Research on Cd Absorption,Accumulation and Tolerance of Taraxacum ohwianum Kitam.

Author:Cheng Hai Tao

Supervisor:ning wei


Degree Year:2019





Taraxacum ohwianum Kitam.,which belongs to Taraxacum of Compositae,is a perennial herb.In recent years,the problem of excessive heavy metals in traditional Chinese medicines has become increasingly prominent,which has become a major factor affecting the quality of medicinal materials and drug safety,and has restricted the development of traditional Chinese medicine industry.As a common Chinese medicine,Taraxacum ohwianum is a widely used in northeast China.However,Cd excess occurs frequently in the production of raw medicinal materials.As a hyperaccumulator,the hyperaccumulation law and tolerance mechanism of Taraxacum ohwianum remain unclear.It is of great theoretical significance to study the law of Cd absorption and enrichment and the mechanism of tolerance to Cd stress in Taraxacum ohwianum in order to guarantee the quality of medicinal materials from the source.From the perspective of plant physiology,this research studied the changes of plant osmotic regulatory substances,antioxidant enzyme activities and non-enzyme antioxidant substances,so as to clarify the physiological response of plants to Cd stress.The absorption and accumulation of Cd and the distribution of Cd in cell ultrastructure were studied from the perspective of cell physiology.In order to reveal the molecular mechanism of plant tolerance to Cd stress,the function of Cd gene in Taraxacum ohwianum was also studied at the molecular level.The main research results of this paper are as follows:1.Differences in Cd tolerance of five dandelions: With the increase of Cd concentration,biomass of the dandelions except for Taraxacum mongolicum increased first and then decreased.Under the same Cd concentration treatment,the biomass of Taraxacum ohwianum was the highest and the reduction was the smallest,while that of Taraxacum mongolicum was the lowest and the decrease was the largest.Under the same concentration,the content of accumulated Cd in tissues of Taraxacum ohwianum was the highest,and the bioconcentration factor and transfer coefficient in roots and aboveground parts were the highest,followed by Taraxacum antungense.The Cd content,bioconcentration factor and transfer coefficient in Taraxacum mongolicum were the lowest.While the Cd contents in tissues of Taraxacum asiaticum and Taraxacum liaotungense were similar.The effects of Cd stress on the contents of soluble sugar and soluble protein in five dandelions were significantly higher than those in the control group.The contents of soluble sugar and soluble protein in the same tissue of Taraxacum ohwianum were significantly higher than those in other dandelions,and the contents of aboveground part and roots were respectively increased by 6.6%-51.8% and 5.2%-62.5%.2.Effects of Cd stress on seed germination and seedling growth of Taraxacum ohwianum: Different Cd concentrations had different effects on seed germination and seedling growth of Taraxacum ohwianum.The Cd concentration of 5 mg/L could promote seed germination and seedling growth.With the increase of Cd concentration,seed germination and seedling growth were inhibited more intensively.The inhibition of Cd stress on radicle growth was greater than that on embryo growth.Additionally,the content of Cd enriched in the seedling also increased,and Cd content in the radicle was much higher than that in the embryo.3.Effects of Cd stress on resistance index of Taraxacum ohwianum: With the increase of Cd concentration,the enzyme activities of SOD,POD,GR,GPX and CAT in the roots and leaves of Taraxacum ohwianum showed an overall increasing trend,and that of APX decreased.The content of nonenzymatic antioxidant ASA and DHA increased and that of GSH decreased.The content of H2O2 and O2.-increased,while the content of MDA and total chlorophyll increased at first and then decreased.4.Cd distribution in tissues and subcellular structure of Taraxacum ohwianum: In the cross section of roots,Cd was mainly stored in the cork layer,inner part of phloem and xylem.Besides,cork cell wall,intercellular space,laticifer group of phloem ducts and cell contents were the main sites for Cd in roots.The epidermal cell wall,laticifer group and intercellular space in the leaves were the main sites where Cd exists.Meanwhile,Cd was abundant in epidermal cell wall,phloem parenchyma cell wall and nonglandular hair in scapes.Under scanning electron microscope,the content of Cd in phloem and phellem was higher in roots.When the concentration was 60 mg/L,there was a decreasing and then increasing trend from the outside to the inside,that is,the content of Cd in epidermal cells and annular laticifer group around xylem was higher in roots.In the ultrastructure of root cells,Cd mainly accumulated in the cell wall,vacuole and intercellular space.In the leaves,large vacuole and cell wall of mesophyll cells contained a large amount of Cd.5.The To NRAMP2 gene sequence cloned from Taraxacum ohwianum contained 482 amino acids and 1640 bp bases.The analysis of NRAMP biological information showed that the closest relative to the To NRAMP2 gene is Lactuca sativa of Compositae.Through the tissue-specificity study of To NRAMP2 gene,it was found that the expression level of the gene in roots was the highest in the root,followed by leaves,scapes and flowers.Within 2 hours,with the extension of Cd treatment time,the gene expressions in roots and leaves showed an upward trend,and then a downward trend.The combination of PGEX4T-1-To NRAMP2 recombinant protein and Cd ion has made the strain more tolerant to Cd and improved the survival rate of the strain.To NRAMP2 subcellular localization analysis revealed that To NRAMP2 protein was located on the cell membrane.