Abstract
Live cell-based SELEX (Systematic Evolution of Ligand EXponential enrichment) is a promising approach for identifying aptamers that can selectively bind to a cell-surface antigen or a particular target cell population. In particular, it offers a facile selection strategy for some special cell-surface proteins that are original glycosylated or heavily post-translationally modified, and are unavailable in their native/active conformation after in vitro expression and purification. In this chapter, we describe evolution of cell-type-specific RNA aptamers targeting the human CCR5 by combining the live cell-based SELEX strategy with high-throughput sequencing (HTS) and bioinformatics analysis.
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Acknowledgments
We thank James O. McNamara II and Paloma H. Giangrande for helpful discussions. We thank Mayumi Takahashi for providing primary PBMCs. The authors would like to thank City of Hope DNA sequencing core and Bioinformatics Core facility (Harry Gao, Xiwei Wu and Jinhui Wang) for Solexa Deep sequencing and data processing. The following reagents were obtained through the NIH AIDS Research and Reference Reagent Program, Division of AIDS, NIAID, NIH: the U373-Magi and U373-Magi-CCR5E cell lines were obtained from Dr. Michael Emerman [14]; CEM-NKr and CEM-NKr-CCR5 cell lines were obtained from Dr. Peter Cresswell [15, 16].
This work was supported by National Institutes of Health grants (grant numbers R01AI29329, R01AI42552, and R01HL07470) to J.J.R. Funding for open access charge: National Institutes of Health.
Conflict of Interest declaration: J.J.R. and J.Z. have an issued patent entitled “Cell-type-specific aptamer-siRNA delivery system for HIV-1 therapy”. USPTO, No. US 8, 222, 226 B2, issued date: July 17, 2012. J. J. R. and J. Z. have a patent pending on “Cell-specific internalizing RNA aptamers against human CCR5 and used therefore”, The United States Patent, and application number: 62/025,368, filed on July 16, 2014.
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Zhou, J., Rossi, J.J. (2016). Evolution of Cell-Type-Specific RNA Aptamers Via Live Cell-Based SELEX. In: Lin, RJ. (eds) RNA-Protein Complexes and Interactions. Methods in Molecular Biology, vol 1421. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3591-8_16
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DOI: https://doi.org/10.1007/978-1-4939-3591-8_16
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