基于cytb引物的鱼粉原料鱼组成分子鉴定方法  

Molecular Identification Method of Raw Fish Species Composition in Fish Meal

作　　者：周萌[1,2,3,4] 詹燕航 刘振宇 梁日深 唐义梅[1,2,3,4] 高红燕 黄燕华 林蠡 ZHOU Meng;ZHAN Yanhang;LIU Zhenyu;LIANG Rishen;TANG Yimei;GAO Hongyan;HUANG Yanhua;LIN Li(Innovative Institute of Animal Healthy Breeding,Zhongkai University of Agriculture and Engineering,Guangzhou 510225,China;College of Animal Sciences and Technology,Zhongkai University of Agriculture and Engineering,Guangzhou 510225,China;Guangdong Provincial Key Laboratory of Waterfowl Healthy Breeding,Guangzhou 510225,China;Guangzhou Key Laboratory of Aquatic Animal Diseases and Waterfowl Breeding,Guangzhou 510225,China)

机构地区：[1]仲恺农业工程学院,健康养殖创新研究院,广州510225 [2]仲恺农业工程学院,动物科技学院,广州510225 [3]广东省水环境与水产品安全工程技术研究中心,广州510225 [4]广州市水产病害与水禽养殖重点实验室,广州510225

出　　处：《动物营养学报》2020年第12期5795-5804,共10页CHINESE JOURNAL OF ANIMAL NUTRITION

基　　金：国家自然科学基金项目(31802300);湛江市科技计划项目(2018A01017)。

摘　　要：本研究旨在开发一种灵敏、准确、简便的分子鉴定方法,实现对鱼粉中原料鱼组成的鉴定。采用cytb和16S rDNA 2对引物,分别对9种海、淡水鱼和7种商品鱼粉的基因组DNA进行扩增后发现,cytb较16S rDNA引物可获得良好的扩增效果,各样品中均出现较清晰明亮的目的条带。将cytb引物扩增鱼粉基因组DNA得到的目的片段分别连接到克隆载体pMD19-T,转化大肠杆菌DH5α感受态细胞后,挑取阳性克隆进行测序,将测序结果进行BlastN比对分析。结果显示:在7种商品鱼粉中共鉴定到33个鱼类物种,主要包括鳀属(Engraulis)、圆鲹属(Decapterus)、沙丁鱼属(Sardina)、竹荚鱼属(Trachurus)、油鲱属(Brevoortia)及罗非鱼属(Tilapia)的各种鱼类,种类数量远超产品标签所示鱼种范围。为验证该方法的可靠性,重复以上步骤,对2种添加已知鱼种的自配鱼粉进行了验证性鉴定,鉴定结果与试验设计吻合。综上可知,基于cytb引物的分子鉴定技术可实现鱼粉中原料鱼组成的鉴定,鉴定结果将为鱼粉质量的准确判断提供丰富信息。This study was to develop a sensitive,accurate and simple molecular method to identify the raw fish species composition of fish meal.Two pairs of primers,cytb and 16S rDNA were used to amplify the genomic DNA of 9 species of marine and fresh water fish,and 7 kinds of commercial fish meal,respectively.It was found that the amplification efficiency of cytb primer were superior to that of 16S rDNA,clear and bright target bands appeared for each sample.The PCR products of 7 fish meal amplified by cytb primer were ligated to pMD 19-T vector,and transformed into Escherichia coli DH5αcompetent cells,the identified positive clones were sequenced and analyzed using BlastN.The results showed that 33 fish species belonging to genera of Engraulis,Decapterus,Sardina,Trachurus,Brevoortia and Tilapia et al.were identified in 7 commercial fish meal samples.The number of fish species was far beyond that labeled on the fish meal products.To verify the reliability of the above method,two samples of self-formulated fish meal with known supplemented fish species were identified with the same methods.The results were consistent with those of the design.In conclusion,the molecular identification method based on cytb primer can be used to identify the raw fish species composition in fish meal,and the results will provide abundant information for accurate judgment of fish meal quality.

关 键 词：鱼粉 原料鱼 cytb引物 分子鉴定 

分 类 号：S963[农业科学—水产养殖]