Customised nucleic acid libraries for enhanced aptamer selection and performance

Pfeiffer, Franziska; Rosenthal, Malte; Siegl, Julia; Ewers, Jörg; Mayer, Günter

doi:10.1016/j.copbio.2017.03.026

Cited by 53 publications

(44 citation statements)

References 48 publications

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“…Base-modified aptamers that incorporate non-natural chemical functional groups provide many advantages as affinity reagents because, like conventional aptamers, they are chemically synthesized and sequence-defined, while also offering a much broader chemical repertoire than their natural DNA and RNA counterparts 1 . This can result in a commensurate expansion in the spectrum of targets that can be recognized with high affinity and specificity, and a number of groups have developed effective strategies for introducing such modifications into nucleic acids over the past two decades.…”

Section: Introductionmentioning

confidence: 99%

Click-PD: A Quantitative Method for Base-Modified Aptamer Discovery

Gordon

Wu²,

Feagin

et al. 2019

Preprint

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Base-modified aptamers that incorporate non-natural chemical moieties can achieve greatly improved affinity and specificity relative to natural DNA or RNA aptamers. However, conventional methods for generating base-modified aptamers require considerable expertise and resources. In this work, we have accelerated and generalized the process of generating basemodified aptamers by combining a click-chemistry strategy with a fluorescence-activated cell sorting (FACS)-based screening methodology that measures the affinity and specificity of individual aptamers at a throughput of ~10^7 per hour. Our "click-PD" strategy offers many advantages. First, almost any chemical modification can be introduced with a commerciallyavailable polymerase. Second, click-PD can screen vast numbers of individual aptamers based on quantitative on-and off-target binding measurements to simultaneously achieve high affinity and specificity. Finally, it requires minimal specialized equipment or reagents besides a FACS instrument which is now widely-available. Using click-PD, we generated a boronic acid-modified aptamer with ~1 μM affinity for epinephrine, a target for which no aptamer has been reported to date. We subsequently generated a mannose-modified aptamer with nanomolar affinity for the lectin concanavalin A (ConA). The strong affinity of both aptamers is fundamentally dependent upon the presence of chemical modifications, and we show that their removal essentially eliminates aptamer binding. Importantly, our ConA aptamer exhibited exceptional specificity, with minimal binding to other structurally-similar lectins. Finally, we show that our aptamer remarkable biological activity. Indeed, to the best of our knowledge, this aptamer is the most potent inhibitor of Con A-mediated hemagglutination reported to date.

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Section: Introductionmentioning

confidence: 99%

Click-PD: A Quantitative Method for Base-Modified Aptamer Discovery

Gordon

Wu²,

Feagin

et al. 2019

Preprint

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“…This is mainly linked to the limited chemical space covered by these libraries given by the canonical set of nucleotides, which lacks the versatility of functional groups of amino acids residues in antibodies. Thus, approaches that increase the chemical versatility of nucleic acid libraries are extensively pursued . (−)‐Δ 9 ‐Tetrahydrocannabinol (THC) represents a ligand that effectively escapes conventional selection approaches (Table S1, Figure S1 in the Supporting Information).…”

Section: Figurementioning

confidence: 99%

A Receptor‐Guided Design Strategy for Ligand Identification

2019

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Biomedical sciences require effective tools to manipulate,d etect, and study biological phenomena. Oligo-(deoxy)nucleotide ligands represent such tools,but the current strategies to generate them are restricted. Their limited availability is insufficient to address the broad range of targets related to biomedical research. Exemplified by targeting the hydrophobic molecule (À)-D 9 -tetrahydrocannabinol (THC), we report areceptor-guided design (RGD) strategy to generate chemically modified oligodeoxynucleotide libraries for the tailored selection of clickmers. Angewandte ChemieCommunications

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“…Short randomized regions ensure representation of each sequence in the pool, albeit with less structural complexity. Although longer libraries do not cover the full sequence space, the possibility to form more complex structural motifs increases [52]. To impart additional physicochemical diversity to aptamers, nucleobase modifications can be introduced to the library [53].…”

Section: Experimental Design Of Selexmentioning

confidence: 99%

A Bottom-Up Approach for Developing Aptasensors for Abused Drugs: Biosensors in Forensics

et al. 2019

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Aptamer-based point-of-care (POC) diagnostics platforms may be of substantial benefit in forensic analysis as they provide rapid, sensitive, user-friendly, and selective analysis tools for detection. Aptasensors have not yet been adapted commercially. However, the significance of the applications of aptasensors in the literature exceeded their potential. Herein, in this review, a bottom-up approach is followed to describe the aptasensor development and application procedure, starting from the synthesis of the corresponding aptamer sequence for the selected analyte to creating a smart surface for the sensitive detection of the molecule of interest. Optical and electrochemical biosensing platforms, which are designed with aptamers as recognition molecules, detecting abused drugs are critically reviewed, and existing and possible applications of different designs are discussed. Several potential disciplines in which aptamer-based biosensing technology can be of greatest value, including forensic drug analysis and biological evidence, are then highlighted to encourage researchers to focus on developing aptasensors in these specific areas.

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Customised nucleic acid libraries for enhanced aptamer selection and performance

Cited by 53 publications

References 48 publications

Click-PD: A Quantitative Method for Base-Modified Aptamer Discovery

Click-PD: A Quantitative Method for Base-Modified Aptamer Discovery

A Receptor‐Guided Design Strategy for Ligand Identification

A Bottom-Up Approach for Developing Aptasensors for Abused Drugs: Biosensors in Forensics

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