Specific interaction between transcription factor (TF) and cis-regulatory elements in the context of chromatin is the key to determining gene expression. Thus, it is important to measure DNA-binding specificity and identify cis-elements of TFs of your interest. In this chapter, we described a microwell-based assay to determine DNA-binding specificity by using translational fusion of a TF with a highly active cellulase (CELD), which hydrolyzes 4-methylumbelliferyl β-d-cellobioside to a fluorescent 4-methylumbelliferone product. The hydrolysis activity arrows us to quantify the binding strength between TF-CELD fusion protein and biotinylated DNA sequences in a 96-well microplate. The high-throughput and quantitative nature of CELD assay enable researchers to test a large number of putative DNA-binding sites of TFs, which subsequently leads to the identification of its direct target genes.
CITATION STYLE
Kalaipandian, S., & Xue, G. P. (2018). Rapid and quantitative CELD assay to measure the specificity of transcription factor-DNA-binding interactions and identify cis-elements. In Methods in Molecular Biology (Vol. 1830, pp. 307–324). Humana Press Inc. https://doi.org/10.1007/978-1-4939-8657-6_18
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