In Vitro Selection and Characterization of DNA Aptamers to a Small Molecule Target

Ruscito, Annamaria; McConnell, Erin M.; Koudrina, Anna; Velu, Ranganathan; Mattice, Christopher; Hunt, Vernon; McKeague, Maureen; DeRosa, Maria C.

doi:10.1002/cpch.28

Cited by 21 publications

(19 citation statements)

References 87 publications

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“…While this is a common phenomenon with SELEX, it may be that the primer binding regions of Malaria.1, which were truncated for the analysis, were required. Indeed, nucleotides in the primer regions are occasionally involved in binding, but often they have little or no effect 30 . Nevertheless, the Malaria.1 sequence tested has several characteristics of a "good" aptamer.…”

Section: Discussionmentioning

confidence: 99%

“…Despite the promise of these elegant studies, in general, these approaches require technology and expertise that is largely unavailable to most researchers. Thus, generic DNA SELEX remains by far the most popular aptamer isolation technique 29,30 . Efforts to identify which steps may contribute to the failure of SELEX are hampered by the complex dynamics of aptamer enrichment and the dearth of deep sequencing information from published studies to inform selections 31,32 .…”

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confidence: 99%

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High-efficiency enrichment enables identification of aptamers to circulating Plasmodium falciparum-infected erythrocytes

Oteng

McKeague

2020

Sci Rep

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Plasmodium falciparum is the causative agent of the deadliest human malaria. New molecules are needed that can specifically bind to erythrocytes that are infected with P. falciparum for diagnostic purposes, to disrupt host-parasite interactions, or to deliver chemotherapeutics. Aptamer technology has the potential to revolutionize biological diagnostics and therapeutics; however, broad adoption is hindered by the high failure rate of the systematic evolution of ligands by exponential enrichment (SELEX). Here we performed parallel SELEX experiments to compare the impact of two different methods for single-strand recovery on the efficiency of aptamer enrichment. Our experimental results and analysis of SELEX publications spanning 13 years implicate the alkaline denaturation step as a significant cause for inefficient aptamer selection. Thus, we applied an exonuclease single-strand recovery step in our SELEX to direct aptamers to the surface of erythrocytes infected with P. falciparum. The selected aptamers bind with high affinity (low nanomolar K d values) and selectivity to exposed surface proteins of both laboratory parasite strains as well isolates from patients in Asia and Africa with clinical malaria. The results obtained in this study potentially open new approaches to malaria diagnosis and surveillance. Malaria is a global health problem, with 228 million clinical episodes and an estimated 405,000 deaths each year 1. Plasmodium falciparum is the causative agent of the deadliest human malaria. A constellation of parasite-encoded proteins is displayed on the surface of infected erythrocytes (IEs) that are important in the life cycle of the parasite and mediate critical host-parasite interactions. One important host-parasite interaction is IE sequestration in the microvasculature which is thought to lead to the most severe manifestations of the disease. Therefore, ligand-binding molecules that can specifically identify and bind to IEs may disrupt important host-parasite interactions, deliver chemotherapeutics to parasitized cells, and have potential to further our understanding of the diversity of proteins expressed on the IE surface 2-5. Indeed, antibodies targeting IEs disrupt pathogenesis and contribute to naturally-acquired immunity to malaria 6-9. Aptamers are an alternative to antibodies that may sufficiently serve as molecular recognition agents of IEs. Several aptamers have been selected that bind to targets and biomarkers of malaria infection (see review 10 for examples). Aptamers are single-stranded nucleic acids that bind target molecules with antibody-like affinity and specificity through shape complementarity 11,12. However, the nucleic acid composition of aptamers presents several advantages over antibodies including increased stability, cell-free synthesis (either by PCR or chemical synthesis), and simple modification for detection and immobilization. Aptamers have been developed against small molecules, peptides, proteins, and whole cells including bacteria and parasites 3,13-16. As such, aptame...

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Section: Discussionmentioning

confidence: 99%

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confidence: 99%

High-efficiency enrichment enables identification of aptamers to circulating Plasmodium falciparum-infected erythrocytes

Oteng

McKeague

2020

Sci Rep

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“…Aptamers have excellent molecular recognition abilities that make them uniquely suited for drug delivery, diagnostic, and therapeutic applications [9,10]. Using systematic evolution of ligands by exponential enrichment (SELEX), DeRosa et al developed and studied different aptamers for a variety of molecular targets, including neurotransmitters, cancer biomarkers, and viruses [11,12], and further developed aptamer-based MRI and computed tomography (CT) contrast agents by conjugating the aptamers and the contrast agents (as seen in Figure 1) for detection of intercranial thrombus in neurotraumatic patients [13,14]. Current research efforts are focused on developing new aptamer-based contrast agents to image fibrin in attempts to localize intercranial blood clots for early detection, as clinical treatment options for intercranial blood clots and prevention of their complications are highly dependent on early detections [15].…”

Section: Contrast Agentsmentioning

confidence: 99%

Advancements in Canadian Biomaterials Research in Neurotraumatic Diagnosis and Therapies

Chen

Auriat

et al. 2019

Processes

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Development of biomaterials for the diagnosis and treatment of neurotraumatic ailments has been significantly advanced with our deepened knowledge of the pathophysiology of neurotrauma. Canadian research in the fields of biomaterial-based contrast agents, non-invasive axonal tracing, non-invasive scaffold imaging, scaffold patterning, 3D printed scaffolds, and drug delivery are conquering barriers to patient diagnosis and treatment for traumatic injuries to the nervous system. This review highlights some of the highly interdisciplinary Canadian research in biomaterials with a focus on neurotrauma applications.

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“…The advantages of using biosensors in metabolic engineering have been extensively reviewed before [1][2][3] and will not be detailed further. Moreover, a wide array of techniques now exists to develop biosensors, from FRET [4] to riboswitches [5,6]: the interested reader is referred to those two excellent reviews that cover the strengths and limitations of the above-mentioned technologies [7•,8••]. In this review, we will focus on transcriptional biosensors in three different aspects.…”

Section: Introductionmentioning

confidence: 99%

Custom-made transcriptional biosensors for metabolic engineering

Koch

Pandi

Borkowski

et al. 2019

Current Opinion in Biotechnology

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Transcriptional biosensors allow screening, selection or dynamic regulation of metabolic pathways, and are therefore an enabling technology for faster prototyping of metabolic engineering and sustainable chemistry. Recent advances have been made, allowing for routine use of heterologous transcription factors, and new strategies such as chimeric protein design allow engineers to tap into the reservoir of metabolite-binding proteins. However, extending the sensing scope of biosensors is only the first step, and computational models can help in fine-tuning properties of biosensors for custom-made behavior. Moreover, metabolic engineering is bound to benefit from advances in cell-free expression systems, either for faster prototyping of biosensors or for whole-pathway optimization, making it both a means and an end in biosensor design. Highlights • Successful examples of transcriptional biosensor implementations are presented • Various engineering strategies extend the space of detectable chemicals • Novel strategies exist to transform metabolite-responding proteins into biosensors • Mathematical models of varying complexity can help tune biosensor properties • Biosensors using or designed for cell-free systems are presented

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In Vitro Selection and Characterization of DNA Aptamers to a Small Molecule Target

Cited by 21 publications

References 87 publications

High-efficiency enrichment enables identification of aptamers to circulating Plasmodium falciparum-infected erythrocytes

High-efficiency enrichment enables identification of aptamers to circulating Plasmodium falciparum-infected erythrocytes

Advancements in Canadian Biomaterials Research in Neurotraumatic Diagnosis and Therapies

Custom-made transcriptional biosensors for metabolic engineering

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