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Selection and Characterization of Cell Surface Specific Aptamer and Development of Fluorescence Assay for Detection of Shigella flexneri from Water Samples

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Abstract

The present study demonstrates, development of ssDNA aptamers against whole cell of S. flexneri employing a whole bacterium-based Systemic Evolution of Ligands by Exponential Enrichment (SELEX). After ten rounds of SELEX, cell surface specific aptamer pool was cloned, sequenced and divided based on sequence similarities and secondary structure. Binding affinity of FITC labelled aptamer from different group were carried out by flow cytometry analysis. The dissociation constant (Kd) values for specific and higher binder were evaluated to range from 144 to 329 nM. Six high binding aptamers with lower dissociation constant was chosen for selectivity study. Aptamer SHI 23, SHI 37 and SHI 42 showed higher selectivity towards S. flexneri in comparison with other related bacteria. Further applicability of selected aptamer was proven by fluorescence assay for convenience detection of target cell from spiked water sample and natural contaminated water samples. Altogether, aptamer generated in this study can be alternative DNA ligands for detection of S. flexneri compared to available ligands.

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Acknowledgments

The first author is indebted to Department of Science and Technology (DST, New Delhi, India), for financial assistance through women scientist program. The second author thankful to Department of Science and Technology (DST, New Delhi, India), for financial assistance through Inspire programme. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. The corresponding author and first author design, data collection and analysis, decision to publish, or preparation of the manuscript.

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  1. Microbiology Division, Defence Food Research Laboratory, Siddharthanagar, Mysore, Karnataka, 570011, India

    Padma Sudharani Lavu, Bhairab Mondal & Shylaja Ramlal

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  1. Padma Sudharani Lavu
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  2. Bhairab Mondal
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  3. Shylaja Ramlal
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Contributions

Dr. Padma Sudharani Lavu was involved in research designing, performing experiment, and manuscript writing. Bhairab Mondal was involved in research designing, performing experiment, data analysis, and manuscript writing. Dr. Shylaja Ramlal was involved in research designing, manuscript writing and correction of manuscript.

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Correspondence to Shylaja Ramlal.

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Lavu, P.S., Mondal, B. & Ramlal, S. Selection and Characterization of Cell Surface Specific Aptamer and Development of Fluorescence Assay for Detection of Shigella flexneri from Water Samples. J Fluoresc 31, 685–693 (2021). https://doi.org/10.1007/s10895-021-02691-7

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  • DOI: https://doi.org/10.1007/s10895-021-02691-7

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