/GTP binding domain.In addition,the GTG/COLD1 proteins are evolutionarily conserved.(2)Zm COLD1-10 A was expressed in root,stem and leaf of maize,and was induced by cold while was inhibited by light signal.Arabidopsis cold1 mutants showed sensitive to cold that WT,and Arabidopsis Zm COLD1-10A-OE plants showed enhanced cold tolerance.Four cis-elements in the promoter of Zm COLD1-10 A were identified as the candidate elements,such as H-box,MYB,ABA and DRE core regulated elements.(3)Zm COLD1-10 protein located in plasma membrane(PM)and endoplasmic reticulum(ER),is the membrane-regulated protein.Analyses through Bi FC showed that Zm COLD1-10 A interacts with Zm CT2,and the core domain of interaction is the HL domain of Zm COLD1-10 A and the C-terminal of Zm CT2.4.Anslyses of function and evolution of cold related proteins in crops revealed that the protein structure and mechanisms of COLD1 proteins are conserved;the structure of ICE and CBF proteins are conserved,but the structure of COR proteins are not conserved in the ICE-CBF-COR pathway;IRI proteins are conserved only in grains,instead of all the crops.Hence,COLD1 protein is the the optimal targeted protein to improve crops cold tolerance through the ways of genetic engineering.All in all,this study provides theoretical basis and important reference for digging excellent resistant materials and resistant genes,as well as improving maize cold tolerance.