Electrophoretic Mobility Shift Assay of DNA and CRISPR-Cas Ribonucleoprotein Complexes.

Autor: Künne T; Laboratory of Microbiology, Department of Agrotechnology and Food Sciences, Wageningen University, Dreijenplein 10, 6703 HB, Wageningen, The Netherlands., Westra ER, Brouns SJ
Jazyk: angličtina
Zdroj: Methods in molecular biology (Clifton, N.J.) [Methods Mol Biol] 2015; Vol. 1311, pp. 171-84.
DOI: 10.1007/978-1-4939-2687-9_11
Abstrakt: The Electrophoretic Mobility Shift Assay is a straightforward and inexpensive method for the determination and quantification of protein-nucleic acid interactions. It relies on the different mobility of free and protein-bound nucleic acid in a gel matrix during electrophoresis. Nucleic acid affinities of crRNA-Cas complexes can be quantified by calculating the dissociation constant (Kd). Here, we describe how two types of EMSA assays are performed using the Cascade ribonucleoprotein complex from Escherichia coli as an example.
Databáze: MEDLINE