Skip to main content

Aptamer-linked immobilized sorbent assay for detecting GMO marker, phosphinothricin acetyltransferase (PAT)

Molecular & Cellular Toxicology volume 16, pages 253–261 (2020)Cite this article

Abstract

Background

Development of genetically modified crops has rapidly increased in last few years. The most widely grown GM crops express genes that confer herbicide tolerance and insect resistance. Detection system of GM crops is important for safety evaluation before its consumption.

Objective

The purpose of this research is to detect GM crops, especially PAT, in food-samples.

Results

The bar gene (PAT protein, herbicide resistant) was cloned in pGEX-4T-1 and expressed by E. coli. The high-affinity PAT-specific single-stranded DNA (ssDNA) aptamers were obtained from a random DNA library. MOE docking study was performed to identify the potential binding region of the selected aptamers on PAT. Aptamer-linked immobilized sorbent assay (ALISA) method was used to detect PAT.

Conclusion

We screened aptamer against PAT for developing an efficient detection method. The selected PAT specific aptamers, HRPA-05 and HRPA-07, showed the distinct target binding behaviors, and detected PAT protein by aptamer-linked immobilized sorbent assay method with high efficiency and selectivity.

This is a preview of subscription content, access via your institution.

Access options

Buy single article

Instant access to the full article PDF.

USD 39.95

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

References

  • Block A et al (2013) The GMOseek matrix: a decision support tool for optimizing the detection of genetically modified plants. BMC Bioinform 14:256

    Article  Google Scholar 

  • Eigenmann PA, Burks AW, Bannon GA, Sampson HA (1996) Identification of unique peanut and soy allergens in sera adsorbed with cross-reacting antibodies. J Allergy Clin Immunol 98:969–978

    CAS  Article  Google Scholar 

  • Ermolli M et al (2006) Development of an innovative immunoassay for CP4EPSPS and Cry1AB genetically modified protein detection and quantification. Food Addit Contam 23:876–882

    CAS  Article  Google Scholar 

  • Gizzarelli F et al (2006) Evaluation of allergenicity of genetically modified soybean protein extract in a murine model of oral allergen-specific sensitization. Clin Exp Allergy 36:238–248

    CAS  Article  Google Scholar 

  • Hermann T, Patel DJ (2000) Adaptive recognition by nucleic acid aptamers. Science 287:820–825

    CAS  Article  Google Scholar 

  • Huber I et al (2013) Development and validation of duplex, triplex, and pentaplex real-time PCR screening assays for the detection of genetically modified organisms in food and feed. J Agric Food Chem 61:10293–10301

    CAS  Article  Google Scholar 

  • Ishii T, Araki M (2016) Consumer acceptance of food crops developed by genome editing. Plant Cell Rep 35:1507–1518

    CAS  Article  Google Scholar 

  • Kim HJ et al (2010) Expression of PAT and NPT II proteins during the developmental stages of a genetically modified pepper developed in Korea. J Agric Food Chem 58:10906–10910

    CAS  Article  Google Scholar 

  • Lee S-H et al (2015) Analytical bioconjugates, aptamers, enable specific quantitative detection of Listeria monocytogenes. Biosens Bioelectron 68:272–280

    CAS  Article  Google Scholar 

  • Lee E-H, Lim HJ, Lee S-D, Son A (2017) Highly sensitive detection of bisphenol A by NanoAptamer assay with truncated aptamer. ACS Appl Mater Interfaces 9:14889–14898

    CAS  Article  Google Scholar 

  • Lin C-H, Pan T-M (2016) Perspectives on genetically modified crops and food detection. J Food Drug Anal 24:1–8

    Article  Google Scholar 

  • McKeague M, Giamberardino A, DeRosa MC (2011) Environmental biosensors. IntechOpen, London

    Google Scholar 

  • Popek S, Halagarda M (2017) Genetically modified foods: consumer awareness, opinions and attitudes in selected EU countries. Int J Consum Stud 41:325–332

    Article  Google Scholar 

  • Sekhon SS et al (2017) Aptabody–aptatope interactions in aptablotting assays. Nanoscale 9:7464–7475

