CN101875976A - Triple PCR-RFLP method for identifying meat species source of yak jerky - Google Patents
Triple PCR-RFLP method for identifying meat species source of yak jerky Download PDFInfo
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- CN101875976A CN101875976A CN 201010226704 CN201010226704A CN101875976A CN 101875976 A CN101875976 A CN 101875976A CN 201010226704 CN201010226704 CN 201010226704 CN 201010226704 A CN201010226704 A CN 201010226704A CN 101875976 A CN101875976 A CN 101875976A
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Abstract
The invention discloses a triple PCR-RFLP method for quickly identifying yak meat, scalper meat and buffalo meat, which belongs to the field of molecular biological detections, and can be used for tracing the source of a yak jerky product and quickly identifying species components of the yak jerky product. The method overcomes the defects of low accuracy and the like of the current method for identifying the yak meat, the scalper meat and the buffalo meat, and provides the PCR-RFLP method for accurately identifying the yak meat, the scalper meat and the buffalo meat. The method can identify the yak meat, the scalper meat and the buffalo meat by the following steps of: extracting the DNA of a sample; adopting a universal primer for amplification and gel electrophoresis for detection; and using HgaI, ApaI and PflMI restriction endonuclease to perform synchronous enzyme digestion and PCR-RFLP analysis to ensure that the yak meat, the scalper meat and the buffalo meat are identified quickly. The PCR-RFLP method has the characteristics of simplicity, convenience, quickness, low cost and high accuracy.
Description
Technical field
The invention belongs to the molecular Biological Detection field, the triple PCR-RFLP that relates to Carnis Bovis grunniens, Carnis Bovis seu Bubali and buffalo meat differentiates fast.
Technical background
Yak is China's distinctive ox kind in Qinghai-Tibet Platean, and its meat is good, delicious, pollution-free, is natural green food.Carnis Bovis grunniens ranks the meat first with high protein and lower fat, and the Carnis Bovis grunniens product is also promptly enlarging in the occupation rate in market, and there is gap greatly in price with other beef.Therefore, a lot of phenomenons with Carnis Bovis seu Bubali (comprising zebu and common ox) and buffalo meat personation Carnis Bovis grunniens product have appearred on the market.In order to ensure consumers in general's rights and interests,, need differentiate fast and accurately Carnis Bovis grunniens, Carnis Bovis seu Bubali and buffalo meat for the law enfrocement official provides technical support.
When fresh meat was carried out the sense organ discriminating, according to the difference of meat at aspects such as color, smell, flavour and fiber thickness, by " see, hear, touch ", comprehensive three judgements that main points draw were carried out sense organ to the meat source and are differentiated.There are the following problems in actual applications for this: owing to be beef all, be difficult to accurately differentiate by the sense organ means.And immunology detection is based on the immune response of soluble antigen and antibody, uses interspecies specific antigen and carries out the method for design studies, but utilize this method, because sample can influence the detection accuracy of ELISA method through the heat treated of differing temps.Particularly after sample was handled through the high temperature deep processing, sex change had taken place in albumen, has changed the specific antigen determinant, therefore was difficult to differentiate and distinguish different animal species.The test kit sensitivity that is used for animal derived detection in addition at present is lower, also is unsuitable for promoting the use of.
And the PCR-RFLP technology is quick, easy, cheap, existing relevant report aspect the kind discriminating of meat product, but do not see Carnis Bovis grunniens is arranged, the report of triple PCR-RFLP aspect that Carnis Bovis seu Bubali and buffalo meat are differentiated fast.
Summary of the invention
The objective of the invention is to solve meat kind source discriminating in the yak jerky, triple PCR-RFLP the method that provides a kind of quick Carnis Bovis grunniens, Carnis Bovis seu Bubali and buffalo meat to differentiate is for the malpractice of hitting with Carnis Bovis seu Bubali (comprising zebu and common ox) and buffalo meat personation yak jerky provides technical support.
