title: A multiplexed transcription activator-like effector system for detecting specific DNA sequences.
abstract: Transcription activator-like effectors (TALEs), originating from the Xanthomonas genus of bacteria, bind to specific DNA sequences based on amino acid sequence in the repeat-variable diresidue (RVD) positions of the protein. By altering these RVDs, it has been shown that a TALE protein can be engineered to bind virtually any DNA sequence of interest. The possibility of multiplexing TALEs for the purposes of identifying specific DNA sequences has yet to be explored. Here, we demonstrate a system in which a TALE protein bound to a nitrocellulose strip has been utilized to capture purified DNA, which is then detected using the binding of a second distinct TALE protein conjugated to a protein tag that is then detected by a dot blot. This system provides a signal only when both TALEs bind to their respective sequences, further demonstrating the specificity of the TALE binding.
authors.affiliation: Department of Biological Sciences, University of Calgary , Calgary, Alberta T2N 1N4, Canada.
authors.forename: Ali
authors.initials: A
authors.lastname: Honarmand
authors.forename: Robert
authors.initials: R
authors.lastname: Mayall
authors.forename: Iain
authors.initials: I
authors.lastname: George
authors.forename: Lisa
authors.initials: L
authors.lastname: Oberding
authors.forename: Himika
authors.initials: H
authors.lastname: Dastidar
authors.forename: Jamie
authors.initials: J
authors.lastname: Fegan
authors.forename: Somshukla
authors.initials: S
authors.lastname: Chaudhuri
authors.forename: Justin
authors.initials: J
authors.lastname: Dole
authors.forename: Sharon
authors.initials: S
authors.lastname: Feng
authors.forename: Denny
authors.initials: D
authors.lastname: Hoang
authors.forename: Ruth
authors.initials: R
authors.lastname: Moges
authors.forename: Julie
authors.initials: J
authors.lastname: Osgood
authors.forename: Taylor
authors.initials: T
authors.lastname: Remondini
authors.forename: Wm Keith
authors.initials: WK
authors.lastname: van der Meulen
authors.forename: Su
authors.initials: S
authors.lastname: Wang
authors.forename: Chris
authors.initials: C
authors.lastname: Wintersinger
authors.forename: Amanda
authors.initials: A
authors.lastname: Zaparoli Zucoloto
authors.forename: Kate
authors.initials: K
authors.lastname: Chatfield-Reed
authors.forename: Mayi
authors.initials: M
authors.lastname: Arcellana-Panlilio
authors.forename: Anders
authors.initials: A
authors.lastname: Nygren
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004247:DNA / Q000032:analysis* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005818:Genetic Engineering; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014974:Xanthomonas / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D004247:DNA