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Fig. 2 | Genome Biology

Fig. 2

From: Mechanisms of enhancer action: the known and the unknown

Fig. 2

Cell-free assays to interrogate EPC and transcription. Four cell-free assays are envisaged where a construct with an enhancer and a promoter, or isolated enhancer and promoter fragments, or a BAC clone can be used as templates to interrogate EPC and transcription activation. Streptavidin-coated M280 magnetic beads are employed to capture EPCs on biotinylated fragments. These assays offer unique advantages not achievable through existing cell-based approaches, such as (1) same-source verification of EPC and transcription, (2) exploring mechanistic and/or causal link between EPC and transcription, (3) proteomics of EPC to identify protein complexes that (i) mediate looping and (ii) ensure transcription, and (4) biochemistry of EPC to elucidate requirements of non-proteinaceous ingredients (e.g., ATP, NTPs, coenzymes, and cofactors) in looping-coupled transcription activation. Arguably, any putative enhancer-promoter pair can be studied using the nuclear extract (NE) prepared from a relevant cell line. See (Panigrahi et al., 2018) for details

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