Carnis Bovis grunniens, Carnis Bovis seu Bubali and buffalo meat are differentiated by the following method: add about 100mg sample in (1) 1.5mL centrifuge tube, shred, add 200 μ L TE solution (pH 8.0), the vortex mixing with scissors; (2) add 400 μ L lysates, vortex mixing; (3) add 500 μ L phenol: chloroform: primary isoamyl alcohol (25: 24: 1), thermal agitation, the centrifugal 10min of 12000rpm; (4) get supernatant liquor, add the equal-volume chloroform: primary isoamyl alcohol (24: 1), thermal agitation, the centrifugal 10min of 12000rpm; (5) get supernatant liquor, add the equal-volume chloroform, thermal agitation, the centrifugal 10min of 12000rpm; (6) get supernatant liquor, add 0.8 times of volume Virahol, the centrifugal 10min of 12000rpm; (7) abandon supernatant liquor, collect DNA, then get 70% ethanol and clean 1 time, again with seasoning under its room temperature; (8) add the DNA that 100 μ L TE (pH 8.0) dissolving is extracted, at last with its-20 ℃ of preservations with standby.(9) pcr amplification and detection: with general upstream primer the F:5 '-TAC CAT GAG GAC AAATAT CAT TCTG-3 ' and general downstream primer the R:5 '-CCT CCT AGT TTG TTA GGG ATT GAT CG-3 ' of all vertebrates Cytb genes that can increase.PCR reaction cumulative volume is 25 μ L, 10 * PCR Buffer, 2.5 μ L wherein, Taq archaeal dna polymerase 0.2 μ L (2.5U/ μ L), dNTPs (2.5mmol/L) 2 μ L, each 2 μ L of the upstream and downstream primer of 10pmol/ μ L, template DNA 2 μ L, sterile purified water is supplied cumulative volume.Amplification condition is: 95 ℃ of sex change 10min, 95 ℃ of sex change 45s, 53 ℃ of annealing 60s, 72 ℃ are extended 60s, after 35 circulations again 72 ℃ continue to extend 7min.Get 4 μ L PCR products and mix with 1 μ L, 6 * loadding buffer, adopt the TAE buffering system, 2% sepharose (adding staining agent Gold view I) about 25min under 5v/cm constant voltage deposition condition observes and record in gel imaging system.(10) get the PCR product 7 μ L of Cytb gene fragment, add each 0.5 μ L of three kinds of restriction enzymes of HgaI, ApaI and PflMI then successively, NEB buffer 2.5 μ L, ddH
209 μ L, 37 ℃ of temperature are bathed 3h.Enzyme is cut product and is detected with 2% agarose gel electrophoresis.Judge according to restriction enzyme mapping then: HgaI is cut into 220bp and 252bp to yak sample enzyme, and ApaI is cut into 173bp and 299bp to common ox sample enzyme, and PflMI is cut into 69bp and 403bp to buffalo sample enzyme.
The present invention adopts the PCR-RFLP method in discrimination process, have easy, quick, cheapness and the high characteristics of accuracy.
Description of drawings
Fig. 1 is the gel electrophoresis figure that step in the embodiment one (9) Carnis Bovis grunniens (1,2), Carnis Bovis seu Bubali (3,4) and buffalo meat (5,6) product amplified production detect; Fig. 2 is that the PCR product that step in the embodiment one (10) is cut step (9) in the embodiment one with triple enzymes carries out the gel electrophoresis figure that PCR-RFLP analyzes the back detection: M:Marker I, 1: control group, 2: Carnis Bovis grunniens, 3: Carnis Bovis seu Bubali, 4: buffalo meat, 5: yak, ox and buffalo mix the meat sample; 6-11: yak jerky.
