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Integrating quantitative proteomics with accurate genome profiling of transcription factors by greenCUT&RUN

Nucleic Acids Res. 2021-05; 
Sheikh Nizamuddin, Stefanie Koidl, Tanja Bhuiyan, Tamara V Werner, Martin L Biniossek, Alexandre M J J Bonvin, Silke Lassmann, HThMarc Timmers
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Catalog Antibody The 40 μg of chromatin were incubated with either 2 μg of NFYA antibody (Santa Cruz, sc11753) or 5 μg of GFP antibody (GenScript, A01704) in IP buffer Get A Quote

Abstract

Genome-wide localization of chromatin and transcription regulators can be detected by a variety of techniques. Here, we describe a novel method 'greenCUT&RUN' for genome-wide profiling of transcription regulators, which has a very high sensitivity, resolution, accuracy and reproducibility, whilst assuring specificity. Our strategy begins with tagging of the protein of interest with GFP and utilizes a GFP-specific nanobody fused to MNase to profile genome-wide binding events. By using a GFP-nanobody the greenCUT&RUN approach eliminates antibody dependency and variability. Robust genomic profiles were obtained with greenCUT&RUN, which are accurate and unbiased towards open chromatin. By integrating greenCUT&RUN w... More

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