Mechanism Of Action And Variation Mining Of ZmCASP4 In Root Development And Mineral Element Absorption Of Maize

As an important grain,warp and feed crop,maize is the crop with the highest total yield and plays an important role in national production.It is an urgent problem to identify the key genes regulating maize growth and development and improve maize yield.It is very important for plants to get the nutrients they need from the changing environment.The endodermis of plant root system is an important barrier for material exchange between plants and the external environment due to the existence of casparian strip.On the one hand,the existence of the casparian strip can prevent water and nutrients from directly entering the stele through the apoplastic loading pathway,making the symplast pathway the essential route of material transport,so as to achieve the selective absorption of nutrients by plants;on the other hand,the casparian strip can prevent the backflow of nutrients in the stele and improve the efficiency of nutrient absorption by plants.The CASP(CASPARIAN STRIP DOMAIN PROTEINs)protein family has been shown to influence plant growth and development by mediating proper localization of the casparian strip.Although the physiological function of casparian strip is clear,its formation mechanism and the role in regulating nutrient absorption and plant growth remain unclear.In this study,the genetic mechanism of ZmCASP4 regulating root development and mineral element absorption was analyzed by mutant and overexpression lines of ZmCASP4,a key gene for the formation of casparian strip.The main results are as follows:1.Single cell sequencing was performed on the primary root of maize,and 11 main root cell types were classified.It was found that ZmCASP4 was specifically expressed in the endodermis cluster.Pseudo-time analysis showed that the cell cluster where the gene was located was mainly in the late stage of development of the endodermis.The transcriptome data of 357 inbred lines were used to analyze the correlation between the expression level of ZmCASP4 and root traits.The expression level of ZmCASP4 was significantly correlated with total length of primary root,lateral root length of primary root,total length of seminal root,lateral root length of seminal root,total root length and lateral root length of root.Gene expression pattern analysis found that ZmCASP4 is expressed specifically in the root system;it was mainly expressed in 48 to 76 h after;the response to salt stress was the highest.2.ZmCASP4 mutant and overexpression lines were constructed and were the root length was evaluated under hydroponics.Compared with the wild type,the growth of primary root length,seminal root length,root length and lateral root length of root of the knockout material was inhibited;while overexpressed lines showed stronger phenotypes in total length of primary root,primary root length,lateral root length of primary root,average length of seminal root,lateral root length of seminal root,total root length,root length and lateral root length of root.The integrity of the Casparian strip in the mutant and the wild type KN5585 was detected by propidium iodide,and the results showed that the Casparian strip was formed later in the knockout mutant.The content of mineral elements in the shoot of the mutant and overexpression lines was determined.The content of N,P,Ca,Na,Mn and Zn increased in the mutant.The contents of N,Ca,Mg and Na were decreased,while the contents of K and Cu were increased in overexpression lines.3.Based on resequencing data of ZmCASP4 gene in CUBIC population and its parents,The ZmCASP4-based candidate gene association analysis identified two SNPs significantly associated with total root length,total length of seminal root and lateral root length of seminal root.Based on the two significant SNPs,the population was divided into three haplotypes,and there were significant differences in total root length,total length of seminal root and lateral root length of seminal root among different haplotypes.