A novel isothermal detection method for the universal element of genetically modified soybean

Abstract

Due to the increasingly heated debate on the potential threat of genetically modified (GM) crops to human health and environment, regulations and laws relevant to GM crops have been issued in many countries and regions to strictly restrict their cultivation and application. Therefore, fast and accurate method to realized on-site detection of GM crops is greatly demanded. In this work, a novel isothermal amplification method termed denaturation bubble-mediated strand exchange amplification (SEA) was proposed first time to detect GM crops. Fluorescence assay based on SEA could accurately distinguished GM and non-GM soybean by detecting agrobacterium tumefaciens nopaline synthase (NOS) terminator, which was widely incorporated in GM crops. Moreover, this feasible and specific method could detect NOS terminator from as low as 200 pg/μL total genomic DNA of GM soybean. In addition, in the actual sample detection, the result of colorimetric assay based on SEA results could be directly observed by the naked eyes within 58 min. Compared with the traditional methods based on PCR, which normally required complex equipment, skilled technicians and long operation time, this simple, fast and precise method is more desirable for the on-site GM crops detection.

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Data availability

We make sure that all data and materials support our published claims and comply with field standards.

Abbreviations

GM:

Genetically modified

NOS:

Nopaline synthase

FDA:

Food and Drug Administration

US EPA:

United States Environmental Protection Agency

USDA:

United States Department of Agriculture

PCR:

Polymerase chain reaction

SEA:

Strand exchange amplification

LAMP:

Loop-mediated isothermal amplification

RCA:

Rolling-circle amplification

RPA:

Recombinase polymerase amplification

DNA:

Deoxyribonucleic acid

HPLC:

High performance liquid chromatography

NCBI:

National Center for Biotechnology Information

dNTPs:

Deoxy-ribonucleoside triphosphates

NTC:

No template control

PAGE:

Polyacrylamide gel electrophoresis

Tt:

Threshold time

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Acknowledgments

The authors sincerely thank Qingdao Grain and Oils Quality Inspection Center for GM bean samples. This work was supported by National Natural Science Foundation of China (No.31670868 and No.21675094), and Shandong Province Natural Science Fund Major Basic Research Project (ZR2017ZC0123).

Funding

This work was funded by National Natural Science Foundation of China (No.31670868 and No.21675094), and Shandong Province Natural Science Fund Major Basic Research Project (ZR2017ZC0123).

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Contributions

Yongxiang Liu and Yang Li performed the experiments; Yongxiang Liu, Yang Li and Mengzhe Li analyzed the data; Cuiping Ma and Chao Shi designed the study; Hongyuan Sun and Qingguo Huo provided the GM bean samples; Yang Li and Yongxiang Liu wrote the manuscript; and all authors contributed to the writing of the paper, had primary responsibility for the final content, and read and approved the final manuscript.

Corresponding author

Correspondence to Chao Shi.

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Liu, Y., Li, Y., Li, M. et al. A novel isothermal detection method for the universal element of genetically modified soybean. Biologia (2020). https://doi.org/10.2478/s11756-020-00541-8

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Keywords

  • Isothermal nucleic acid amplification
  • SEA
  • Colorimetric detection
  • GM soybean
  • On-site detection