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Screening, Post-SELEX Optimization and Application of DNA Aptamers Specific for Tobramycin

  • Nandi ZhouEmail author
  • Rongfeng Cai
  • Xuyan Han
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 2070)

Abstract

Tobramycin (TOB) is an aminoglycoside antibiotic. The residue of TOB in animal-derived foods and environment will be harmful to human health, and therefore the specific detection of TOB residue in food and water is of great importance. Herein, through magnetic beads-based SELEX, overall 37 ssDNA aptamers specific for TOB were screened after ten rounds of selection. The affinity and specificity of these aptamers were evaluated, among which No. 32 aptamer (Ap 32) exhibits excellent performance. Then a post-SELEX optimization of Ap 32 was carried out based on rational design, through which a truncated aptamer with the length of 34 nucleotides (Ap 32-2) was identified as the best aptamer for TOB. Finally, the application of the screened aptamer was explored. A colorimetric assay of TOB was established based on the aptamer-modified gold nanoparticles (AuNPs). In the range from 100 to 1400 nM, the absorbance of AuNPs solution at 520 nm was linearly decreased with the increased concentration of TOB. The detection limit was estimated to be 37.9 nM. The assay was applied to detect TOB residue in honey samples.

Key words

Magnetic beads Single-stranded DNA aptamer Dissociation constant SELEX Truncation Tobramycin Gold nanoparticles Colorimetric detection 

Notes

Acknowledgments

This work was supported by the National Natural Science Foundation of China (no. 31271860).

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2020

Authors and Affiliations

  1. 1.The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of BiotechnologyJiangnan UniversityWuxiChina

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