Uji Deteksi Adulterasi Daging Babi (Sus scrofa domestica) pada Bakso Metode Loop-Mediated Isothermal Amplification


  • Rahma Micho Widyanto Department of Nutrition, Faculty of Medicine, Universitas Brawijaya
  • Cleonara Yanuar Dini
  • Irma Sarita Rahmawati
  • Sekar Ramadhanti Putri
  • Amelia Nurdini Rozana
  • Sakinah Hilya Abida
  • Yunimar Yunimar




adulteration, loop-mediated isothermal amplification, pork, mitochondrial DNA, halal


Composition adulteration in a product is still a serious problem. For that we need a DNA-based test method that is fast, specific and simple. In this study, a rapid test technique with the loop-mediated isothermal amplification method was applied to identify the presence of pork DNA in meatballs. Six primers were used for the identification of mitochondrial DNA cytochrome oxidase subunit II. Samples were prepared in the form of mixed beef meatballs containing pork with a concentration of 50; 5; 0.5; 0.05%, respectively. The results showed that the optimal concentration for primers was 4 pmol inner primers, 2 pmol loop primers, and 1 pmol outer primers. The LAMP reaction showed a change in color in the meatball sample with the pork mixture to the lowest concentration. The LAMP technique provides a faster test with the final result of on-the-spot color visualization so that it is potential to be developed as a quick detection kit for identification of halal food processed products


Ulca P, Balta H, Çaǧin I, Senyuva HZ. Meat species identification and halal authentication using pcr analysis of raw and cooked traditional turkish foods. Meat Sci. 2013; 94(3):280–4.

Cawthorn DM, Steinman HA, Hoffman LC. A high incidence of species substitution and mislabelling detected in meat products sold in South Africa. Food Control. 2013; 32(2):440–9.

Doosti A, Ghasemi Dehkordi P, Rahimi E. Molecular assay to fraud identification of meat products. J Food Sci Technol. 2014; 51(1):148–52.

Aida AA, Man YBC, Wong CMVL, Raha AR, Son R. Analysis of raw meats and fats of pigs using polymerase chain reaction for Halal authentication. Meat Sci. 2005; 69(1):47–52.

Ali ME, Hashim U, Mustafa S, Che Man YB, Yusop MHM, Kashif M, et al. Nanobiosensor for detection and quantification of DNA sequences in degraded mixed meats. J Nanomater. 2011; 2011:1–11.

Farrokhi R, Jafari Joozani R. Identification of pork genome in commercial meat extracts for Halal authentication by SYBR green I real-time PCR. Int J Food Sci Technol. 2011; 46:951–5.

Ali ME, Hashim U, Mustafa S, Che Man YB, Dhahi TS, Kashif M, et al. Analysis of pork adulteration in commercial meatballs targeting porcine-specific mitochondrial cytochrome b gene by TaqMan probe real-time polymerase chain reaction. Meat Sci. 2012; 91(4):454–9.

Notomi T, Okayama H, Masubuchi H, Yonekawa T, Watanabe K, Amino N, et al. Loop-mediated isothermal amplification of DNA. Nucleic Acids Res. 2000; 28(12):E63.

Tomita N, Mori Y, Kanda H, Notomi T. Loop-mediated isothermal amplification (LAMP) of gene sequences and simple visual detection of products. Nat Protoc. 2008; 3(5):877–82.

Dooley JJ, Paine KE, Garrett SD, Brown HM. Detection of meat species using TaqMan real-time PCR assays. Meat Sci. 2004; 68:431–8.

Dawnay N, Ogden R, McEwing R, Carvalho GR, Thorpe RS. Validation of the barcoding gene COI for use in forensic genetic species identification. Forensic Sci Int. 2007; 173(1):1–6.

Kesmen Z, Yetiman AE, Åžahin F, Yetim H. Detection of Chicken and Turkey Meat in Meat Mixtures by Using Real-Time PCR Assays. J Food Sci. 2012; 77:C167-173.

Cho AR, Dong HJ, Cho S. Meat species identification using loop-mediated isothermal amplification assay targeting species-specific mitochondrial DNA. Korean J Food Sci Anim Resour. 2014; 34(6):799–807.

Firahmi F, Dharmawati S, Aldrin M. Sifat fisik dan organoleptik bakso yang dibuat dari daging sapi dengan lama pelayuan berbeda. J Al Ulum Sains dan Teknol. 2015; 1(1).

Ran G, Ren L, Han X, Liu X, Li Z, Pang D, et al. Development of a Rapid Method for the Visible Detection of Pork DNA in Halal Products by Loop-Mediated Isothermal Amplification. Food Anal Methods. 2016; 9:565–70.

Koh B-R-D, Kim J-Y, Na H-M, Park S-D, Kim Y-H. Development of species-specific multiplex PCR assays of mitochondrial 12S rRNA and 16S rRNA for the identification of animal species. Korean J Vet Serv. 2011; 34(4):417–28.

Avise JC, Arnold J, Ball RM, Bermingham E, Lamb T, Neigel JE, et al. Intraspecific Phylogeography: The Mitochondrial DNA Bridge Between Population Genetics and Systematics. Annu Rev Ecol Syst. 1987; 18(1):489–522.

Koetsier G, Cantor E. A practical guide to analyzing nucleic acid concentration andpurity with microvolume spectrophotometers. New England BioLabs. Inc.: Ipswich,MA, USA; 2019,

Gadkar VJ, Goldfarb DM, Gantt S, Tilley PAG. Real-time Detection and Monitoring of Loop Mediated Amplification (LAMP) Reaction Using Self-quenching and De-quenching Fluorogenic Probes. Sci Rep. 2018; 8(1).

