碩士 / 亞洲大學 / 保健營養生技學系碩士班 / 98 / Cytochrome b gene (Cyt b) is a functional gene between tRNAGlu and tRNAThr in mitochondria DNA (mtDNA), and which was usually used to study about phylogeny and evolution in vertebrate. In 2003, the so-called “DNA barcode” was proposed and allocated to identify organisms based on the sequences of cytochrome c oxidase subunit I (COI) gene in the mitochondria.
In order to identify the original species of raw materials and processed products, the complete Cyt b (1137 bp) and COI from the 14 pufferfish species including Lagocephalus gloveri、L. wheeleri、L. lunaris、L. inermis、Takifugu xanthopterus、T. oblongus、T. poecilonotus、T. pardalis、T. vermicularis、T. stictonotus、T. niphobles、Sphoeroides pachygaster、Ostracion cubicus and Arothron hispidus were amplified by PCR (Polymerase chain reaction) technique in this study. Six pairs of primers were successfully used to reconstruct the complete Cyt b sequences and the complete COI from the 14 pufferfish species. The homogeneous of the complete Cyt b and the complete COI among the 14 pufferfish species are from 66.94% to 95.75%, and from 76.49% to 99.94% individually. By the way, these results also shows that the Cyt b gene is more suitable for reconstructing species phylogeny of pufferfish than the COI gene.
In order to apply the DNA barcode technique in identification of various processed products, the primers were designed to amplify the short fragment within the length limitation of the confident sequencing. Than the DNA barcode technique was also applied to identify the species of L. gloveri、L. wheeleri、L. lunaris、L. inermis、T. xanthopterus、T. oblongus、T. poecilonotus、T. pardalis、T. vermicularis、T. stictonotus、T. niphobles、S. pachygaster、O. cubicus and A. hispidus in the simulated producte and the commercial processed products. The results indicated that DNA barcode of the simulated samples could be amplified under the condition 121 ℃ for 10 ~ 80 mim. In contrast, that were not DNA barcode fragament be amplified under the sterilize condition 121 ℃ for 90 mim. Therefore, the new design DNA barcode fragament amplified by primers TLS-LC and TLS-HC were recommended as DNA barcodes for raw materials and processed products.
Keywords: DNA barcode, COI, Cyt b, pufferfish, processed products, species identification, PCR.
| Identifer | oai:union.ndltd.org:TW/098THMU8115015 |
| Date | January 2010 |
| Creators | Chung Yun-Lung, 鐘云隆 |
| Contributors | 謝承紘 |
| Source Sets | National Digital Library of Theses and Dissertations in Taiwan |
| Language | zh-TW |
| Detected Language | English |
| Type | 學位論文 ; thesis |
| Format | 129 |
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