In vitro selection of DNA aptamers binding ethanolamine

Mann, D E; Reinemann, Christine; Stoltenburg, Regina; Strehlitz, Beate

doi:10.1016/j.bbrc.2005.10.172

Cited by 125 publications

(96 citation statements)

References 14 publications

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“…Aptamers have been generated against many classes of molecular targets, including small molecules (5,6), proteins (7,8), and cell-surface markers (9,10), for a wide range of applications including diagnostics (11,12), molecular imaging (13,14), targeted therapeutics (15), gene delivery (16), and drug delivery (17). Systematic evolution of ligands by exponential enrichment (SELEX) offers an effective in vitro selection method for the isolation of aptamers from random libraries of nucleic acids.…”

mentioning

confidence: 99%

Quantitative selection of DNA aptamers through microfluidic selection and high-throughput sequencing

Cho

Xiao

Nie

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

239

244

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We describe the integration of microfluidic selection with high-throughput DNA sequencing technology for rapid and efficient discovery of nucleic acid aptamers. The Quantitative Selection of Aptamers through Sequencing method tracks the copy number and enrichment-fold of more than 10 million individual sequences through multiple selection rounds, enabling the identification of high-affinity aptamers without the need for the pool to fully converge to a small number of sequences. Importantly, this method allows the discrimination of sequences that arise from experimental biases rather than true high-affinity target binding. As a demonstration, we have identified aptamers that specifically bind to PDGF-BB protein with K d < 3 nM within 3 rounds. Furthermore, we show that the aptamers identified by Quantitative Selection of Aptamers through Sequencing have ∼3–8-fold higher affinity and ∼2–4-fold higher specificity relative to those discovered through conventional cloning methods. Given that many biocombinatorial libraries are encoded with nucleic acids, we extrapolate that our method may be extended to other types of libraries for a range of molecular functions.

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mentioning

confidence: 99%

Quantitative selection of DNA aptamers through microfluidic selection and high-throughput sequencing

Cho

Xiao

Nie

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

239

244

View full text Add to dashboard Cite

show abstract

“…Multiple rounds of selection (typically [8][9][10][11][12][13][14][15] are generally necessary to isolate aptamers with sufficient specificity and binding affinity (e.g., nanomolar dissociation constant (K d ) for proteins and micromolar for small molecules), which therefore requires significant resources and time (17). To accelerate this lengthy discovery process, a wide variety of molecular separation techniques-beyond traditional filter-binding assays (18) and affinity chromatography (19)-have been explored as means to enhance the efficiency of aptamer selection, including surface plasmon resonance (20), flow cytometry (21), and capillary electrophoresis (CE) (22)(23)(24)(25). In particular, CE-based separation methods have shown remarkable selection efficiencies for protein targets that exhibit significant shifts in the electrophoretic mobility ( e ) upon binding to aptamers; proteinbinding aptamers with K d in the low-nanomolar range have been isolated after a small number (Ϸ1-4) of selection rounds (22)(23)(24)(25).…”

mentioning

confidence: 99%

“…To accelerate this lengthy discovery process, a wide variety of molecular separation techniques-beyond traditional filter-binding assays (18) and affinity chromatography (19)-have been explored as means to enhance the efficiency of aptamer selection, including surface plasmon resonance (20), flow cytometry (21), and capillary electrophoresis (CE) (22)(23)(24)(25). In particular, CE-based separation methods have shown remarkable selection efficiencies for protein targets that exhibit significant shifts in the electrophoretic mobility ( e ) upon binding to aptamers; proteinbinding aptamers with K d in the low-nanomolar range have been isolated after a small number (Ϸ1-4) of selection rounds (22)(23)(24)(25). However, the method is less effective for other classes of targets that do not induce sufficient e shift of aptamers (e.g., small molecules and cell surfaces), or those that cannot be accommodated in the separation apparatus (e.g., whole organisms).…”

mentioning

confidence: 99%

Micromagnetic selection of aptamers in microfluidic channels

Lou

Qian

Xiao

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

320

284

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Aptamers are nucleic acid molecules that have been selected in vitro to bind to their molecular targets with high affinity and specificity. Typically, the systematic evolution of ligands by exponential enrichment (SELEX) process is used for the isolation of specific, high-affinity aptamers. SELEX, however, is an iterative process requiring multiple rounds of selection and amplification that demand significant time and labor. Here, we describe an aptamer discovery system that is rapid, highly efficient, automatable, and applicable to a wide range of targets, based on the integration of magnetic bead-based SELEX process with microfluidics technology. Our microfluidic SELEX (M-SELEX) method exploits a number of unique phenomena that occur at the microscale and implements a design that enables it to manipulate small numbers of beads precisely and isolate high-affinity aptamers rapidly. As a model to demonstrate the efficiency of the M-SELEX process, we describe here the isolation of DNA aptamers that tightly bind to the light chain of recombinant Botulinum neurotoxin type A (with low-nanomolar dissociation constant) after a single round of selection.microchannel ͉ recombinant Botulinum neurotoxin type A ͉ systematic evolution of ligands by exponential enrichment

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“…33 Most of them have a common conserved G-rich section of 16 nucleotides at the 5 0 -end of the core region. It may imply that the conserved 16-mer G-rich section might be the real binding motif to ethanolamine.…”

Section: Specific G-quadruplex Sequences Binding To Diethanolamine Anmentioning

confidence: 99%

Specific DNA G‐quadruplexes bind to ethanolamines

et al. 2009

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A significant number of G-quadruplex-forming sequences have been revealed in human genome by bioinformatic searches, implying that G-quadruplexes may be involved in important biological processes and may be new chemotherapeutic targets. Therefore, it is important to discover the potential interactions of G-quadruplexes with other molecules or groups. Here we describe a class of G-quadruplexes, which can bind to ethanolamine groups that widely exist in biomolecules and drug molecules. The specific interaction of these G-quadruplexes with ethanolamine groups was identified by high performance affinity chromatography (HPAC) using immobilized ethanolamine and diethanolamine as stationary phase reagents. The circular dichroism (CD) spectra and native polyacrylamide gel electrophoresis (PAGE) show that these ethanolamine binding quadruplexes adopt an intramolecularly parallel structure. The relationship of ethanolamine binding and G-quadruplexe structure provides new clues for the G-quadruplex-related studies as well as for the molecular designs of therapeutic reagents.

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In vitro selection of DNA aptamers binding ethanolamine

Cited by 125 publications

References 14 publications

Quantitative selection of DNA aptamers through microfluidic selection and high-throughput sequencing

Quantitative selection of DNA aptamers through microfluidic selection and high-throughput sequencing

Micromagnetic selection of aptamers in microfluidic channels

Specific DNA G‐quadruplexes bind to ethanolamines

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