Cloning And Functional Characterization Of ZmMCM9,A Endoreplication-Related Gene In Maize

Maize(Zea mays L.)is one of the important food crops and industrial raw material crops in our country and the world.Its safety production plays an important role in national economic and social development.More than 70% of dry matter in maize grains comes from endosperm,so the development and formation of endosperm is directly related to maize yield.Endoreplication refers to cell cycle in which only DNA replicates but cells do not diviede,resulting in multiple ploidy co-existing in the same cell.Endosperm cells of maize have endoreplication.During endoreplication,the volume of endosperm cells increases and nutrients such as starch accumulate continuously,while the time cycle of endosperm filling is almost synchronous with that of endosperm replication.Therefore,endoreplicationmay be the key driving force of maize grain morph and yield formation,and mini-chromosome maintenance(MCM)protein family is closely related to endoreplication mechanism.Therefore,the study of MCM protein family related genes is expected to provide a basis for enriching the theoretical basis of endoreplication and improving maize yield by regulating the occurrence time and duration of endoreplication.In this study,based on previous studies,bioinformatics analysis was conducted on the MCM family,and ZmMCM9 gene was cloned,and the function of the gene was further explored.The main results were as follows:1.Through screening the maize whole genome database,it was found that the ZmMCM family consisted of 21 genes,which were named ZmMCM1-21 according to the distribution order.Physical and chemical properties of the 21 members were analyzed and subcellular localization prediction was made.The molecular weight and isoelectric point are different.These 21 genes were distributed in 9 branches,respectively corresponding to MCM2-10 protein,containing a variety of the same motif and a variety of cis-acting elements on the promoter,indicating that ZmMCMs are highly conserved and have diverse functions.2.The expression pattern analysis of ZmMCM9 showed that ZmMCM9 was mainly expressed in maize leaves,endosperm,and filaments with tissue specificity.The expression level of ZmMCM9 was similar to that of the endosperm endorplication,and it could respond to abiotic stresses such as ABA and PEG.3.The overexpression vector of ZmMCM9 was constructed and transformed into Arabidopsis Thaliana by Agrobacterium inflorescence.It was found that overexpression of ZmMCM9 could affect the growth and reproductive development of Arabidopsis thaliana.Flow cytometry showed that the endogenous replication level of Arabidopsis leaves was significantly improved,indicating that ZmMCM9 was involved in the regulation of endoreplication.4.Subcellular localization results showed that ZmMCM9 was localized in the nucleus.5.The results of yeast diheterozyme showed that ZmMCM9 interacts with Zm CDKA1,Zm CDKB in pairs,indicating that ZmMCM9 may bind with Zm CDKA1 and Zm CDKB to form specific protein-dependent kinase complexes,which can regulate the cell cycle and jointly affect the process of endoreplication.