Unknown

Dataset Information

0

BET-seq: Binding energy topographies revealed by microfluidics and high-throughput sequencing.


ABSTRACT: Biophysical models of transcriptional regulation rely on energetic measurements of the binding affinities between transcription factors (TFs) and target DNA binding sites. Historically, assays capable of measuring TF-DNA binding affinities have been relatively low-throughput (measuring ~103 sequences in parallel) and have required significant specialized equipment, limiting their use to a handful of laboratories. Recently, we developed an experimental assay and analysis pipeline that allows measurement of binding energies between a single TF and up to 106 DNA species in a single experiment (Binding Energy Topography by sequencing, or BET-seq) (Le et al., 2018). BET-seq employs the Mechanically Induced Trapping of Molecular Interactions (MITOMI) platform to purify DNA bound to a TF at equilibrium followed by high coverage sequencing to reveal relative differences in binding energy for each sequence. While we have previously used BET-seq to refine the binding affinity landscapes surrounding high-affinity DNA consensus target sites, we anticipate this technique will be applied in future work toward measuring a wide variety of TF-DNA landscapes. Here, we provide detailed instructions and general considerations for DNA library design, performing BET-seq assays, and analyzing the resulting data.

SUBMITTER: Aditham AK 

PROVIDER: S-EPMC7531582 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

altmetric image

Publications

BET-seq: Binding energy topographies revealed by microfluidics and high-throughput sequencing.

Aditham Arjun K AK   Shimko Tyler C TC   Fordyce Polly M PM  

Methods in cell biology 20181023


Biophysical models of transcriptional regulation rely on energetic measurements of the binding affinities between transcription factors (TFs) and target DNA binding sites. Historically, assays capable of measuring TF-DNA binding affinities have been relatively low-throughput (measuring ~10<sup>3</sup> sequences in parallel) and have required significant specialized equipment, limiting their use to a handful of laboratories. Recently, we developed an experimental assay and analysis pipeline that  ...[more]

Similar Datasets

| S-EPMC4441486 | biostudies-other
| S-EPMC5561429 | biostudies-literature
| S-EPMC3665311 | biostudies-other
| S-EPMC6120635 | biostudies-literature
| S-EPMC10263358 | biostudies-literature
| S-EPMC2984161 | biostudies-other
| S-EPMC3607113 | biostudies-literature
| S-EPMC9382637 | biostudies-literature
| S-EPMC3988572 | biostudies-literature
| S-EPMC9750877 | biostudies-literature