Embodiment
Embodiment one: Carnis Bovis grunniens, Carnis Bovis seu Bubali and buffalo meat are differentiated by the following method: add about 100mg sample in (1) 1.5mL centrifuge tube, shred, add 200 μ L TE solution (pH 8.0), the vortex mixing with scissors; (2) add 400 μ L lysates, vortex mixing; (3) add 500 μ L phenol: chloroform: primary isoamyl alcohol (25: 24: 1), thermal agitation, the centrifugal 10min of 12000rpm; (4) get supernatant liquor, add the equal-volume chloroform: primary isoamyl alcohol (24: 1), thermal agitation, the centrifugal 10min of 12000rpm; (5) get supernatant liquor, add the equal-volume chloroform, thermal agitation, the centrifugal 10min of 12000rpm; (6) get supernatant liquor, add 0.8 times of volume Virahol, the centrifugal 10min of 12000rpm; (7) abandon supernatant liquor, collect DNA, then get 70% ethanol and clean 1 time, again with seasoning under its room temperature; (8) add the DNA that 100 μ L TE (pH 8.0) dissolving is extracted, at last with its-20 ℃ of preservations with standby.(9) pcr amplification and detection: with general upstream primer the F:5 '-TAC CAT GAG GAC AAATAT CAT TCT G-3 ' and general downstream primer the R:5 '-CCT CCTAGT TTG TTA GGG ATT GAT CG-3 ' of all vertebrates Cytb genes that can increase.PCR reaction cumulative volume is 25 μ L, 10 * PCR Buffer, 2.5 μ L wherein, Taq archaeal dna polymerase 0.2 μ L (2.5U/ μ L), dNTPs (2.5mmol/L) 2 μ L, each 2 μ L of the upstream and downstream primer of 10pmol/ μ L, template DNA 2 μ L, sterile purified water is supplied cumulative volume.Amplification condition is: 95 ℃ of sex change 10min, 95 ℃ of sex change 45s, 53 ℃ of annealing 60s, 72 ℃ are extended 60s, after 35 circulations again 72 ℃ continue to extend 7min.Get 4 μ L PCR products and mix with 1 μ L, 6 * loadding buffer, adopt the TAE buffering system, 2% sepharose (adding staining agent Gold view I) about 25min under 5v/cm constant voltage deposition condition observes and record in gel imaging system.(10) get the PCR product 7 μ L of Cytb gene fragment, add each 0.5 μ L of three kinds of restriction enzymes of HgaI, ApaI and PflMI then successively, NEB buffer 2.5 μ L, ddH
209 μ L, 37 ℃ of temperature are bathed 3h.Enzyme is cut product and is detected with 2% agarose gel electrophoresis.Judge according to restriction enzyme mapping then: HgaI is cut into 220bp and 252bp to yak sample enzyme, and ApaI is cut into 173bp and 299bp to common ox sample enzyme, and PflMI is cut into 69bp and 403bp to buffalo sample enzyme.
Claims (3)
1. triple PCR-RFLP method of differentiating fast Carnis Bovis grunniens, Carnis Bovis seu Bubali and buffalo meat, it is characterized in that and to differentiate Carnis Bovis grunniens, Carnis Bovis seu Bubali and buffalo meat by the following method: add about 100mg sample in (1) 1.5mL centrifuge tube, shred with scissors, add 200 μ LTE solution (pH 8.0), the vortex mixing; (2) add 400 μ L lysates, vortex mixing; (3) add 500 μ L phenol: chloroform: primary isoamyl alcohol (25: 24: 1), thermal agitation, the centrifugal 10min of 12000rpm; (4) get supernatant liquor, add the equal-volume chloroform: primary isoamyl alcohol (24: 1), thermal agitation, the centrifugal 10min of 12000rpm; (5) get supernatant liquor, add the equal-volume chloroform, thermal agitation, the centrifugal 10min of 12000rpm; (6) get supernatant liquor, add 0.8 times of volume Virahol, the centrifugal 10min of 12000rpm; (7) abandon supernatant liquor, collect DNA, then get 70% ethanol and clean 1 time, again with seasoning under its room temperature; (8) add the DNA that 100 μ LTE (pH 8.0) dissolving is extracted, at last with its-20 ℃ of preservations with standby.