摘要
岩牡蛎(Crassostrea nippona)与长牡蛎(Crassostrea gigas)是世界上2种重要的海产双壳经济贝类。两种牡蛎形态相似,较易混淆,目前没有快速准确的鉴别方法。本研究采用PCR-RFLP技术首次从分子水平上鉴定了岩牡蛎与长牡蛎。利用通用引物与本研究开发引物扩增2种牡蛎的线粒体16SrDNA与COI基因片段,并对扩增出的目的基因片段进行了测序。对测序结果进行限制性内切酶特异性位点分析,选取了同一种限制性核酸内切酶AluI对PCR产物进行酶切,并用2.0%琼脂糖凝胶检测PCR产物的酶切结果。结果表明,岩牡蛎与长牡蛎PCR产物经AluI酶切后产生了大小不同的酶切片段,根据酶切图谱可有效鉴别岩牡蛎与长牡蛎。这种简单、可靠、经济的PCR-RFLP鉴定方法,能够为岩牡蛎养殖新品种的开发提供技术支撑。
The Iwagaki oyster(Crassostrea nippona)and the Pacific oyster(C.gigas)are important species of shellfish in the world.C.nippona and C.gigas are similar in shell characteristics and difficult to discriminate morphologically.There was no quick and reliable method to discriminate these two oysters.In this study,we developed a PCR-RFLP assay for the rapid and effective identification of C.nippona and C.gigas at the molecular level for the first time.Amplifications of the 16S rDNA and COI genes were performed using the universal primers and the primers newly developed in this study,respectively.The amplified target fragments were sequenced and sequences were further analyzed for the selection of restriction enzymes using a software.The PCR product was digested by the selected restriction enzyme Alu I and the digestion result were examined through electrophoresis in 2%agarose gel.Digestion of 16S rDNA and COI with Alu I generated distinct restriction patterns for C.nippona and C.gigas,allowing a clear identification.In conclusion,this simple,reliable and cost-effective method can provide technical support for the development of C.nippona.
作者
刘凯凯
李琪
LIU Kai-Kai;LI Qi(The Key Laboratory of Mariculture(Ocean University of China),Ministry of Education,Qingdao 266003,China)
出处
《中国海洋大学学报(自然科学版)》
CAS
CSCD
北大核心
2018年第A02期17-22,共6页
Periodical of Ocean University of China
基金
山东省科技发展计划项目(2016ZDJS06A06)
青岛市产业培育计划项目(17-3-3-64-nsh)资助~~