Protocol for monitoring and analyzing single nucleotide incorporation by S. cerevisiae RNA polymerases

Autor: Ruth Q. Jacobs, Nathan F. Bellis, Aaron L. Lucius, David A. Schneider
Jazyk: angličtina
Rok vydání: 2023
Předmět:
Zdroj: STAR Protocols, Vol 4, Iss 2, Pp 102191- (2023)
Druh dokumentu: article
ISSN: 2666-1667
DOI: 10.1016/j.xpro.2023.102191
Popis: Summary: Here we present an optimized protocol for monitoring and analyzing single nucleotide incorporation by RNA polymerases. This protocol describes the assembly of Saccharomyces cerevisiae RNA polymerase I elongation complexes in a promoter-independent system in vitro. We describe how to collect a time course using a quench-flow, a rapid mixing instrument, and subsequently resolve reactions on a polyacrylamide gel. Finally, we detail how to quantify the gel images.For complete details on the use and execution of this protocol, please refer to Appling et al. (2015).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.
Databáze: Directory of Open Access Journals