title: Nucleotide bias observed with a short SELEX RNA aptamer library.
abstract: Systematic evolution of ligands by exponential enrichment (SELEX) is a powerful in vitro selection process used for over 2 decades to identify oligonucleotide sequences (aptamers) with desired properties (usually high affinity for a protein target) from randomized nucleic acid libraries. In the case of RNA aptamers, several highly complex RNA libraries have been described with RNA sequences ranging from 71 to 81 nucleotides (nt) in length. In this study, we used high-throughput sequencing combined with bioinformatics analysis to thoroughly examine the nucleotide composition of the sequence pools derived from several selections that employed an RNA library (Sel2N20) with an abbreviated variable region. The Sel2N20 yields RNAs 51 nt in length, which unlike longer RNAs, are more amenable to large-scale chemical synthesis for therapeutic development. Our analysis revealed a consistent and early bias against inclusion of adenine, resulting in aptamers with lower predicted minimum free energies (ΔG) (higher structural stability). This bias was also observed in control, "nontargeted" selections in which the partition step (against the target) was omitted, suggesting that the bias occurred in 1 or more of the amplification and propagation steps of the SELEX process.
authors.affiliation: Department of Internal Medicine, University of Iowa, Iowa City, IA 52242, USA.
authors.forename: William H
authors.initials: WH
authors.lastname: Thiel
authors.forename: Thomas
authors.initials: T
authors.lastname: Bair
authors.forename: Kristina
authors.initials: K
authors.lastname: Wyatt Thiel
authors.forename: Justin P
authors.initials: JP
authors.lastname: Dassie
authors.forename: William M
authors.initials: WM
authors.lastname: Rockey
authors.forename: Craig A
authors.initials: CA
authors.lastname: Howell
authors.forename: Xiuying Y
authors.initials: XY
authors.lastname: Liu
authors.forename: Adam J
authors.initials: AJ
authors.lastname: Dupuy
authors.forename: Lingyan
authors.initials: L
authors.lastname: Huang
authors.forename: Richard
authors.initials: R
authors.lastname: Owczarzy
authors.forename: Mark A
authors.initials: MA
authors.lastname: Behlke
authors.forename: James O
authors.initials: JO
authors.lastname: McNamara
authors.forename: Paloma H
authors.initials: PH
authors.lastname: Giangrande
mesh_terms: D000818:Animals; D052157:Aptamers, Nucleotide / Q000737:chemistry*; D001482:Base Composition; D001483:Base Sequence; D002460:Cell Line; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D051379:Mice; D011743:Pyrimidines / Q000737:chemistry*; D036104:Receptor, EphA2 / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry; D052156:SELEX Aptamer Technique; D017423:Sequence Analysis, RNA; D013816:Thermodynamics; D014158:Transcription, Genetic
chemical_list: D052157:Aptamers, Nucleotide; D011743:Pyrimidines; D011994:Recombinant Proteins; D036104:Receptor, EphA2