(9) pcr amplification and detection: with general upstream primer the F:5 '-TAC CAT GAG GAC AAA TAT CAT TCT G-3 ' and general downstream primer the R:5 '-CCT CCT AGT TTG TTA GGG ATT GAT CG-3 ' of all vertebrates Cytb genes that can increase.PCR reaction cumulative volume is 25 μ L, 10 * PCRBuffer, 2.5 μ L wherein, Taq archaeal dna polymerase 0.2 μ L (2.5U/ μ L), dNTPs (2.5mmol/L) 2 μ L, each 2 μ L of the upstream and downstream primer of 10pmol/ μ L, template DNA 2 μ L, sterile purified water is supplied cumulative volume.Amplification condition is: 95 ℃ of sex change 10min, 95 ℃ of sex change 45s, 53 ℃ of annealing 60s, 72 ℃ are extended 60s, after 35 circulations again 72 ℃ continue to extend 7min.Get 4 μ LPCR products and mix with 1 μ L, 6 * loadding buffer, adopt the TAE buffering system, 2% sepharose (adding staining agent Gold view I) about 25min under 5v/cm constant voltage deposition condition observes and record in gel imaging system.(10) get the PCR product 7 μ L of Cytb gene fragment, add each 0.5 μ L of three kinds of restriction enzymes of HgaI, ApaI and PflMI then successively, NEB buffer 2.5 μ L, ddH
209 μ L, 37 ℃ of temperature are bathed 3h.Enzyme is cut product and is detected with 2% agarose gel electrophoresis.Judge according to restriction enzyme mapping then: HgaI is cut into 220bp and 252bp to yak sample enzyme, and ApaI is cut into 173bp and 299bp to common ox sample enzyme, and PflMI is cut into 69bp and 403bp to buffalo sample enzyme.
2. a kind of PCR-RFLP method of differentiating Carnis Bovis grunniens, Carnis Bovis seu Bubali and buffalo meat according to claim 1 is characterized in that the amplimer in the step 8 is:
General upstream primer F:5 '-TAC CAT GAG GAC AAA TAT CAT TCT G-3 '
General downstream primer R:5 '-CCT CCT AGT TTG TTA GGG ATT GAT CG-3 '
3. a kind of triple PCR-RFLP method of differentiating Carnis Bovis grunniens, Carnis Bovis seu Bubali and buffalo meat according to claim 1 is characterized in that step (10) carries out PCR-RFLP synchronously with HgaI, ApaI and three restriction enzymes of PflMI and analyze.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102559919A (en) * | 2012-02-29 | 2012-07-11 | 上海出入境检验检疫局动植物与食品检验检疫技术中心 | Real-time PCR (Polymerase Chain Reaction) detection method of buffalo components in food and feed |
| CN103397101A (en) * | 2013-08-22 | 2013-11-20 | 山东省农业科学院生物技术研究中心 | Fluorescently-labeled gene multiplex amplification method for identifying sources of goat, sheep, pig and duck meat simultaneously |
| CN105132526A (en) * | 2015-06-25 | 2015-12-09 | 西南民族大学 | Detection method for identification of yak meat authenticity |
| CN107858442A (en) * | 2017-11-15 | 2018-03-30 | 四川省轻工业研究设计院 | Yak, the detection primer and method of three kinds of derived components of ox and buffalo in a kind of beef food |
| CN109266752A (en) * | 2017-07-17 | 2019-01-25 | 中国科学院成都生物研究所 | It is a kind of for detecting the kit and method of yak derived component |
| US10986816B2 (en) | 2014-03-26 | 2021-04-27 | Scr Engineers Ltd. | Livestock location system |
| US10986817B2 (en) | 2014-09-05 | 2021-04-27 | Intervet Inc. | Method and system for tracking health in animal populations |
