Molecular Identification Meath For Fish And Soft-shelled Turtle

With the rapid development of the global economy,the consumption of seafood products are increasing,some retailers seek to maximize economic benefits,who sold a less expensive species,which is substituted for a more expensive species.It has seriously effected the interests of consumers and the normal market order.Howere,there is not a prefect national standard or method for the aquatic product variety identification.It means that the regulaor is lack of important technical basis for aquatic product.According to this situation,edible fishes and soft-shelled turtles are as the research object,we build a sequence library of mitochondrial Cytochrome C oxidase subunit(COI)for salmon and cod,establishment of real-time PCR method is highly specific for the detection of soft-shelled turtle,and using DNA barcode dentification fish products,which carry on a beneficial exploration for species identification and protection and provide technical support for aquatic product.1.Structure COI sequence for salmon and codDNA barcoding,based on the sequencing of a standardization region of the cytochrome c oxidase I(COI)gene,has received considerable attention as an accurate and broadly applicable tool for animal species identifications.Avoiding to edible fishes and soft-shelled turtle are substituted for cheap species,the real salmon and cod collected and structure them COI sequence are very importance.This study collected the salmonidae sample of 3 subfamily,7 genus,a total of 17 species of salmon;Gadidae and Merlucciidae,a total of 8 species of cod.All samples were PCR amplified and sequenced,following structure COI sequence for salmon and cod.The results show that the most of samples can accuracy identification species from NCBI or BOLD,except for Oncorhynchus masou,Coregonus Berg,C.Peled,C.nasus,C.muksun,Thymallus arcticus yaluensis,T.arcticus grubei and T.brevirostris,analysis of this sample provenance,Which can find this samples almostly from our country.It is probably that our country has not research COI sequence of salmon,then lead to can not accurately identify which from production areas in China.2.Using DNA barcode dentification fish productsIn this study,we used DNA barcode dentification fish products.Sixty-three seafood samples were PCR amplified and sequenced,following which species identification by querying Gen Bank and Barcode of Life Data(BOLD)DNA sequence records,then compare with their advertisement name.Of the 63 samples,16 samples were mislabeled,and the cases of mislabeling were some from import and similar in appearance fishes.The results show that our country have a high mislabelling of fish species,and the existing phenomenon of legislative deficiency or lax enforcement on labelling of fisheries products.Therefore,we recommend that the regulatory bodies can utilize of DNA barcoding to authenticate fish species,and then enhancing transparency and fair trade on the domestic fisheries market.3.Establishment of real-time PCR method for the soft-shelled turtlesIn order to prevent the soft-shelled turtles are imitated with othe cheap species,we set up a real-time PCR method,mt DNA Cytochrome b(Cyt b)sequences of Trionychidae were download from Genebank,and compard by Primer 5 software,designed PCR primers and probe.After optimizing the reaction conditions,a novel quantitative real-time PCR method for soft-shelled turtles detection was established.The results indicated that the developed realtime PCR method was high specific soft-shelled turtles detection,the limit of absolute detection(LOD)were 1pg,the standard deviation(SD)and the relative standard deviation(RSD)were all in the acceptable range.Therefore,the established real-time PCR method for soft-shelled turtles detection has high specificity and good sensitivity,and is suitable for identification of soft-shelled turtles ingredient in mixed samples quickly.