牛肉真伪鉴别荧光RPA现场快检方法建立
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摘要
目的:为实现现场快速检测牛肉及其制品中牛DNA成分,建立基于荧光RPA技术的快速检测方法。方法:在细胞色素C氧化酶亚型Ⅰ中的黄牛、水牛、牦牛特异性DNA序列上设计RPA引物以及exo探针,对引物对进行组合筛选,结合核酸快速提取方法,建立荧光RPA快速检测方法,并对其特异性、灵敏度测试,通过方法比对实验验证其检测准确性。结果:所建立的检测方法具有很好的特异性,检测灵敏度低至1%(w/w),并且与标准方法检测结果一致。结论:研究成功建立了牛肉真伪鉴别荧光RPA现场快速检测方法。
Purpose: Construct beef authentication on-site rapid detection method based on fluorescent RPA technology, in order to realize rapid detection of bovine DNA components in beef and its products. Methods: Design RPA primer pairs and exo probe according to COI gene sequences specific for cow, buffalo and yak. Construct bovine authentication on-site rapid detection method through primer pairs screening, specificity and sensitivity test. Results: This method has good specificity, and the sensitivity can be reached 1%(w/w). The testing results were the same with standard methods. Conclusions: This study successfully construct beef authentication on-site rapid detection method based on fluorescent RPA technology.
Purpose: Construct beef authentication on-site rapid detection method based on fluorescent RPA technology, in order to realize rapid detection of bovine DNA components in beef and its products. Methods: Design RPA primer pairs and exo probe according to COI gene sequences specific for cow, buffalo and yak. Construct bovine authentication on-site rapid detection method through primer pairs screening, specificity and sensitivity test. Results: This method has good specificity, and the sensitivity can be reached 1%(w/w). The testing results were the same with standard methods. Conclusions: This study successfully construct beef authentication on-site rapid detection method based on fluorescent RPA technology.
引文
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[14]王耕,张薇,陈安东,等.肉类食品的动物物种基因鉴别及等温扩增荧光(IAFT)快速检测技术的建立[J].生物化工,2016,5(2):21-25
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[3]迪澳生物.大润发超市被曝光肉类掺假何时能休[J].食品安全导刊,2016,151(28):56-56
[4]汪子芳.猪肉+添加剂做成“牛肉干”老板被判15年[J].广西质量监督导报,2016,(5):35-35
[5]Kim T H,Park J,Kim C J,et al.Fully integrated lab-on-adisc for nucleic acid analysis of food-borne pathogens[J].Analytical Chemistry,2014,86(8):3841
[6]Lutz S,Weber P,Focke M,et al.Microfluidic lab-onafoil for nucleic acid analysis based on isothermal recombinase polymerase amplification(RPA)[J].Lab on AChip,2010,10(7):887
[7]Lorraine L,Dara L,Singhal M C,et al.Non-Instrumented Incubation of a Recombinase Polymerase Amplification Assay for the Rapid and Sensitive Detection of Proviral HIV-1 DNA[J].Plos One,2014,9(9):e108189
[8]Xu C,Li L,Jin W,et al.Recombinase Polymerase Amplification(RPA)of CaMV-35S Promoter and nos Terminator for Rapid Detection of Genetically Modified Crops[J].International Journal of Molecular Sciences,2014,15(10):18197-18205
[9]邓婷婷,黄文胜,程奇,等.重组酶聚合酶扩增技术检测转基因水稻中的Cry1Ab/c基因[J].中国食品学报,2015,15(3):187-193
[10]郭燕华,王德莲,王强,等.重组酶介导等温扩增技术快速检测牛肉及牛肉制品中的牛源性成分[J].食品安全质326量检测学报,2017,8(5):1745-1749
[11]郭燕华,陈遂,王德莲,等.基于重组酶等温扩增技术快速检测生鲜肉中猪源性成分[J].食品安全质量检测学报,2017,8(6):2012-2016
[12]吴昊,谢文佳,王艳丽,等.利用重组酶聚合酶扩增技术快速鉴定肉及肉制品中的猪源性成分[J].食品科技,2017,42(10):318-322
[13]李向丽,刘垚,谭贵良,等.基于LAMP法快速检测羊肉及其制品中的猪、鸭和羊源性成分[J].中国食品卫生杂志,2015,27(3):247-252
[14]王耕,张薇,陈安东,等.肉类食品的动物物种基因鉴别及等温扩增荧光(IAFT)快速检测技术的建立[J].生物化工,2016,5(2):21-25
[15]Piepenburg O,Williams C H,Stemple D L,et al.DNA detection using recombination proteins[J].Plos Biology,2006,7(4):e204