Primary Study Of Identification Of Several Sea Cucumbers Based On Mitochondrial DNA Sequences And Its' Application On Productions

The sea cucumbers, class Holothuroidea, constitute one of the six extant classes of the exclusively marine phylum Echinodermata. The approximately 1200 species of sea cucumbers are partitioned into six orders. Sea cucumber orders are distinguished on the basis of gross morphological characters such as the morphology of the tentacles, the presence or absence of tentacular retractor muscles, the presence or absence of an internal respiratory tree, and the presence and distribution of podia on the trunk. At the familial level, taxonomic distinctions are dependent on a finer level of characters such as the morphology of dermal ossicles, the form of the esophageal calcareous ring, the distribution and morphology of the tube feet, etc. At the species level, the differences in morphological can be quite subtle, obscuring taxonomic distinctions. Because of the lack of the scientific, methods for detecting processed dried cucumbers, it's necessary to find a simple and accurate way to resolve this problem.Amplified the mitochondrial COâ… genes extracted from Apostichopus japonicus, Actinopyga miliaris, Acaudina leucoprocta, Holothuria atra, Holothuria edulis, Parastichopus californicus, Thelenota ananas by PCR.691-692bp-size products were detected by electrophoresis,then purified for cloning and sequencing.The results of sequencing show that the average GC contain is 40.0% to 44.6% and there are no obvious differences among COâ… fragment of the seven samples.Take the sequencing data of Riben cucumbers as the standard,we find that Actinopyga miliaris, Acaudina leucoprocta, Holothuria atra, Holothuria edulis, Parastichopus californicus, Thelenota ananas have more transition sites in all samples by using MEGA software and Thelenota ananas has 124 transition sites,while all the Apostichopus japonicus only have 18 transition sites. The difference of hereditary distance between 7 sea cucumbers is few, while between Acaudina leucoprocta, Holothuria atra, Holothuria edulis, Thelenota ananas and other samples is obvious(0.196-0.212,0.197-0.225,0.196-0.243,0.179-0.213). Parastichopus californicus is 0.095-0.229,and others is 0.003-0.012.According to the NJ and MP systerm trees based on the calculation of the hereditary distances of samples, we can have a result that Apostichopus japonicus from Russia, Apostichopus japonicus , Apostichopus japonica from GuanXi, Apostichopus japonica from BieHaiDao, Apostichopus japonicus from DaLian formed a close relationship.Amplified the mitochondrial 16Sr RNA genes extracted from Apostichopus japonicus, Actinopyga miliaris, Acaudina leucoprocta, Holothuria coluber semper, Holothuria mexicana, Parastichopus californicus, Actinopyga mauritaana by PCR.330bp-size products were detected by electrophoresis, then purified for cloning and sequencing. The results of sequencing show that the average GC contain is 39.4% to 48.1% and there are no obvious differences among COâ… fragment of the seven samples.Take the sequencing data of Riben cucumbers as the standard, we find that Acaudina leucoprocta, Holothuria coluber semper,Holothuria mexicana have more transition sites in all samples by using MEGA software. The difference of hereditary distance between 7 sea cucumbers is few, while between Acaudina leucoprocta, Holothuria coluber semper,Holothuria mexicana and other samples is obvious(0.150-0.291,0.140-0.282,0.140-0.277). According to the NJ and MP system trees based on the calculation of the hereditary distances of samples, we can have a result that Apostichopus japonicus from Russia, Apostichopus japonicus , Apostichopus japonica from GuanXi, Apostichopus japonicus from DaLian, Actinopyga miliaris, Actinopyga mauritaana, Acaudina leucoprocta, Parastichopus californicus formed a close relationship.We find that the aberrance of COâ… genes is more obvious than 16Sr RNA genes through the analysis of COâ… genes and 16Sr RNA genes,so COâ… genes are used in identifying the products of the sea cucumber. mitochondria of the sea cucumber nutrient solution extracted with optimized method. Agar gelose electrophoresis and PCR were carried out, then purified for cloning and sequencing. The results of sequencing show that the sea cucumber in the sea cucumber nutrient solution is Apostichopus japonicus from Qingdao.The successful amplification of the COâ… and16Sr RNA fragment reconfirms the primers are applicable for Echinoderm. In this paper, we set up a method that is able to identify a number species of sea cucumbers directly, and it is helpful to the study of the identification and molecular evolution on the sea cucumber.