文蛤COI基因片段序列分析

食品科學論文編號:SP016  論文字數:9299,頁數:21

目    錄

1  引言……………………………………………………………………………………… 1
文蛤的生物學特性………………………………………………………………………… 1
文蛤的藥用價值………………………………………………………………… 2
本試驗的目的意義…………………………………………………………………… 2
2  材料和方法…………………………………………………………………………… 3
2.1 材料 ………………………………………………………………………………… 3
2.2方法…………………………………………………………………………………… 3
3  結果和分析……………………………………………………………………………… 5
3.1文蛤基因組總DNA電泳圖……………………………………………………………… 5
3.2 文蛤PCR電泳結果………………………………………………………………………… 6
3.3測序及序列組裝…………………………………………………………………………… 6
3.4 序列比對…………………………………………………………………………………… 7
3.5構建系統發育進化樹…………………………………………………………………… 10
4  討論 ………………………………………………………………………………… 11
4.1 DNA提取……………………………………………………………………………… 11
4.2 PCR中遇到的問題……………………………………………………………………… 12
4.3 COI基因序列的應用…………………………………………………………………… 12
結論 …………………………………………………………………………………… 14
致謝 …………………………………………………………………………………… 15
參考文獻……………………………………………………………………………… 16

摘  要：本試驗以文蛤（M.meretrix）閉殼肌為材料，采用CTAB裂解法提取基因組總DNA，根據COI基因的保守序列設計引物，PCR擴增文蛤的COI基因片段序列。PCR產物送上海生工進行純化回收，并采用擴增引物進行正反雙向測序。用DNAStar （version 5.01）中的SeqMan軟件對正反序列進行組裝，得到COI基因片段的一致序列，長度為709bp（包括引物在內）。用EditSeq對所得到的序列進行編輯和分析，序列中A=21.44%， G=20.17%，T=44.85%，C=13.54%，A+T=66.29%，G+C=33.71%，使用無脊椎動物線粒體遺傳密碼把COI基因片段翻譯成氨基酸序列，此片段共編碼235個氨基酸。進入GenBank對得到的序列進行BLAST，以確定所得到的序列是COⅠ基因序列，并與親緣關系在74%以上的簾蛤科的12個不同種COⅠ基因序列用MegAlign軟件進行比對，并以住石蛤科的住石蛤相應序列數據為外群構建進化樹。系統發育樹顯示：文蛤和同屬的斧文蛤和麗文蛤聚在一起，這和傳統分類吻合。
關鍵詞： 文蛤 COI基因 序列分析

Sequence Analysis of Cytochrome Oxidase Subunit I
Gene Fragment of Meretrix meretrix

Abstract: The whole DNA of M. meretrix was extracted using CTAB method from adductor muscle. And the COI gene fragment were amplified by PCR and sequenced in Shanghai Sangon. The result showed that size of COI gene fragment was 709bp (contain the primer), the A, T, G and C contents in this fragment were 21.44%, 44.85%, 20.17% and 13.54% respectively. The A+T content was higher than G+C content. The same sequences of 12 Veneridae clams were downloaded from GenBank which have above 74% homology with M. meretrix.The homologus sequences were aligned, and the molecular phylogenetic tree was constructed with the sequence information of petricola lapicida as outgroup using MegAlign software. Tree topologies indicated that M. meretrix shared a close relationship with Meretrix lusoria and Meretrix lamarckii. It is consistent with the morphological and taxonomic results.
Keywords: M. meretrix; Cytochrome Oxidase Subunit I Gene; Sequence analysis.