title: An In Vitro Selection Protocol for Threose Nucleic Acid (TNA) Using DNA Display.
abstract: Threose nucleic acid (TNA) is an unnatural genetic polymer composed of repeating threofuranosyl sugars linked by 2' and 3' phosphodiester bonds. TNA is capable of forming antiparallel Watson-Crick duplex structures in a self-pairing mode, and can also cross-pair opposite complementary strands of DNA and RNA. The solution NMR structure of a self-complementary TNA duplex reveals that TNA adopts an A-form helical structure, which explains its ability to exchange genetic information with natural genetic polymers. In a recent advance, a TNA aptamer was evolved from a pool of random sequences using an engineered polymerase that can copy DNA sequences into TNA. This unit details the steps required to evolve functional TNA molecules in the laboratory using a method called DNA display. Using this approach, TNA molecules are physically linked to their encoding double-stranded DNA template. By linking TNA phenotype with DNA genotype, one can select for TNA molecules with a desired function and recover their encoding genetic information by PCR amplification. Each round of selection requires ∼3 days to complete and multiple rounds of selection and amplification are required to generate functional TNA molecules.
authors: School of Life Sciences, Arizona State University, Tempe, Arizona; The Biodesign Institute at Arizona State University, Tempe, Arizona.
authors: Matthew R
authors: MR
authors: Dunn
authors: John C
authors: JC
authors: Chaput
mesh_terms: D004247:DNA / Q000737:chemistry*; D019020:Directed Molecular Evolution / Q000379:methods*; D009690:Nucleic Acid Conformation; D016133:Polymerase Chain Reaction / Q000379:methods*; D013780:Tetroses / Q000737:chemistry*
chemical_list: D013780:Tetroses; D004247:DNA; C073321:erythrose
keywords: DNA display; aptamer; in vitro selection; oligonucleotide; threose nucleic acid (TNA)