title: In vitro selection with artificial expanded genetic information systems.
abstract: Artificially expanded genetic information systems (AEGISs) are unnatural forms of DNA that increase the number of independently replicating nucleotide building blocks. To do this, AEGIS pairs are joined by different arrangements of hydrogen bond donor and acceptor groups, all while retaining their Watson-Crick geometries. We report here a unique case where AEGIS DNA has been used to execute a systematic evolution of ligands by exponential enrichment (SELEX) experiment. This AEGIS-SELEX was designed to create AEGIS oligonucleotides that bind to a line of breast cancer cells. AEGIS-SELEX delivered an AEGIS aptamer (ZAP-2012) built from six different kinds of nucleotides (the standard G, A, C, and T, and the AEGIS nonstandard P and Z nucleotides, the last having a nitro functionality not found in standard DNA). ZAP-2012 has a dissociation constant of 30 nM against these cells. The affinity is diminished or lost when Z or P (or both) is replaced by standard nucleotides and compares well with affinities of standard GACT aptamers selected against cell lines using standard SELEX. The success of AEGIS-SELEX relies on various innovations, including (i) the ability to synthesize GACTZP libraries, (ii) polymerases that PCR amplify GACTZP DNA with little loss of the AEGIS nonstandard nucleotides, and (iii) technologies to deep sequence GACTZP DNA survivors. These results take the next step toward expanding the power and utility of SELEX and offer an AEGIS-SELEX that could possibly generate receptors, ligands, and catalysts having sequence diversities nearer to that displayed by proteins.
authors.affiliation: Department of Chemistry, University of Florida, Gainesville, FL 32611.
authors.forename: Kwame
authors.initials: K
authors.lastname: Sefah
authors.forename: Zunyi
authors.initials: Z
authors.lastname: Yang
authors.forename: Kevin M
authors.initials: KM
authors.lastname: Bradley
authors.forename: Shuichi
authors.initials: S
authors.lastname: Hoshika
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Jiménez
authors.forename: Liqin
authors.initials: L
authors.lastname: Zhang
authors.forename: Guizhi
authors.initials: G
authors.lastname: Zhu
authors.forename: Savita
authors.initials: S
authors.lastname: Shanker
authors.forename: Fahong
authors.initials: F
authors.lastname: Yu
authors.forename: Diane
authors.initials: D
authors.lastname: Turek
authors.forename: Weihong
authors.initials: W
authors.lastname: Tan
authors.forename: Steven A
authors.initials: SA
authors.lastname: Benner
mesh_terms: D001483:Base Sequence; D017931:DNA Primers; D007256:Information Systems; D016133:Polymerase Chain Reaction; D052156:SELEX Aptamer Technique; D012641:Selection, Genetic
chemical_list: D017931:DNA Primers
keywords: cancer cell SELEX; next generation sequencing; nucleic acids; synthetic biology