    CAS  Article  Google Scholar 

  • Shim Y, Shin W, Moon G, Kim K (2007) Quantitative analysis of phosphinothricin-N-acetyltransferase in genetically modified herbicide tolerant pepper by an enzyme-linked immunosorbent assay. J Microbiol Biotechnol 17:681

    CAS  PubMed  Google Scholar 

  • Shin W-R et al (2018a) Aptamer-based pathogen monitoring for Salmonella enterica ser. Typhimurium. J Biomed Nanotechnol 14:1992–2002

    CAS  Article  Google Scholar 

  • Shin W-R et al (2018b) Aptamer-based paper strip sensor for detecting Vibrio fischeri. ACS Comb Sci 20:261–268

    CAS  Article  Google Scholar 

  • Smyth SJ, Kerr WA, Phillips PW (2015) Global economic, environmental and health benefits from GM crop adoption. Glob Food Secur 7:24–29

    Article  Google Scholar 

  • Stoltenburg R, Reinemann C, Strehlitz B (2007) SELEX—a (r) evolutionary method to generate high-affinity nucleic acid ligands. Biomol Eng 24:381–403

    CAS  Article  Google Scholar 

  • Thompson CJ et al (1987) Characterization of the herbicide-resistance gene bar from Streptomyces hygroscopicus. EMBO J 6:2519–2523

    CAS  Article  Google Scholar 

  • Wilbanks B et al (2019) Optimization of a 40-mer antimyelin DNA aptamer identifies a 20-mer with enhanced properties for potential multiple sclerosis therapy. Nucleic Acid Ther 29:126–135

    CAS  Article  Google Scholar 

  • Xu W, Huang K, Zhao H, Luo Y (2005) Application of immunoaffinity column as cleanup tool for an enzyme linked immunosorbent assay of phosphinothricin-N-acetyltransferase detection in genetically modified maize and rape. J Agric Food Chem 53:4315–4321

    CAS  Article  Google Scholar 

Download references

Acknowledgements

This work was supported by the National Research Foundation of Korea (NRF) Grant funded by the Korea government (MEST) (no. NRF-2019R1A2C1010860).

Author information

Author notes

  1. Woo-Ri Shin and Mun-Jong Lee contributed equally to this work.

Authors and Affiliations

  1. School of Biological Sciences, Chungbuk National University, 1 Chungdae-Ro, Seowon-Gu, Cheongju, 28644, Republic of Korea

    Woo-Ri Shin, Mun-Jong Lee, Simranjeet Singh Sekhon, Ji-Young Ahn & Yang-Hoon Kim

  2. Department of Biological Sciences, Korea Advanced Institute of Science and Technology, 291 Daehak-ro, Yuseong-gu, Daejeon, 34141, Republic of Korea

    Ji Hun Kim, Sun Chang Kim & Byung-Kwan Cho

Authors
  1. Woo-Ri Shin
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Mun-Jong Lee
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Simranjeet Singh Sekhon
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Ji Hun Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Sun Chang Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Byung-Kwan Cho
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Ji-Young Ahn
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Yang-Hoon Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

Investigation and data analysis, WRS and MJL: experiments design and results analysis; JHK, SSS, JYA, YHK, SCK: Molecular docking analysis; BKC, YHK: All authors have read and agreed to the published version of the manuscript.

Corresponding authors

Correspondence to Byung-Kwan Cho, Ji-Young Ahn or Yang-Hoon Kim.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Shin, WR., Lee, MJ., Sekhon, S.S. et al. Aptamer-linked immobilized sorbent assay for detecting GMO marker, phosphinothricin acetyltransferase (PAT). Mol. Cell. Toxicol. 16, 253–261 (2020). https://doi.org/10.1007/s13273-020-00087-5

Download citation

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s13273-020-00087-5

Keywords

  • Genetically modified organism (GMO)
  • Phosphinothricin acetyltransferase (PAT)
  • ssDNA aptamer
  • Aptamer-linked immobilized sorbent assay (ALISA)