• HARI WIDADA Post Graduate Program, Faculty of Pharmacy, Universitas Gadjah Mada, Yogyakarta, Indonesia 55281.
  • ABDUL ROHMAN Post Graduate Program, Faculty of Pharmacy, Universitas Gadjah Mada, Yogyakarta, Indonesia 55281.
  • RIRIS ISTIGHFARI JENIE Post Graduate Program, Faculty of Pharmacy, Universitas Gadjah Mada, Yogyakarta, Indonesia 55281.
  • SISMINDARI Post Graduate Program, Faculty of Pharmacy, Universitas Gadjah Mada, Yogyakarta, Indonesia 55281.


Objective: The objective of this study was to perform aptamer selection using systematic evolution of ligands by exponential enrichment (SELEX) method which assisted by graphene oxide against target of porcine gelatin (non-halal gelatin).

Methods: The aptamer selection was carried out using SELEX method without target immobilization. Selection of aptamer capable of binding porcine gelatin by applying grafen oxide (GO) was known as GO-SELEX. The selection process was initially carried out by incubation of single-stranded DNA (ssDNA) libraries targeting on porcine gelatin with the addition of graphene oxide. The selected ssDNA was then purified by several stages namely; symmetric PCR amplification, purification of products with DNA purification kits, asymmetric PCR amplification, and continued purification of DNA with native PAGE. The analysis of each stage was done by agarose gel electrophoresis.

Results: the results showed that aptamer targeting porcine DNA could be selected. This was indicated by the results of DNA analysis using native polyacrylamide gel electrophoresis (PAGE) in which sharp separation band with a base length equivalent to the marker of the ssDNA library (about 80 base pair) was obtained.

Conclusion: Aptamer targeting on porcine gelatin has been successfully developed using GO-SELEX method. GO can increase selectivity in developing aptamer which will be used as a biosensor to detect porcine gelatin. The method could be proposed as a standard of apatamer based method for porcine gelatin detection on halal products authentication.

Keywords: Graphene oxide, Aptamer, Porcine gelatin, GO-SELEX


1. Gibbs BF, Kermasha S, Alli I, Mulligan CN. Encapsulation in the food industry: a review. Int J Food Sci Nutr 1999;50:213-24.
2. Mondal N. The role of matrix tablet in drug delivery system. Int J Appl Pharm 2010;10:1-6.
3. Khatavkar UN, Kumar KJ, Shimpi SL. Novel approaches for the development of oral controlled release compositions of galantamine hydrobromide and paroxetine hydrochloride hemihydrate: a review. Int J Appl Pharm 2016;8:1-6.
4. Gomez Guillen MC, Perez Mateos M, Gomez Estaca J, Lopez Caballero E, Gimenez B, Montero P. Fish gelatin: a renewable material for the development of active biodegradable films. Trends Food Sci Technol 2009;20:3-16.
5. Ladislaus M, Kasankala YX, Weilong Y, Sun D, Hong Q. Optimization of gelatin extraction from grass carp (Catenopharyngodon idella) fish skin by response surface methodology. Biores Technol 2007;98:3338-43.
6. Murugaiah C, Noor ZM, Mastakim M, Bilung LM, Selamat J, Radu S. Meat species identification and Halal authentication analysis using mitochondrial DNA. Meat Sci 2014;83:57–61.
7. Hashim DM, Che Man YB, Norakasha R, Shuhaimi M, Salmah Y, Syahariza ZA. Potential use of fourier transform infrared spectroscopy for differentiation of bovine and porcine gelatins. Food Chem 2010;118:856-60.
8. Azilawati MI, Hashim DM, Jamilah B, Amin I. RP-HPLC method using 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate incorporated with normalization technique in the principal component analysis to differentiate the bovine, porcine and fish gelatins. Food Chem 2015;172:368-76.
9. Zhang GF, Liu T, Wang Q, Chen L, Lei JD, Luo J, et al. Mass spectrometric detection of marker peptides in tryptic digests of gelatin: a new method to differentiate between bovine and porcine gelatin. Food Hydrocoll 2009;23:2001–7.
10. Rodriguez MA, Garcia T, Gonzalez I, Hernandez PE, Martin R. TaqMan real-time PCR for the detection and quantitation of pork in meat mixtures. Meat Sci 2005;70:113–20.
11. Chen SY, Liu YP, Yao YG. Species authentication of commercial beef jerky based on PCR–RFLP analysis of the mitochondrial 12S rRNA gene. J Genetics Genom 2010;37:763–9.
12. Ghovvati S, Nassiri MR, Mirhoseini S, Moussavi AH, Javadmanesh A. Fraud identification in industrial meat products by multiplex PCR assay. Food Control 2009;20:696–9.
13. Man YBC, Aida AA, Raha AR, Son R. Identification of pork derivatives in food products by species-specific polymerase chain reaction (PCR) for halal. Food Control 2007;18:885-9.
14. Tuerk C, Gold L. Sistematic evolution of ligands by exponential enrichment: RNA ligands to bacteriophage t4 DNA polymerase. Science 1990;249:505–10.
15. Kim YS, dan GuMB. Advances in aptamer screening and small molecule aptasensors. Adv Biochem Eng Biotechnol 2014;140:29-67.
16. Wang Y. Nitrogen-doped graphene and its application in electrochemical biosensing. ACS Nano 2010;4:1790–8.
17. Varghese N, Mogera U, Govindaraj A, Das A, Maiti PK, Sood AK, et al. Binding of DNA nucleobases and nucleosides with graphene. Chem Phys 2009;10:206–10.
18. Shabani H, Mehdizadeh M, Mousavi SM, Dezfouli EA, Solgi T, Khodaverdi M, et al. Halal authenticity of gelatin using species-specific PCR. Food Chem 2015;184:203–6.
19. Pfeiffer F, Mayer G. Selection and biosensor application of aptamers for small molecules. Front Chem 2016;4:1-25.
20. Citartan M, Tang TH, Tan SC, Hoe CH, Saini R, Tominaga J, Gopinath SCB. Asymmetric PCR for good quality ssDNA generation towards DNA aptamer production. Songklanakarin J Sci Technol 2012;34:125-31.
21. Joeng CB, Niazi JH, Lee SJ, Gu MB. ssDNA aptamers that recognize diclofenac and 2-anilinophenylacetic acid. Biorg Med Chem 2009;17:5380–7.
22. Stoltenburg R, Reinemann C, Strehlitz B. SELEX—A (r) evolutionary method to generate high-affinity nucleic acid ligands. Biomol Eng 2007;24:381–403.
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