| With the development of the industrialization process,heavy metal cadmium gradually penetrates into China’s agricultural land,and land cadmium pollution will not only seriously affect the growth and development of plants,Eventually,these chromium elements will accumulate in the human body,causing irreversible harm to the human body.Therefore,utilizing scientific and efficient hyperaccumulation plants to repair cadmiumcontaminated agricultural soil can ensure the safety of food production in China,and has numerous ecological and economic implications.As one of the main oilseed crops in China,Brassica napus has a wide planting base in all parts of China,and can be used as a plant resource for restoring cadmium polluted soil.In this study,using Arabidopsis and Brassica napus as the research objects,the cadmium-tolerant candidate genes screened based on genome-wide association analysis were analyzed,and 11 genes were verified,and two genes were identified to significantly improve the cadmium tolerance of the transgenic material.The main conclusions can be briefly stated as follows:1.The functional analysis of the candidate gene for high cadmium tolerance in cabbage rape showed that these 11 genes had the functions from divalent cation transporter and ion channel to glutathione transfer and antioxidant response,respectively,and may be involved in the cadmium detoxification mechanism of plants from these three directions.2.Candidate gene expression in response to cadmium stress.Cadmium stress treatment and fluorescence quantitative analysis were performed on hydroponic seedlings of rapeseed Westar material plants to determine the gene expression of candidate genes under stress.It was found that the expression of Bna A06g04090D(Bnamp LPD1)and Bna A09g29540D(Bna GSTU13)genes was significantly elevated and stable,indicating that these two genes are probably related to the growth regulation under cadmium stress.3.Construction of overexpression vectors and expression identification of transformed plants.Eleven Brassica napus cadmium-related candidate genes were constructed as overexpression vectors,with col-0 as the receptor material,genetic transformation,and then cadmium-stressed treatment of Arabidopsis-thaliana positive strains,and two mutants OEBnamp LPD1 and OE Bna GSTU13,which were significantly superior to WT at all levels of cadmium treatment,were screened from 11 mutants.4.Determination of physiological indexes of overexpressed strains.The fresh weight and root length characteristics of OE Bnamp LPD1 and OE Bna GSTU13 strains under different concentrations of cadmium stress were investigated,and the results showed that the fresh weight and root length of plants in the experimental group were significantly higher than those of wild Arabidopsis thaliana at different concentrations,indicating that the overexpressed strains had the ability to tolerate cadmium.5.Cadmium ion content determination.The cadmium content of OE Bnamp LPD1,OE Bna GSTU13 and WT was determined at different treatment levels,and the results showed that the cadmium content in OE Bnamp LPD1 and OE Bna GSTU13 was significantly higher than that of WT.6.Genetic transformation of Brassica napus.The overexpression vectors of Bna A06g04090 D and Bna A09g29540 D genes were constructed using Westar cabbage as the recipient material for genetic transformation.7.Analysis of protein physicochemical properties.The physicochemical properties of Bna A06g04090 D and Bna A09g29540 D genes were analyzed,and the results showed that both genes were relatively stable and hydrophilic proteins.This study can provide new germplasm resources for the cultivation of cadmium-tolerant varieties of rapeseed,and new plant resources for phytoremediation technology,which has positive significance for the remediation of cadmium-contaminated land. |
PDF Full Text Request