海参(Holothurians)DNA条形码的构建及初步应用.docVIP

海参DNA条形码的构建及初步应用 律迎春（1988-），硕士研究生，研究方向为水产品化学。Email：oasis0212@163.com 通讯作者：唐庆娟（1971-），博士，硕士生导师 联系方式053282032957 email：tangqingjuan@， 基金资助情况:海参加工新技术，国家863计划，2007AA091805； 海珍品营养保持技术研究开发与产业化示范，国家科技支撑计划，2008DAD94B05 共5981字 图 5个，表格3个 海参（Holothurians）DNA条形码的构建及初步应用 律迎春，唐庆娟*，左涛，段高飞，王春霞，薛长湖 （中国海洋大学食品科学与工程学院，青岛 266003） 摘要：本文研究了线粒体COⅠ基因作为DNA条形码用于海参品种鉴定的可行性。研究中得到了7种海参43个个体的COⅠ基因序列，利用DNAstar、DNAMAN和MEGA4.1软件计算了7种海参的碱基组成，不同海参之间的种间遗传距离和种内遗传距离，利用邻接法和最大简约法分别构建了分子系统树。研究结果表明：海参的种间遗传距离显著大于种内遗传距离，同种海参不同个体在系统树中分别形成各自独立的分支，提示以COⅠ作为海参DNA条形码进行品种鉴定具有一定的可行性。在建立海参DNA条形码的基础上，设计了针对仿刺参（Apostichopus japonicas）的特异性探针。对四种海参（仿刺参Apostichopus japonicas，冰岛参Cucumaria frondosa，加州拟刺参Parastichopus californicus，梅花参Thelenota ananas）进行斑点杂交实验，结果表明:该探针具有较好的特异性和较高的灵敏度,能够实现对仿刺参的鉴定。本研究为后续开展斑点杂交及基因芯片法鉴定海参品种的研究奠定了基础。 关键词：海参；线粒体COⅠ基因；DNA条形码；品种鉴定；斑点杂交 Construction and application of DNA barcoding of sea cucumber（Holothurians） LV Ying-chun, TANG Qing-juan, ZUO Tao, DUAN Gao-fei, WANG Chun-xia, Xue Chang-hu (Academy of Food Science and Engineering, Ocean University of China, Qingdao, 266003) Abstract: DNA barcoding is a diagnostic technique in which short DNA sequences can be used for species identification. The mitochondrial cytochrome c oxidase submit 1(COⅠ) sequence is used in DNA barcoding for diverse species of organisms. In this paper, the validation of COⅠ gene as DNA barcoding for species identification in sea cucumber was discussed. The COⅠ gene in seven species of sea cucumber was amplified and sequenced. The data were analyzed by using DNAstar, DNAMAN and MEGA 4.1. The DNA base composition was analyzed, and the genetic variation among species and within species were calculated. It was found that genetic distance among 7 species was higher than the distance within a species. Phylogenetic analysis was carried out with neighbor-joining and maximum parsimony methods. The topology of either tree revealed that all individuals of each species formed a strong monophyletic group. These results s