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RNA Ligation for Mono and Dually Labeled RNAs.
Depaix, Anaïs; Mlynarska-Cieslak, Agnieszka; Warminski, Marcin; Sikorski, Pawel J; Jemielity, Jacek; Kowalska, Joanna.
Afiliación
  • Depaix A; Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Pasteura 5, 02-093, Warsaw, Poland.
  • Mlynarska-Cieslak A; Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Pasteura 5, 02-093, Warsaw, Poland.
  • Warminski M; Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Pasteura 5, 02-093, Warsaw, Poland.
  • Sikorski PJ; Centre of New Technologies, University of Warsaw, Banacha 2c, 02-097, Warsaw, Poland.
  • Jemielity J; Centre of New Technologies, University of Warsaw, Banacha 2c, 02-097, Warsaw, Poland.
  • Kowalska J; Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Pasteura 5, 02-093, Warsaw, Poland.
Chemistry ; 27(47): 12190-12197, 2021 Aug 19.
Article en En | MEDLINE | ID: mdl-34114681
Labeled RNAs are invaluable probes for investigation of RNA function and localization. However, mRNA labeling remains challenging. Here, we developed an improved method for 3'-end labeling of in vitro transcribed RNAs. We synthesized novel adenosine 3',5'-bisphosphate analogues modified at the N6 or C2 position of adenosine with an azide-containing linker, fluorescent label, or biotin and assessed these constructs as substrates for RNA labeling directly by T4 ligase or via postenzymatic strain-promoted alkyne-azide cycloaddition (SPAAC). All analogues were substrates for T4 RNA ligase. Analogues containing bulky fluorescent labels or biotin showed better overall labeling yields than postenzymatic SPAAC. We successfully labeled uncapped RNAs, NAD-capped RNAs, and 5'-fluorescently labeled m7 Gp3 Am -capped mRNAs. The obtained highly homogenous dually labeled mRNA was translationally active and enabled fluorescence-based monitoring of decapping. This method will facilitate the use of various functionalized mRNA-based probes.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Azidas / ARN Idioma: En Revista: Chemistry Asunto de la revista: QUIMICA Año: 2021 Tipo del documento: Article País de afiliación: Polonia Pais de publicación: Alemania
Texto completo
Añadir a Mi BVS
Imprimir
XML
PubMed Links
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Azidas / ARN Idioma: En Revista: Chemistry Asunto de la revista: QUIMICA Año: 2021 Tipo del documento: Article País de afiliación: Polonia Pais de publicación: Alemania