碩士 / 國立臺灣海洋大學 / 食品科學系 / 94 / Approximately 88,000 species of gastropod mollusk distribute in the world. However, several species of Naticidae, Nassariidae and Olividae have been detected to contain marine toxins, and outbreaks of food poisoning cases in Taiwan occurred. Species is usually identified by morphological characteristics of marine snail. But it is difficult to identify without shell. Recently, species identification is developed based on DNA analysis, especially the polymerase chain reaction-restriction fragment length polymorphism ( PCR-RFLP ).
In order to identification of several toxic gastropod species in Taiwan, the direct sequencing and the PCR-RFLP technology were used to identify gastropod species in fresh, frozen, cooked ( 100℃, 30-60 min ) and steam sterilized ( 121℃, 15-30 min ) meat of Natica lineata, N. vitellus, Polinices didyma, P. tumidus,. Nassarius glans, N. papillosus, Niotha clathrata, Zeuxis scalaris, Oliva lignaria and O. reticulata.
Judging from the data of this study, high molecular weight DNA（＞1,000 bp） was obtained from fresh, frozen and cooked meat, while relatively low molecular weight DNA was obtained from samples treated by high temperature. The PCR primers of 16S L22/R22 specifically amplify 383-396 bp of fragments from fresh, frozen and cooked meats; smaller fragments of 244-257 bp are used for steam sterilized meats amplified with primers of 16S LS22/RS22.
After analyzing the different sequences amplified with primers 16S L22/R22 and 16S LS22/RS22, five restriction enzymes with specific cutting sites, including Apo I, Pac I, Hinc II, Acc I and Vsp I, were used. The restriction enzyme Apo I could differentiate the species of Natica lineata, N. vitellus, Polinices didyma and P. tumidus. Pac I, Hinc II and Acc I could differentiate the species of Nassarius glans, N. papillosus, Niotha clathrata and Zeuxis scalaris. Vsp I could differentiate the species of Oliva lignaria and O. reticulata. The polymorphic pattern in the DNA electrophoretic gel could support to identify these species precisely and quickly. By the same restriction enzyme could differentiate species of fresh, frozen, cooked and sterilized meats simultaneously.
Furthermore, applying above method to identify the gastropod species in the residue of cooked gastropod meat from food poisoning incident in Hsiau Liouchiou Island, after shell identification, sequence comparing and PCR-RFLP analyzing, it indicated that the species of gastropod was Nassarius papillosus. Therefore, by this study could provide useful technique in identifying the gastropod species of different heating meat.
| Identifer | oai:union.ndltd.org:TW/094NTOU5253018 |
| Date | January 2006 |
| Creators | Shin-Yuan Lin, 林欣緣 |
| Contributors | Deng-Fwu Hwang, 黃登福 |
| Source Sets | National Digital Library of Theses and Dissertations in Taiwan |
| Language | zh-TW |
| Detected Language | English |
| Type | 學位論文 ; thesis |
| Format | 92 |
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