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Molecular Diagnosis for Changes of Methylation of MLH1 Promoter of Arabidopsis Thaliana Induced by Cadmium Stress

Author: DanCunHai
Tutor: ZhongMing;LiuWan
School: Shenyang Agricultural University
Course: Biochemistry and Molecular Biology
Keywords: Cd mismatch repair gene biomarker promoter methylation
CLC: X173
Type: Master's thesis
Year: 2011
Downloads: 7
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Abstract

It is one of the most wide area and damaging environmental problems about soil heavy metal pollution in current. It has been the key factor to effect our sustainable agriculture and ecological environmental quality and has received general attention. However, diagnosis using only chemical methods could not reveal whole spectrum of the characteristics of soil quality. Therefore, assays of ecotoxicological diagnosis are required to complement chemical ones, and in which one of the research focuses is of the technique of biomarkers. Using bisulfite DNA sequencing and combined bisulfite restriction analysis technique, effects of cadmium (Cd) stress on the methylation of MutL-homologue 1 (MLH1) promoter of mismatch repair gene of Arabidopsis thaliana seedlings are studied and the biomarkers sensitive to Cd stress are chose linking to morphological, physiological indices and condition of base mutation of MLH1 promoter. The main results are as follows:1. The different levels of Cd stress (0,0.125,0.25,1.0,3.0 mg·L-1) for 3 days have different toxic effects on the growth of A. thaliana seedlings. No significant changes are observed in leaf number and fresh mass of aerial parts. They response slowly under Cd stress, and the growth of roots of A. thaliana seedlings are restrained obviously. It can clearly be seen that heavy metal first act on the root system, and the growth of roots is more sensitive to Cd stress.2. After treatment with different levels of Cd stress (0,0.125,0.25,1.0,3.0 mg-L-1) for 3 days, total soluble proteins and chlorophyll contents in aerial parts of A. thaliana seedlings are changed to a certain extent. Compared with the control, total soluble protein contents of aerial parts decreased significantly with increasing Cd concentration (P<0.05), and no significant changes are observed in chlorophyll contents of aerial parts. It can clearly be seen that total soluble protein content are more sensitive to Cd stress than chlorophyll content of aerial parts.3. Base mutation of MLH1 promoter in mismatch repair gene of A. thaliana seedlings is changed to a certain extent under different levels of Cd stress (0,0.125,0.25,1.0,3.0 mg·L-1) for 3 days, but the percentage of mutation is too small, suggesting that they are not sensitive to low dose of Cd stress.4. Under different levels of Cd stress (0,0.125,0.25,1.0,3.0 mg-L-1) for 3 days, MLH1 promoter methylation of seedlings is generally increased with increasing Cd concentration. It indicates that they are sensitive to low dose of Cd stress.5. Total soluble protein contents of aerial parts and changes of MLH1 promoter methylation of A. thaliana seedlings are sensitive to Cd stress and both them with low doses of Cd pollutant concentration significantly exist dose-effect relationship. The above results suggest that they can serve as the potential and useful biomarker for the detection of genotoxic effects of Cd pollution on plants. Although specific regional DNA methylation of MLH1 promoter in certain degree of soil heavy metal pollution stress for sensitive response characteristics of performance, the application of these biomarkers in eco-toxicology is needed to be further investigated.

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