2D layout of bead based thioaptamer/aptamer selection platform for therapeutics and diagnostics

Simple item page
dc.contributor.advisorBruce A. Luxonen_US
dc.contributor.committeeMemberWilliam D. Willisen_US
dc.contributor.committeeMemberL.Lilian Chanen_US
dc.creatorXu Zhaoen_US
dc.date.accessioned2011-12-20T16:05:04Z
dc.date.accessioned2014-02-19T22:05:26Z
dc.date.available2008-06-17en_US
dc.date.available2011-12-20T16:05:04Z
dc.date.available2014-02-19T22:05:26Z
dc.date.created2007-07-25en_US
dc.date.issued2007-07-20en_US
dc.description.abstractNucleic acid research has expanded the way we can intervene with biological systems. Especially, oligonucleotide agents (ODN or aptamers) are believed to affect cell function via complementary recognition or binding to specific proteins by forming tertiary structure. This opens new ways in therapeutics and diagnostics. The phosphoro- mono-/di- thioate substitutions in the backbone (termed “thioaptamer”) grants ODN nuclease resistance and higher binding affinity. \r\nA bead-based combinatorial library, in which every bead contains a unique species of aptamers, provides a promising platform for selection of aptamers and thioaptamers. To successfully screen the bead-based library, 2D layout of beads in gel and on bead screening model is proposed. To develop the 2D layout of beads and its corresponding functional assays, a model system is first established: NF-kappa B proteins were expressed, purified and characterized. Thioaptamer XBY6, which specifically targets NF-kappa B protein, and its natural origin, I-kappa B were synthesized and verified. Thioaptamer purification using FPLC and HPLC was also investigated, and several 5’-funtionalized thioaptamers were successfully purified. Electrophoretic mobility shifting assay (EMSA) has been used to verify XBY6 binding, and ELISA assay has been used to verify I-kappa B binding towards human recombinant NF-kappa B protein. Preliminary study of bead in 2D gel showed applicability of bead-based selection and thus on bead functional assays were developed. Both double strand one species library with I-kappa B sequence and a 212=4096 different species beads library were constructed and verified. The library was then tested using on-bead EMSA like assay and ELISA assay. Both assays showed encouraging results for 2D layout selection and further enhancement of visualization (signal/noise improvement) is discussed. \r\nThe project suggested that 2D layout of beads in gel (PAGE) is well suited for parallel high-throughput selection of thioaptamers and aptamers, thus paving a new way for drug discovery and future therapeutics and diagnostics.\r\nen_US
dc.format.mediumelectronicen_US
dc.identifier.otheretd-07252007-082849en_US
dc.identifier.urihttp://hdl.handle.net/2152.3/187
dc.language.isoengen_US
dc.rightsCopyright © is held by the author. Presentation of this material on the TDL web site by The University of Texas Medical Branch at Galveston was made possible under a limited license grant from the author who has retained all copyrights in the works.en_US
dc.subjectThioaptamer XBY6en_US
dc.subjectnucleic aciden_US
dc.subjectNuclear Factor Kappa Ben_US
dc.subjectdrug developmenten_US
dc.subjectcombinatorial chemistryen_US
dc.subjectbead-based selectionen_US
dc.subjectAptameren_US
dc.title2D layout of bead based thioaptamer/aptamer selection platform for therapeutics and diagnosticsen_US
dc.type.genrethesisen_US
dc.type.materialtexten_US
thesis.degree.departmentHuman Biological Chemistry and Geneticsen_US
thesis.degree.grantorThe University of Texas Medical Branchen_US
thesis.degree.levelMasteren_US
thesis.degree.nameMaster of Scienceen_US

Files

Collections

University of Texas Medical Branch