Effects Of Cd²⁺ Stress On The Biological Characteristics And Differential Expression Of Some Antioxidant Genes In Bostrychus Sinensis

As an important metabolic regulation system,antioxidant defense system can remove organism's oxygen free radicals from themselves generated or external stimulus.There are many antioxidant genes in the antioxidant defense system and they form various defense lines according to the function of action site.The first defense line mainly includes superoxide dismutase?SOD?,catalase?CAT?,glutathione peroxidase?GPX?.They are the most important and indispensable part of the whole antioxidant defense strategy.The antioxidant defense mechanism and antioxidant genes about SOD,CAT and GPX have been extensively studied in other teleost but of the Bostrychus sinensis is almost blank.Bostrychus sinensis is an economic dominant species in the southeast coast of China.It is delicious in taste and rich in nutrition,and has the effect of promoting wound healing recognized in traditional medicine.In recent years,with the pollution of the coastal environment,especially the presence of heavy metals or toxic substances,the habitat of Bostrychus sinensis has been destroyed,which aggravates the oxidation pressure in the body,and leads to immune and antioxidant reactions in fish.Based on the current research background,my research is divided into three parts:1.Literature review,through the introduction of three aspects of Bostrychus sinensis,cadmium and anti-oxidation defense system,as well as the general situation of research progress,leads to the starting point of the paper and the purpose and significance of the research.2.The acute toxic effect of cadmium ion on Bostrychus sinensis.This chapter mainly focuses on the 96-LC₅₀ of Bostrychus sinensis under cadmium ion stress:first,carry out pre experiment,set up seven different concentration gradients with large span,use 10 fish seedlings for each concentration,record the data every 24 hours,record four times,i.e.the total experiment time is 96 hours,and find out the 100%lethal concentration and the minimum lethal concentration of Bostrychus sinensis under cadmium ion stress Dead concentration.Then,a formal experiment was carried out.Five concentration groups were set up?the concentration was in the sequence of equal difference?,one control group,and three parallel groups were set up at the same time.The results showed that the fish had higher resistance to cadmium ion than other hard bone fish in terms of acute toxicity of heavy metal cadmium,and made several inference about the results,which may be related to the strong regulation of antioxidant genes in vivo or the high content of non enzymatic cell antioxidants and cell PUFA in fish.In addition,the experimental environment was poor differences also affect the measured experimental values.3.cDNA cloning and expression of four key antioxidant genes of ophiocephalus sinensis.In this study,we cloned and obtained the coding sequence?CDS?sequences of two genes?BsCu/Zn-SOD and BsMn-SOD?,catalase gene?BsCAT?and glutathione peroxidase gene family?BsGPx1a and BsGPx1a?in Bostrychus sinensis.The open reading frame of BsCu/Zn-SOD gene contains 465bp nucleotides and encodes 154 amino acids;the open reading frame of Mn-SOD gene contains 678bp nucleotides and encodes 225 amino acids;the total length of cat core sequence is 1578bp and 525 amino acids are deduced;the total length of GPx1a core sequence is 576bp and 190 amino acids are deduced;the total length of GPx1b core sequence is 567bp,187 amino acids were deduced.Sequence analysis showed that BsCu/Zn-SOD gene had no signal peptide and was an intracellular protein;Mn-SOD gene contained 27aa mitochondrial targeting sequence at N-terminal;bscat gene had high homology with other hard bone fishes,especially for catalase near active site and catalase near heme ligand signature sequence,which were very conservative and three amino acid catalytic sites His⁷⁵,The active sites of His⁷⁵ and Tyr³⁵⁸;GPx1a and GPx1b core sequences all found selenocysteine residues encoded in a TGA codon.These key anti-oxidation genes were proved to be conservative.4.The effect of cadmium ion on the gene expression of BsCu/Zn-SOD,Mn-SOD and CAT.In this study,we detected the gene expression of five main immune and external contact tissues of Bostrychus sinensis under three gradients of cadmium stress based on 96-LC₅₀.We found that cadmium stress induced the gene expression of BsCu/Zn-SOD,BsMn-SOD and cat in liver,spleen,gill,skin and muscle of Bostrychus sinensis.Compared with the other three tissues,these two tissues are the main antioxidant organs of Bostrychus sinensis.The expression of BsMn-SOD gene in Bostrychus sinensis increased with time,which is more obvious than that of BsCu/Zn-SOD.It is speculated that in the oxidative damage of heavy metals,BsMn-SOD gene can prevent life from receiving gold It is stronger than BsCu/Zn-SOD gene in terms of toxic stress and protecting cell from oxidative damage.