Identification Of Maize HSP70 Family Members And Screening And Functional Analysis Of Drought Resistance Genes

As one of the three major grain crops,the growth of maize is seriously affected by drought stress.Heat shock protein 70(HSP70)is a highly conserved molecular chaperone that is activated when the organism is subjected to stress.It has been found in previous studies that HSP70 plays an important role in plants responding to stress.However,little is known about the function of maize HSP70.In this study,the maize genomic data were analyzed to identify the HSP70 family members,and the family members were further analyzed.The HSP70 family members involved in maize drought resistance were screened out,and then some members' preliminary function were investigated.Firstly,41 maize HSP70 family members were identified using bioinformatics methods.The phylogenetic tree was generated with Arabidopsis and rice HSP70 family members.The results showed that the clusters of ZmHSP70 in the phylogenetic tree were in consistent with two subfamilies of HSP70.Among them,25 members belonged to the DnaK subfamily and 16 members belonged to the HSP110/SSE subfamily.The subcellular localization prediction results showed that23 of the 41 maize HSP70 members were located in the cytoplasm,8 ZmHSP70 s located in the endoplasmic reticulum,8 ZmHSP70 s located in the chloroplast,and 2ZmHSP70 s located in the mitochondria.By analyzing the protein structure of maize HSP70 members,it was found that each member contained a typical conserved motif of HSP70,and this result validated the accuracy of the identified family members.Secondly,candidate genes responding to drought stress were screened by analysis of gene promoter region stress and hormone-related elements.Further,under drought stress,semi-quantitative PCR and real-time quantitative PCR were used to detect the maize candidate gene's expression.The results showed that the expression of ZmHSP70-1,ZmHSP70-2.1,ZmHSP70-8 enhanced significantly,which suggesting these members maight be involved in the response to signaling drought stresse..Thirdly,tobacco was transiently transformed to observe the localization of ZmHSP70-1,ZmHSP70-2.1,and ZmHSP70-8 in the cells.Laser confocal microscopy observations showed that all three members were located in the cytoplasm,which was consistent with bioinformatics predictions and consistent with the related localization motifs contained in the protein sequence.Through transformation ZmHSP70 s to Arabidopsis thaliana,ZmHSP70-1 and ZmHSP70-8 overexpression lines were constructed.The results of soil drought showed that both overexpression lines grow better than wild type and mutants under drought stress.The transplanted seedlings under mannitol treatment also showed that overexpression plants were better than wild type and mutants,indicating that ZmHSP70-1 and ZmHSP70-8 genes were involved in the drought stress response in the seedling period.The germination experiments under salt treatment showed that ZmHSP70-1 and ZmHSP70-8 overexpression lines showed salt-tolerant phenotype,and the germination rate was significantly higher than that of wild type and mutants.In summary,the maize HSP70 family was a multi-gene family.ZmHSP70-1 and ZmHSP70-8 played a positive regulatory role in response to drought stress in maize.This study has a certain significance for the screening of drought-resistance related genes in maize,which was helpful for further research on the functional mechanisms of maize drought-resistant genes.