| Aim: To further clarify its genetic background and its biomedical application of carassius auratus red variety inbred strain, we were going to establish AFLP marks to carassius auratus red variety inbred strain and study toxicity on carassius auratus red variety inbred strain recessiveness about cadmium and analysis on application in environmental toxicology of carassius auratus red variety inbred strain, , discusses the advantages and disadvantages of fish as laboratory recessiveness toxicity test materials.Methods: In experiments with ordinary recessiveness recessiveness for experimental material, the study of polymorphism recessiveness AFLP, and mark the AFLP markers recessiveness experiment, Through the cadmium ions to experiment recessiveness toxicity testing, test, the main tissue cells observed and pathological damage biochemical studies to determine the experiment of cadmium red hybridaze sensitivity and toxic injury and cadmium ions concentration without relevance between.Results:In order to further clarify the genetic background and application in biological medicine of the experimental carassius auratus red variety of C1HD, this study took experimental carassius auratus red variety and common carassius auratus red variety as the experimental materials, and studied AFLP polymorphic markers to experimental carassius auratus red variety and the biological toxicity of cadmium ions on experimental carassius auratus red variety of C1HD. The major research findings are as follows:1. AFLP Markers to carassius auratus red variety inbred strain C1HDThe research applied AFLP technology, and totally adopted 64 primer combinations to amplify 13 carassius auratus red variety inbred strain C1HD and 12 common Carassius auratus red variety, and 10 primer combinations with rich electrophoresis general bands and high polymorphism site proportion were selected; the 10 selected primer combinations detected five gene specific bands which existed in Carassius auratus red variety inbred strain C1HD and/or common Carassius auratus red variety. E3M6 primer combinations detected a band around 428bp which was specific to normal Carassius auratus red variety, and E4M5 primer combinations detected a fragment around the 293bp which was specific to Carassius auratus red variety, and E4M6 detected a fragment around 85bp which was specific to Carassius auratus red variety, and E4M6 detected a fragment around 127bp which is specific to Carassius auratus red variety inbred strain C1HD, and E6M6 detected a fragment around 317bp which is specific to Carassius auratus red variety inbred strain C1HD.2. Acute Toxicity Experiment of Cadmium Ions on Carassius auratus red variety inbred strain C1HDThe static bioassay method was adopted for conducting acute experiment of cadmium ions on the larvae red carp, and 24h, 48h, 72h and 96h median lethal concentrations and the safe concentration of cadmium ions on red carp were determined. The results show that the 24h, 48h, 72h and 96h median lethal concentrations of cadmium ions on the experimental red carp are respectively: 15.85mg / L, 9.22 mg / L, 7.02 mg / L and 6.01 mg / L, and the safe concentration is 0.6mg / L. According to the toxicity evaluation criteria of toxic substances on fish, the cadmium ions are highly toxic substances to Carassius auratus red variety inbred strain C1HD.3. Research on Mutagenic EffectsMicronucleus test was utilized to detect the damages of red blood cells of Carassius auratus red variety inbred strain C1HD, results show that the red blood cell micronuclei rate of the cadmium treatment group was significantly higher than of the control group (P <0.01), and with the increased concentration of cadmium ions, red blood cell micronuclei rate of Carassius auratus red variety inbred strain C1HD was increased, which showed a clear dose-effect relationship.4. Influence on histopathology of Carassius auratus red variety inbred strain C1HDMucus on the body surface and gills of poisoned Carassius auratus red variety inbred strain C1HD were increased and the gills are attached with slightly black floc. The carps showed neurological symptoms and breathing difficulties and died in severe cases. Epithelial cells of small pieces of gill were swelled and capillaries were gradually separated. Liver cells and renal tubular epithelial cells showed granular degeneration and vacuolar degeneration, and even showed dissolved necrosis. Splenic sinuses were expanded and congested, and generated hemorrhage, splenic parenchyma reticular cells were swelled and proliferated, macrophages were increased and their volumes were increased ,and the splenic intercellular intervals were increased, and more hemosiderin was deposited. Myocardial cell generated edema, intercellular interval were increase, and the cells were arranged loose and disordered, inflammatory cells were infiltrated, and even myocardial fibers were seen dissolved and fractured. All the Lesion degrees were positively correlated with cadmium ion concentration.5. Influence on Blood Biochemical Parameters of Carassius auratus red variety inbred strain C1HDDifferent concentrations of cadmium group compared with controls, TP(the total protein), ALB (albumin), GLB(Globin) increased slightly, GLU (glucose) concentrations is not changed much; The dye cadmium group with cadmium ions increased concentrations , AST (aspertate aminotransferase) and LDH (lactate dehydrogenase) in blood in both significantly higher, ALT (alanine aminotransferase) and AKP (alkaline phosphatase) increased slightly; With the increase of the concentration of cadmium ions, K+, no significant changes, andNa+ andCl-only in 3.00 mg/L group reduce ,but Ca2+continued to decline. This reflects that cadmium to acute injury and damage degree on the main recessiveness tissue organ ,among these biochemical parameters, the parameters of the enzyme is superior to other parameters, because the parameters of the enzyme not only has certain organization specificity, but also with cadmium ions concentration increased ,degree of the enzyme activity are gradually increasing, and the rise of the enzyme activity can reflect the extent of the damage of main organization of Carassius auratus red variety inbred strain C1HD.In conclusion, AFLP markers to Carassius auratus red variety inbred strain C1HD were found that E4M6 has a band around 293bp and E6M6 has a band around 317bp and they can be used as the characteristic markers of Carassius auratus red variety inbred strain C1HD; The acute toxicity experiment of cadmium on Carassius auratus red variety inbred strain C1HD shows that Carassius auratus red variety inbred strain C1HD can reflect the cadmium pollution in water environment; results of micronucleus test and major histopathological damage inspection show that the red blood cells of Carassius auratus red variety inbred strain C1HD are damaged and the pathological damage degree of major tissues show concentration-effect relationship with the quality concentration of cadmium ions and therefore can reflect the severity degree of cadmium ion pollution; Biochemical analysis results of the enzyme can reflect the pathological damage of main organization. Therefore we can believe that the Carassius auratus red variety inbred strain C1HD can be used as the biological monitor for monitoring water cadmium ion pollution.Conclusions: Through the analysis of Carassius auratus red variety inbred strain and established the experiment AFLP on carassius auratus red variety inbred strain molecular markers of AFLP, Through the experiments of cadmium to recessiveness toxicity study that can be used for monitoring recessiveness experiment in water environment pollution of cadmium ions of biological monitoring. |
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