| US11071279B2 (en) | 2014-09-05 | 2021-07-27 | Intervet Inc. | Method and system for tracking health in animal populations |
| USD990063S1 (en) | 2020-06-18 | 2023-06-20 | S.C.R. (Engineers) Limited | Animal ear tag |
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Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101712996A (en) * | 2009-12-14 | 2010-05-26 | 中国科学院昆明动物研究所 | Method for quickly identifying categories of meat and dried meat products of five domestic animals |
-
2010
- 2010-07-15 CN CN 201010226704 patent/CN101875976A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101712996A (en) * | 2009-12-14 | 2010-05-26 | 中国科学院昆明动物研究所 | Method for quickly identifying categories of meat and dried meat products of five domestic animals |
Non-Patent Citations (4)
| Title |
|---|
| 《Molecular Ecology Notes》 20031231 S.K.VERMA等 Novel universal primers establish identity of an enormous number of animal species for forensic application 第28-31页 1-3 第3卷, 2 * |
| 《中国学术文献网络出版总库》 20100601 何建文 牦牛肉产品的分子追溯研究 1-3 , 2 * |
| 《西北农林科技大学学报》 20070228 蔡欣等 中国3个牛种cytb基因多态性及其系统发育研究 第43-46及第52页 1-3 第35卷, 第2期 2 * |
| 《食品科学》 20071231 胡强等 用通用引物扩增细胞色素b基因进行牦牛肉的鉴定 第319-322页 1-3 第28卷, 第11期 2 * |
Cited By (16)
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| CN102559919A (en) * | 2012-02-29 | 2012-07-11 | 上海出入境检验检疫局动植物与食品检验检疫技术中心 | Real-time PCR (Polymerase Chain Reaction) detection method of buffalo components in food and feed |
| CN103397101A (en) * | 2013-08-22 | 2013-11-20 | 山东省农业科学院生物技术研究中心 | Fluorescently-labeled gene multiplex amplification method for identifying sources of goat, sheep, pig and duck meat simultaneously |
| CN103397101B (en) * | 2013-08-22 | 2016-01-13 | 山东省农业科学院生物技术研究中心 | A kind of fluorescent marker gene composite amplification method simultaneously identifying goat, sheep, pig and duck source property |
| US11963515B2 (en) | 2014-03-26 | 2024-04-23 | S.C.R. (Engineers) Limited | Livestock location system |
| US10986816B2 (en) | 2014-03-26 | 2021-04-27 | Scr Engineers Ltd. | Livestock location system |
| US10986817B2 (en) | 2014-09-05 | 2021-04-27 | Intervet Inc. | Method and system for tracking health in animal populations |
| US11071279B2 (en) | 2014-09-05 | 2021-07-27 | Intervet Inc. | Method and system for tracking health in animal populations |
| CN105132526A (en) * | 2015-06-25 | 2015-12-09 | 西南民族大学 | Detection method for identification of yak meat authenticity |
| CN109266752A (en) * | 2017-07-17 | 2019-01-25 | 中国科学院成都生物研究所 | It is a kind of for detecting the kit and method of yak derived component |
| CN107858442A (en) * | 2017-11-15 | 2018-03-30 | 四川省轻工业研究设计院 | Yak, the detection primer and method of three kinds of derived components of ox and buffalo in a kind of beef food |
| US11832584B2 (en) | 2018-04-22 | 2023-12-05 | Vence, Corp. | Livestock management system and method |
| US11864529B2 (en) | 2018-10-10 | 2024-01-09 | S.C.R. (Engineers) Limited | Livestock dry off method and device |
| USD990062S1 (en) | 2020-06-18 | 2023-06-20 | S.C.R. (Engineers) Limited | Animal ear tag |
| US11832587B2 (en) | 2020-06-18 | 2023-12-05 | S.C.R. (Engineers) Limited | Animal tag |
| USD990063S1 (en) | 2020-06-18 | 2023-06-20 | S.C.R. (Engineers) Limited | Animal ear tag |
| US11960957B2 (en) | 2020-11-25 | 2024-04-16 | Identigen Limited | System and method for tracing members of an animal population |
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