Jenkins DM, Kubota R, Dong J, Li Y, Higashiguchi D. Handheld device for real-time, quantitative, LAMP-based detection of Salmonella enterica using assimilating probes. Biosens Bioelectron. 2011; 30(1):255–60.

R. Kubota, A. M. Alvarez, W.W. Su, D. M. Jenkins. FRET-Based Assimilating Probe for Sequence-Specific Real-Time Monitoring of Loop-Mediated Isothermal Amplification (LAMP). Biol Eng Trans. 2011; 4(2):81–100.

Thiessen LD, Keune JA, Neill TM, Turechek WW, Grove GG, Mahaffee WF. Development of a grower-conducted inoculum detection assay for management of grape powdery mildew. Plant Pathol. 2016; 65(2):238–49.

Cardoso TC, Ferrari HF, Bregano LC, Silva-Frade C, Rosa ACG, Andrade AL. Visual detection of turkey coronavirus RNA in tissues and feces by reverse-transcription loop-mediated isothermal amplification (RT-LAMP) with hydroxynaphthol blue dye. Mol Cell Probes. 2010; 24(6):415–7.

Fischbach J, Xander NC, Frohme M, Glökler JF. Shining a light on LAMP assays- a comparison of LAMP visualization methods including the novel use of berberine. Biotechniques. 2015; 58(4):189–94.

Haines AM, Tobe SS, Kobus HJ, Linacre A. Properties of nucleic acid staining dyes used in gel electrophoresis. Electrophoresis. 2015; 36(6):941–4.

Jensen MA, Fukushima M, Davis RW. DMSO and betaine greatly improve amplification of GC-rich constructs in de novo synthesis. PLoS One. 2010; 5(6):e11024.

Musso M, Bocciardi R, Parodi S, Ravazzolo R, Ceccherini I. Betaine, dimethyl sulfoxide, and 7-deaza-dGTP, a powerful mixture for amplification of GC-rich DNA sequences. J Mol Diagnostics. 2006; 8(5):544–50.

Yeh HY, Shoemaker CA, Klesius PH. Evaluation of a loop-mediated isothermal amplification method for rapid detection of channel catfish Ictalurus punctatus important bacterial pathogen Edwardsiella ictaluri. J Microbiol Methods. 2005; 63(1):36–44.

Zhou D, Guo J, Xu L, Gao S, Lin Q, Wu Q, et al. Establishment and application of a loop-mediated isothermal amplification (LAMP) system for detection of cry1Ac transgenic sugarcane. Sci Rep. 2014; 4(1).

Chen R, Tong Q, Zhang Y, Lou D, Kong Q, Lv S, et al. Loop-mediated isothermal amplification: Rapid detection of Angiostrongylus cantonensis infection in Pomacea canaliculata. Parasites and Vectors. 2011; 4(1):204.

Jevtuševskaja J, Krõlov K, Tulp I, Langel Ü. The effect of main urine inhibitors on the activity of different DNA polymerases in loop-mediated isothermal amplification. Expert Rev Mol Diagn. 2017; 17(4):403–10.

Dhama K, Karthik K, Chakraborty S, Tiwari R, Kapoor S, Kumar A, et al. Loop-mediated isothermal amplification of DNA (LAMP): A new diagnostic tool lights the world of diagnosis of animal and human pathogens: A review. Pakistan J Biol Sci. 2014; 17(2):151–66.

Woniakowski G, Kozdruń W, Samorek-Salamonowicz E. Loop-mediated isothermal amplification for the detection of goose circovirus. Virol J. 2012; 9(1):110.

Karthik K, Rathore R, Thomas P, Arun TR, Viswas KN, Dhama K, et al. New closed tube loop mediated isothermal amplification assay for prevention of product cross-contamination. MethodsX. 2014; 1:137–43.

Tao ZY, Zhou HY, Xia H, Xu S, Zhu HW, Culleton RL, et al. Adaptation of a visualized loop-mediated isothermal amplification technique for field detection of Plasmodium vivax infection. Parasites and Vectors. 2011; 4(1):115.

Njiru ZK, Mikosza ASJ, Armstrong T, Enyaru JC, Ndung’u JM, Thompson ARC. Loop-mediated isothermal amplification (LAMP) method for rapid detection of Trypanosoma brucei rhodesiense. PLoS Negl Trop Dis. 2008; 2(2):e147.

Barakat H, El-Garhy HAS, Moustafa MMA. Detection of pork adulteration in processed meat by species-specific PCR-QIAxcel procedure based on D-loop and cytb genes. Appl Microbiol Biotechnol. 2014; 98:9805–16.

Dincer B, Spearow JL, Cassens RG, Greaser ML. The effects of curing and cooking on the detection of species origin of meat products by competitive and indirect ELISA techniques. Meat Sci. 1987; 20:253–65.

Arslan A, Ilhak OI, Calicioglu M. Effect of method of cooking on identification of heat processed beef using polymerase chain reaction (PCR) technique. Meat Sci. 2006; 72(2):326–30.

Ran G, Ren L, Han X, Liu X, Li Z, Pang D, et al. Development of a Rapid Method for the Visible Detection of Pork DNA in Halal Products by Loop-Mediated Isothermal Amplification. Food Analytical Methods. 2015; 9(3):565–70.

Åžakalar E, Abasiyanik MF, Bektik E, Tayyrov A. Effect of Heat Processing on DNA Quantification of Meat Species. J Food Sci. 2012; 77(9):N40-44.