基于细胞色素C氧化酶亚基Ⅰ序列的DNA微条形码技术鉴别11种生鲜肉制品掺假的研究  

Identification of adulteration in 11 fresh meat products by DNA mini-barcoding methods based on cytochrome C oxidase subunitⅠ(COⅠ)sequence

作　　者：励炯[1] 吴琼 扈明洁 金朦娜 邱红钰[1] LI Jiong;WU Qiong;HU Mingjie;JIN Mengna;QIU Hongyu(Hangzhou Institute for Food and Drug Control,Hangzhou 310017,China)

机构地区：[1]杭州市食品药品检验研究院,杭州310017

出　　处：《浙江大学学报（农业与生命科学版）》2021年第1期52-59,共8页Journal of Zhejiang University:Agriculture and Life Sciences

基　　金：浙江省公益技术应用研究计划(LGN19C200001)。

摘　　要：建立并优化了基于细胞色素C氧化酶亚基Ⅰ(cytochrome C oxidase subunitⅠ,COⅠ)序列的DNA微条形码(DNA mini-barcoding)技术,以检测生鲜肉制品中11种常见肉类的掺假情况。样品经真空冷冻干燥处理、提取DNA后,采用通用引物进行聚合酶链式反应(polymerase chain reaction,PCR)扩增,目标条带扩增产物经切胶、回收、纯化后,进行克隆测序,并将测序结果提交到GenBank数据库(http://blast.ncbi.nlm.nih.gov/Blast.cgi)中进行Blast比对,筛选出适合猪肉、牛肉、羊肉、鸡肉、鸭肉、鸽子肉、马肉、驴肉、鹅肉、兔肉、鼠肉等11种肉类扩增的通用引物COⅠ-A,并对PCR扩增条件进行优化,建立18个掺假模型,以对低经济价值肉类的最低掺入比例进行考察验证。结果表明:11种肉类的PCR扩增效率均为100%。在这18个掺假模型中,牛肉和羊肉中掺入6种低经济价值肉类(猪肉、鸡肉、鸭肉、马肉、驴肉、鼠肉)的最低检出比例均为5%,而在鹅肉、鸽子肉和兔肉中其最低检出比例为10%。本方法前处理简单,灵敏度高,重现性好,可作为高经济价值生鲜肉制品中掺假低经济价值肉类的有效检测方法。DNA min-barcoding methods based on cytochrome C oxidase subunitⅠ(COⅠ)sequence were developed to detect adulteration in 11 fresh meat products.DNA extraction was carried out with the DNeasy blood and tissue kit after samples were treated with vacuum freeze drying.The amplification of mini-barcoding region from DNA extracts was carried out using universal primer sets.Polymerase chain reaction(PCR)products were purified with DNA purification kit and analyzed by clone sequencing.The sequencing results were queried in GenBank(http://blast.ncbi.nlm.nih.gov/Blast.cgi)using the Blast align.The primer set COⅠ-A was found to be optimal,showing 100%amplification efficiency in 11 fresh meats(including pork,beef,mutton,chicken,duck,pigeon meat,horseflesh,ass meat,goose,rabbit meat and rat meat)with optimized PCR procedure.Eighteen meat adulteration models were founded to test the minimum detected adulteration rate by low economic value meat,and the results showed that the minimum detected adulteration rates of beef and mutton were 5%,and the rates of pigeon meat,goose,and rabbit meat were 10%,when added by the six low economic value meats (including pork, chicken, duck, horseflesh, ass meat and rat meat). To sum up, the method is simple, sensitive andreproducible, which is suitable for the detection of adulteration in the 11 fresh meat products.

关 键 词：DNA微条形码 肉制品 细胞色素C氧化酶亚基Ⅰ序列 掺假 

分 类 号：TS251.7[轻工技术与工程—农产品加工及贮藏工程]