Functional Verification Of The Salt Stress-responsive Gene ZmSTG1 In Maize

As one of the most important abiotic stresses in agricultural production,soil salinization has become an important adverse factor restricting agricultural development.As an important C4 plant,maize（Zea mays L.）has become an important industrial raw material from a primary forage crop.Therefore,the change of its yield and quality will have an impact on world food security and economic development.However,maize is moderately sensitive to salt,and the germplasm resources of maize salt-tolerant are extremely scarce.Therefore,it is of great importance to excavate maize salt-inducing genes,clarify their regulatory relationship with maize salt-tolerance ability,and explain the mechanism of maize salt-tolerance,so as to cultivate salt-tolerant maize varieties to deal with the yield and quality problems caused by land salinization.In the early stage,our research group constructed a pair of near-isogenic lines with significant differences in salt tolerance.Through transcriptome sequencing after processing them under normal and salt stress conditions,we screened a gene with significant differences in salt stress-induced expression in the two materials,because this gene has not been reported.We tentatively named it maize salt-tolerent gene 1（ZmSTG1）.In this study,salt stress induced expression analysis of ZmSTG1 gene,upstream promoter element prediction analysis,characteristics analysis of its coding protein,plant apparent and physiological characteristics analysis of its overexpression and knockout mutant plants,and at the same time,Transcriptome sequencing was performed on the overexpressed plants,and the downstream genes affected by ZmSTG1 were preliminarily analyzed based on this,and the possible regulatory pathways of ZmSTG1 in maize salt stress response were speculated.The main results are as follows:（1）The expression characteristics of ZmSTG1 gene in two maize inbred lines（082and Ye107）under 200 m M Na Cl stress were evaluated by qRT-PCR.The results showed that in the salt-tolerant maize inbred line 082,the expression of ZmSTG1 gene was strongly induced by salt stress,and the expression was generally up-regulated in the salt-tolerant inbred line 082.（2）The 2000 bp upstream promoter sequence of ZmSTG1 gene was predicted and analyzed,and the results showed that there were 8 ABA-inducible regulatory elements（ABRE）in the promoter region of this gene.In addition,it also includes other plant endogenous hormones（auxin,ethylene,methyl jasmonate）inducible regulatory elements,injury and anaerobic inducible regulatory elements,light-responsive elements and a small amount of meristem expression and protein metabolism regulatory elements.（3）The characteristic analysis of ZmSTG1 protein showed that ZmSTG1 protein did not contain any known protein domain,but had a transmembrane domain and a complex helix region.Its homologous proteins and phylogenetic relationships showed that ZmSTG1 protein was relatively close to monocotyledons and its N-terminal was highly conserved.（4）Overexpression of ZmSTG1 can enhance salt tolerance of maize seedlings.Under different levels of Na Cl stress,compared with the wild type,ZmSTG1 overexpressed maize seedlings were less inhibited,and their aboveground and primary root traits were better.Meanwhile,POD activity increased and H₂O₂ content decreased in overexpressed plants.（5）ZmSTG1 gene knockout enhanced salt sensitivity of transgenic maize seedlings.Without horizontal Na Cl treatment,compared with the wild type,ZmSTG1 knockout mutant was more inhibited in plant growth,and its aboveground and primary rooting traits were worse.At the same time,POD activity was decreased and H₂O₂ content was increased in knockout mutant plants.（6）Transcriptome sequencing results of ZmSTG1 overexpressed plants and wild-type plants showed that the change of ZmSTG1 gene expression level caused 14 DEGs to respond to salt stress.Therefore,ZmSTG1 may play a role by regulating the expression of salt stress responsive genes.（7）Combined with the results of promoter element prediction analysis and transcriptome sequencing,we preliminarily speculated that ZmSTG1 gene may rely on ABA-mediated plant hormone signaling pathway in response to salt stress.