Research On The Function Of Key Genes In Oxalate Metabolism In Spinach

Oxalic acid plays an important role in the growth and development of plants.However,excessive accumulation of oxalic acid in the human body can lead to a variety of kidney-related diseases and neurological diseases.Spinach has a high content of oxalic acid,it will be helpful to understand the mechanism of oxalic acid biosynthesis and degradation,due to study the functions of key genes in oxalate metabolism and to explore its oxalate metabolism regulation network in spinach,and it will explore ways to reduce the oxalate content and improve nutritional quality in spinach.In this study,transgenic Arabidopsis of key genes of spinach oxalate metabolism,was used to determine the changes of various physiological indicators under different environmental stresses(high temperature stress,low temperature stress,and salt stress),and to explore the role of spinach oxalate metabolism key genes overexpressed in Arabidopsis in response to different environmental stresses.To analyze the biological function of spinach oxalate decarboxylase gene(SoOXDC),the bioinformatics analysis of spinach OXDC proteins,the root growth of transgenic Arabidopsis under oxalic acid and oxidative stress,as well as the changes of antioxidant enzyme gene expression and activity under oxidative stress were studied to explore the gene function of SoOXDC2 and SoOXDC3.The expression of stress-responsive genes and transcriptome sequencing analysis of the transgenic line OXDC3-5 treated with oxalic acid stress were carried out to further explore the gene function of spinach SoOXDC3.The main conclusions of this study are as follows:1.At 1 h of high temperature stress,the soluble sugar content of the transgenic line OXDC2-10 increased by 1.37 times and the CAT activity increased 5.5 times as compared with the control.The transgenic Arabidopsis responded to high temperature stress by affecting the soluble sugar metabolism and CAT activity.The soluble protein content of OXDC2-10 decreased by 70.2% compared with the control under low temperature stress for 3 h,and the nitrate nitrogen content of OXDC3-5 decreased by44% compared with the control after low temperature treatment for 1h.Transgenic Arabidopsis responds to low temperature stress by affecting the metabolism of nitrate nitrogen and soluble protein.At 1 h of salt stress,the soluble sugar content of OXS-1is 1.19 times higher than that of the control;the ?-carotene content of OXDC3-5 is 55%lower than that of the control;At 12 h of salt stress,the Vc content of OXDC2-10 is higher than that of the control.The control decreased by 74%.Salt stress caused the accumulation of soluble sugar in transgenic Arabidopsis,and decreased the content of?-carotene and Vc.2.The gene structure,conserved motifs and evolutionary relationship of the three SoOXDCs genes identified in the spinach genome were analyzed.Through phylogenetic tree analysis,it is found that the similarity between SoOXDC3 and Bv OXDC1 is 100%.3.The oxalic acid content of OXDC2-10 and OXDC3-5 were 42.4% and 64.9%lower than that of the wild type,respectively.Overexpression of the SoOXDC gene can reduce the oxalate content of transgenic Arabidopsis;the transgenic plants expressing the SoOXDC gene are more resistant to exogenous oxalate than the wild type.SoOXDC may affect the activity of CAT and APX by reducing the content of oxalic acid,thereby enhancing the plant's antioxidant capacity.4.The gene expression of COR47,RD29 A,and NCED3 of the transgenic line OXDC3-5 decreased under salt stress;the transgenic Arabidopsis might increase its resistance to low temperature stress by increasing the expression of the three genes COR47,RD29 A,and NCED3;SoOXDC3 Genes are sensitive to salt stress and low temperature stress,but less sensitive to high temperature stress.5.Transcriptome analysis was performed to study the gene function of SoOXDC3.It was found that the main difference between the over-expressed SoOXDC3 line and wild-type Arabidopsis is the photosynthesis process and the specific binding of protein domains.Regulating the decrease of oxalate content may be related to RNA Modifications,apoplasts and coenzyme binding related genes,and affect carbon metabolism and glyoxylic acid and dicarboxylic acid metabolism.Wild-type Arabidopsis mainly responds to exogenous oxalic acid stress by affecting photosynthesis-related genes,starch and sucrose metabolism,phenylpropane biosynthesis,and purine metabolism.Transgenic Arabidopsis may enhance resistance by affecting genes related to plastid tissue,carbon metabolism pathways and amino acid biosynthesis pathways.After oxalic acid stress,the responsive of overexpression SoOXDC3 gene in Arabidopsis is mainly invovled in the chitin response,plant cell wall and oligopeptide transmembrane transport activity related gene expression,etc.The differential genes are mainly enriched in the oxidative phosphorylation pathway.The increased resistance of transgenic Arabidopsis to oxalate may also be related to these genes and oxidative phosphorylation metabolism.