Research On The Speciesidentification Of Processed Shark Ceratotrichia Based On DNA Bar-Coding Technique

Processed shark ceratotrichia is a kind of traditional Chinese seafood,andbelongs to thehigh-value processed category. With the rapid growth ofdomestic GDPlevel, shark ceratotrichia consumption has reached toa relatively high point, bringingabout severalfraud cases in the seafood market. Gelatinmaterialsor ray fins are madeas “processed shark ceratotrichia” to confuse consumers. What’s more serious isthatendangered shark species are caught illegally to obtain fins.Theseimproperbehaviors threaten the existing endangered shark species. Thephenomenonabovesetsenormous obstacle in shark ceratotrichia tradeand calls for anurgent need of species identification methods. Usual ways of sharkceratotrichiaidentification (mainly morphological identification) may lose accuracysince processing progresses cause the loss of morphological features correspondingtocertain shark species. DNA bar-coding technique has now applied widely in speciesidentification of processed aquatic food or food material due to its accuracy. Thus webase species identification work mainly on DNA bar-coding technique.Shark ceratotrichiashould be divided asdense connective tissue, which isdeficient in cell amount. Samples for identification in this research is a kind ofproducts named“Ming Chi”, kind of which is considered as represent of processedshark ceratotrichia. Processing procedures of “Ming Chi”includes skinning, boning,removing remains as well as bleaching by hydrogen peroxide and then sun-drying.For the consideration of the above reasons, special DNA extracting method isrequired to meet quantity and quality of DNA to PCR requirement. Before digesting,we put forward a rising and vacuum freeze drying step for samples grinding, whichcontributes to produce a highly degree of tissue comminution. DNA extractingmethods chosen for comparison are Chelex-100method, CTAB method, STE method, improved CTAB method and Omega DNA purification kit. Improved CTAB methodincluding modification of replacing PCI with Tris-HCl balanced phenol and replacingisopropylalcohol(IPA)with new precipitatingagent(IPA:5M NaCl=3:1). DNA qualityvalue as well as PCR amplification result are all considered to evaluate the efficiencyof different extracting method, which shows that improved CTAB is a suitablemethodfor DNA extracting.As to reveal the species consistence and distribution of domestic processed sharkceratotrichia products, Tsingtao and Guangdong province are chosen. The former oneis the trading center of processed shark ceratotrichia products for the northern China,and the latter one is the place where more than90%of processed shark ceratotrichiaproducts trading is taken place. This research are conducted on107shark ceratotrichiasamples, and accuracy of identification work based on16S rRNA sequence isconducted as well. According to the result, shark ceratotrichia products are obtainedfrom various shark species, while in this study, covering10shark species, for instance,I. oxyrinchus, S.lewini, C. falciformis and P.glauca. And the results from twosequences is same. Most samples joins the reference samples from Gen Bank as asingle branch in the phylogenetic relationship. Ambiguous identification results arefound among5samplesbased on16S rRNA sequence. These samples areidentification based on CO Ⅰ sequences.And further study shows that thisphenomenon is caused by the lack of16SrRNA or mitochondrion sequenceinformation of two kinds of shark (C. tilstoni andC. dussumieri) in the Gen Bank. Andthus we can reach to a conclusion that16S rRNA sequence is suitable for sharkceratotrichia identification.According to the analysis of shark species information, we found two kinds ofSphyrnidae shark (S. lewini and S.zygaena) is under protect of CITES (consists as14.01%of the overall samples), the trade activity of these two kinds of shark and itsrelevant products is forbidden. According to China Law of the People’s Republic ofChina on the protection of Wild Life, these kinds of shark is classified as the second protection classification. Othe shark species like the A. superciliosus and I. oxyrinchusbelong to the “VU” category corresponding to the IUCN red list. Despite the fact thatthere is lacking of mandatory law to limit the trade activity of these two kind of sharkand its relevant products, it’s strongly suggested that Bureau of FisheriesAdministration take special care of these two kinds of shark. And in order to acquiresustainable development of shark industry, species based measure should be taken toguarantee the legitimacy of shark trading activity and particularly maintain theecological status of threatened or “near-threatened” shark species.