| Summary: | 碩士 === 國立臺灣海洋大學 === 食品科學系 === 95 === Cephalopoda is an important commercial catch in Taiwan. They play important roles not only in ecological area but in business sector. The distinctive morphological or physiological parameters provide the useful information to the blood relationship. The processing usually involved the separation of head, arms, and fins, evisceration, skinning, and freezing, which resulted in the change of morphological parameters. This fact makes it desirable to have methods for identifying cephalopod species, even in those cases when morphological characteristics have been removed. In order to establish the gene identification of cephalopod species, direct sequencing and the polymerase chain reaction – restriction fragment length polymorphism (PCR-RFLP) technology were used to determine the genetic variation of seven cephalopod species in this study. The gene probe was also developed to identify the cephalopod species of frozen, fresh, cooked and related products.
The primers 16S L22/R21 were used to amplify partial mitochondrial gene of seven cephalopod species, including Suborder Oegopsida: Ommastrephes bartrami, Dosidicus gigas, Todarodes pacificus; Suborder Myopsida: Loligo chinensis, Sepioteuthis lessoniana; Order Sepiida: Sepia esculenta, and Order Octopoda: Octopus ocellatus. After analyzing the different sequences amplified with primers 16S L22/R21, two restriction enzymes including Ssp I and Ase I with specific cutting sites were used. The restriction enzyme Ssp I could differentiate the species of Ommastrephes bartrami, Dosidicus gigas, Sepioteuthis lessoniana, Sepia esculenta and Octopus ocellatus. Ase I could differentiate the species of Todarodes pacificus and Loligo chinensis. The polymorphic pattern in the DNA electrophoretic gel could support to identify these species precisely and quickly.
Furthermore, applying above methods to identify the cephalopod species of frozen, cooked, sterilized and related products. The seven cephalopod meats were frozen at -20℃ for 3 months and heated at 100℃ for 30 minutes to obtain 14 samples. All samples were found to be amplified successfully. However when the cephalopod meats were sterilized at 121℃ for 15 minutes, DNA was degraded seriously and the samples could not be amplified. Besides, we analyzed 10 commercial cephalopod processed products sold in the market, of which 9 samples were determined successfully. The processed product of Sepia esculenta was failure in amplifying the target sequence. Therefore, this study provids useful technique to identify the species of cephalopod processed products.
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