中国南海两种新发现牡蛎物种鉴定方法
Identification Method of Two Oyster Species in the South China Sea
DOI: 10.12677/OJNS.2018.61004, PDF,    国家自然科学基金支持
作者: 周玉兰, 江连英, 杨漪霞：广东医科大学附属医院临床医学研究中心，广东 湛江；闫守泉, 王家丰*：广东医科大学附属医院干细胞研发与临床转化中心，广东 湛江
关键词: 湛江牡蛎；电白牡蛎；物种鉴定；细胞色素C氧化酶I基因；HRM；C. zhanjiangensis； C. dianbaiensis； Species Identification； COI； HRM
摘要: 最近在我国南海沿岸一带发现两种新的牡蛎物种湛江牡蛎(C. Zhanjiangensis)和电白牡蛎(C. Dianbaiensis)。除了对条形码基因测序外，这两种牡蛎很难用以往的物种鉴定方法区分。本研究拟开发一种基于高分辨率熔解曲线(HRM)的新的区分这两种新牡蛎物种的方法。设计引物扩增该两种牡蛎条形码基因COI(细胞色素C氧化酶I基因)序列显著差异片段，并利用HRM技术对扩增片段进行精细区分从而区分物种信息。结果我们筛选到一组引物能够有效区分这两种牡蛎新物种。经验证该方法精确有效，简单易行，是一般实验室或水产生产单位易于掌握推广的方法。为进一步研究牡蛎物种遗传信息和牡蛎养殖具有重要意义。
Abstract: Two kinds of new oyster species (C. zhanjiangensis and C. dianbaiensis) were identified recently in the South China Sea. However, there was no identification method that could distinguish the two new oyster species except barcode gene sequencing. In this study, we aimed to develop a new identification method for the two new oyster species based on HRM. Amplifying the COI sequence by PCR, the HRM analysis was employed to distinguish the different species. One pair of HRM primers were screened out which could identify the two new oyster species accurately. Sanger sequencing certified that this method was accurate and simple which can be easily used in laboratories or aquaculture departments. This method is of great significance to further genetic study and aquaculture of oyster.
文章引用：周玉兰, 闫守泉, 江连英, 杨漪霞, 王家丰. 中国南海两种新发现牡蛎物种鉴定方法[J]. 自然科学, 2018, 6(1): 17-23. https://doi.org/10.12677/OJNS.2018.61004

参考文献

[1] 王海艳. 中国近海常见牡蛎分子系统演化和分类的研究[D]: [博士学位论文]. 北京: 中国科学院研究生院(海洋研究所), 2004.
[2] 杜玄, 郭希明, 钱鲁闽. 福建沿海巨蛎属牡蛎的主要种类及其分布[J]. 应用海洋学学报, 2009, 28(3): 399-404.
[3] Wang, H. and Guo, X. (2008) Identification of Crassostrea ariakensis and Related Oysters by Multiplex Species-Specific PCR. Journal of Shellfish Research, 27, 481-487. [Google Scholar] [CrossRef] 
[4] Wang, J.F., Xu, F., Li, L., et al. (2014) A New Identification Method for Five Species of Oysters in Genus Crassostrea from China Based on High-Resolution Melting Analysis. Chinese Journal of Oceanology and Limnology, 32, 419-425.
[5] Wu, X.Y., Xiao, S. and Yu, Z.N. (2013) Mitochondrial DNA and Morphological Identification, of Crassostrea zhanjiangensis sp. nov. (Bivalvia: Ostreidae): A New Species in Zhanjiang, China. Aquatic Living Resources, 26, 273-280. [Google Scholar] [CrossRef] 
[6] Xia, J.J., Wu, X.Y., Xiao, S., et al. (2014) Mitochondrial DNA and morphological identification of a new cupped oyster species Crassostreadianbaiensis (Bivalvia: Ostreidae) in the South China Sea. Aquatic Living Resources, 27, 41-48. [Google Scholar] [CrossRef] 
[7] 陈军, 李琪, 孔令锋, 等. 基于COI序列的DNA条形码在中国沿海缀锦蛤亚科贝类中的应用分析[J]. 动物学研究, 2010, 31(4): 345-352.
[8] Dawnay, N., Ogden, R., Mcewing, R., et al. (2007) Validation of the Barcoding Gene COI for Use in Forensic Genetic Species Identification. Forensic Science International, 173, 1-6. [Google Scholar] [CrossRef] [PubMed]
[9] 阙华勇, 刘晓, 王海艳, 等. 中国近海牡蛎系统分类研究的现状和对策[J]. 动物学杂志, 2003, 38(4): 110-113.
[10] Simenc, J. and Potocnik, U. (2011) Rapid Differentiation of Bacterial Species by High Resolution Melting Curve Analysis. Prikladnaia Biokhimiia I Mikrobiologiia, 47, 256-263.
[11] Gurtler, V., Grando, D., Mayall, B.C., et al. (2012) A Novel Method for Simultaneous Enterococcus Species Identification/Typing and van Genotyping by High Resolution Melt Analysis. Journal of Microbiological Methods, 90, 167-181. [Google Scholar] [CrossRef] [PubMed]
[12] Wongkamchai, S., Monkong, N., Mahannol, P., et al. (2013) Rapid Detection and Identification of Brugia malayi, B. pahangi, and Dirofilaria immitis by High-Resolution Melting Assay. Vector-Borne & Zoonotic Diseases, 13, 31-36. [Google Scholar] [CrossRef] [PubMed]