The Function Of MMDH2 Gene In Regulating The Cadmium Stress Response In Arabidopsis Thaliana

Environmental pollution is a challenging threat to the modern world.Cadmium heavy metal contributes to the Soil pollution,hence the treatment and anticipation of these heavy metal pollutants will minimize the adverse effects on the environment and upturn to the crop yield.This experimental work focused on mitochondrial malate dehydrogenase gene mMDH2 in Arabidopsis,It has been found that the expression level of mMDH2 can be inhibited by CdCl₂.However,some of the functions under the cadmium stress signal and response mechanisms are not yet clear.Therefor,we conducted research on the basis of these questions,the results are as follows:1.In CdCl₂ stress treatment conditions,The T-DNA mutants insertion to mmdh2-1and mmdh2-2 showed significant tolerance compared to WT.This result indicates that the mMDH2 gene play an important role in cadmium stress.2.The mMDH2 overexpression vector was constructed.Using the specific primers,the mMDH2 gene was cloned by PCR and ligated into a vector with a 35S promoter,and then the 35S::mMDH2 vector was transferred into the Agrobacterium tumefaciens GV3101 strain,which has been finally transferred to the wild type plants,the positive overexpressing plants were obtained by screening.3.Under the Cd stress conditions,mMDH2 overexpressing plants showed a sensitive phenotype compared with WT,indicating that the mMDH2 gene plays a negative regulatory role in cadmium stress response in Arabidopsis thaliana.4.The results of DAB staining showed that the H₂O₂ content in mmdh2 mutant plants was significantly lower compared with WT under the cadmium stress condition,while the overexpressing plants were significantly higher than WT;Further,the results of MDA assay showed that the MDA content in mmdh2 mutant plants was also lower than WT under the condition of cadmium stress;However,the MDA content of overexpressing plants was higher than WT.It was indicated that mMDH2 gene may participate in cadmium transportation and regulation by ROS pathway.5.The expression level of cadmium stress-related genes was analyzed by qRT-PCR.It was found that in cadmium treatment,the expression level of PDR8 gene in mmdh2 mutant plants was higher than WT,while the overexpressing plants was lower than WT,indicating the change of mMDH2 gene expression level affect the expression level of PDR8.6.Determination of cadmium content in roots and stems of WT,mmdh2 mutant and overexpressing plants under CdCl₂ stress,The results showed that the cadmium content in roots and stems of mutants was significantly lower compared with WT,while the overexpressing plants was higher than WT.It was showed that mMDH2 gene affect the gene expression of PDR8,and further participate in the regulation of Cd stress response.In conclusion,the mMDH2 gene negatively regulates cadmium stress response in Arabidopsis thaliana.It not only reveals a new mechanism of plant tolerance to heavy metal cadmium stress,but also provides new genetic resources for phytoremediation genetic engineering.