Klinbunga S, Khamnamtong N, Tassanakajon A, Puanglarp N, Jarayabhand P, Yoosukh W
Marine Biotechnology Research Unit, National Center for Genetic Engineering and Biotechnology (BIOTEC), National Science and Technology Development Agency, Bangkok 10400, Thailand.
Mar Biotechnol (NY). 2003 Jan-Feb;5(1):27-36. doi: 10.1007/s10126-002-0047-4.
Molecular genetic keys for identification of 3 commercially cultured oysters (Crassostrea belcheri, Crassostrea iredalei, and Saccostrea cucullata) in Thailand were developed based on restriction analysis of 18S ribosomal DNA and cytochrome oxidase subunit I (COI). Digestion of the amplified 18S rDNA with Hinf I unambiguously differentiated Crassostrea oysters from Saccostrea oysters and Striostrea (Parastriostrea) mytiloides. In addition, species-specific restriction fragment length polymorphism patterns of C. belcheri, C. iredalei, and S. cucullata were consistently observed when the gel-eluted COI was digested with Mbo I and Dde I. Thirty composite haplotypes were observed across all individuals. Species-specific composite haplotypes were found in C. belcheri (AAAA and AAAB), C. iredalei (AABC and AABU), and S. cucullata (BBCD and BBCE), respectively. The most common composite haplotype of COI in C. belcheri (AAAA), C. iredalei (AABC), and S. cucullata (BBCD) was amplified, cloned, and sequenced. Detection of C. belcheri and C. iredalei based on polymerase chain reaction was further developed using more specific primers (HCO2198 and R372) followed by digestion of a 372-bp product with Mbo I.
基于对18S核糖体DNA和细胞色素氧化酶亚基I（COI）的限制性分析，开发了用于鉴定泰国3种商业养殖牡蛎（贝尔彻巨牡蛎、艾氏巨牡蛎和僧帽牡蛎）的分子遗传密钥。用Hinf I消化扩增的18S rDNA可明确区分巨牡蛎属牡蛎与僧帽牡蛎属牡蛎以及类条纹牡蛎（副条纹牡蛎）。此外，当用Mbo I和Dde I消化凝胶洗脱的COI时，始终能观察到贝尔彻巨牡蛎、艾氏巨牡蛎和僧帽牡蛎的物种特异性限制性片段长度多态性模式。在所有个体中观察到30种复合单倍型。分别在贝尔彻巨牡蛎（AAAA和AAAB）、艾氏巨牡蛎（AABC和AABU）和僧帽牡蛎（BBCD和BBCE）中发现了物种特异性复合单倍型。扩增、克隆并测序了贝尔彻巨牡蛎（AAAA）、艾氏巨牡蛎（AABC）和僧帽牡蛎（BBCD）中最常见的COI复合单倍型。使用更特异的引物（HCO2198和R372）进一步开发了基于聚合酶链反应检测贝尔彻巨牡蛎和艾氏巨牡蛎的方法，随后用Mbo I消化372 bp的产物。
