Article ; Online: Protocol to prepare doubly labeled fluorescent nucleosomes for single-molecule fluorescence microscopy
STAR Protocols, Vol 4, Iss 2, Pp 102229- (2023)
2023
Abstract: Summary: Single-molecule fluorescence microscopy (SMFM) has been shown to be informative in understanding the interaction of chromatin-associated factors with nucleosomes, the basic building unit of chromatin. Here, we present a protocol for preparing ... ...
Abstract | Summary: Single-molecule fluorescence microscopy (SMFM) has been shown to be informative in understanding the interaction of chromatin-associated factors with nucleosomes, the basic building unit of chromatin. Here, we present a protocol for preparing doubly labeled fluorescent nucleosomes for SMFM. We describe steps for over-expression in E. coli and purification of recombinant human core histones. We then detail fluorescent labeling of histones and nucleosomal double-stranded DNA followed by octamer refolding and nucleosome reconstitution. : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. |
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Keywords | Biophysics ; Single-molecule Assays ; Microscopy ; Molecular Biology ; Molecular/Chemical Probes ; Protein Biochemistry ; Science (General) ; Q1-390 |
Language | English |
Publishing date | 2023-06-01T00:00:00Z |
Publisher | Elsevier |
Document type | Article ; Online |
Database | BASE - Bielefeld Academic Search Engine (life sciences selection) |
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