Reasearch And Application Of Carp Molecular Biomarkers

Hun River,an important tributary of the Liaohe basin, plays an importantregulatory role for regional economic development and ecological environmentalhealth. In recent years, Hun River Basin Water Environment survey shows that thetype and quantity of chemical pollutants in Hun River wbasin pollution increasessignificantly every year, in order to asreveal the potential ecological risk of chemicalcontaminants in water Hun River basin, the study investigates the Hun River basinwild fish population structure, on this basis select carp as an environmental indicatororganism, and further to evaluate the Hunhe main steam environment quality used13carp gene biomarkers such as antioxidase, expected to warm the Hunhe waterenvironment risk from the molecular level.Main contents are as follows：In the Hun River Basin wild fish community structure, there are10families,34species, the species of cyprinids account for59%of total species, and the numberaccount for90%of the total catch, so cyprinids is the main dominant species. Incyprinids, carp has a higher frequency, and widely distributed in the upstream anddownstream, easy to capture the male and female sexual characteristics, so determinethe carp as the environment bioindicator of the Hun River basin.Firstly, screen13molecular markers for the evaluation of water environmentquality in Hun Rive. Query other biological target gene and protein sequences by theNCBI database (http://www.ncbi.nlm.nih.gov/), then compare the same genesbetween different organisms used Bioedit7.0Primer Primer5.0software and getconserved sequence of amino acids, on this basis, design the degenerate primers ofcarp target gene, next clone13molecular markers of carp using degenerate primersand Real-time quantitative PCR method to quantify target gene expression level isestablished(amplification efficiency is among90%-110%, quantitative curvecorrelation (R2)>0.99), which makes the molecular biology information of carp richer. Quantify the gene expression levels of13biomarkers in summer and autumnHunhe wild carp liver tissue. Compared wth the upstream point (S1), SODmRNAgene expression is significantly inhibited while HSP30,CYP1A,Vtg mRNAexpression is significantly induced in downsteam points; calculate the integratedbiomarker response index (IBR) of the sampling points in summer and autumn, resultsshow that IBR values of the middle and lower reaches are greater than the upstreamfor both seasons,which illustrates the water quality is poorer in the middle and lowerreaches, and the pollutants had an impact on the health of wild carp; compare IBR ofsummer and autumn, IBR value in summer is significantly higher than that in the fallin every sample points, which indicates the influence of environmental contaminationon the summer organism is significantly increased.For IBR value of each point,, different marker genes has different contributionrate, in summer, the contribution rate of SOD (2.99~98.84%), CAT (1.88~68.13%),the Vtg (0.14~28.65%), CYP1A (0.02~16.77%), HSP70(0.02~27.81%), TGF(0.05~24.26%) and MT (0.014~10.10%) are larger, it can be seen that there aremany kinds of pollutants in the environment, and pollutants has caused strong stresson organisms. In autumn, CAT (0~57.30%), SOD (0~59.48), CYP1A (0~31.74), GPx(1.03~52.64%), GST (0~21.08%) and HSP70(0~14.21%) have higher contributionrate,which illustrates environmental pollutants has a strong role in the induction onbiological metabolism and stress. However, the relative contribution of the differentmarkers to IBR values is different in each point, which also reflects the difference ofevery point in the degree of water pollution and type of water contaminants. Based onthe size of the contribution rate of biomarkers to the IBR value, while taking intoaccount the Specific reaction of organisms on different pollutants, CAT, SOD,CYP1A, HSP70, Vtg and MT is recommended as effective biomarker in testing waterquality of the environment.