FRep: A Fluorescent Protein-Based Bioprobe for <i>in Vivo</i> Detection of Protein–DNA Interactions
Bibliographic record
Abstract
We describe a bacterial reporter system, FRep, for rapid and facile detection of protein-DNA recognition. The bioprobe reporter comprises genes of two fluorescent proteins (FPs) separated by a potential DNA target. If a coexpressed transcription factor binds the DNA target, transcription of the second FP is impeded, resulting in loss of FRET partner. Using ratiometric FRET, we show that evaluation of protein-DNA recognition can be reliably made on bZIP and bHLHZ transcription factors and their DNA targets. FRep displays similar thresholds of detection regarding protein-DNA binding affinities, as compared to well-established electrophoretic and yeast assays, although we observed variations in the intensity of fluorescence signals and detection thresholds that may depend on differences between DNA-binding protein production levels and/or stability in the cell, or the expressed bioprobe linker between the two FPs. FRep can potentially be applied to high-throughput searches of both protein and DNA libraries; in a mock library screen, binding and nonbinding complexes can even be distinguished by visual inspection of colonies on plates. FRep presents notable advantages over existing technologies when applied to assessing protein-DNA interactions in vivo, and this approach has the potential for applications in assaying protein-protein interactions and screening molecules that influence specific macromolecular interactions.
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How this classification was reachedexpand
Full frame distilled prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. Learned from the 10,348 direct Codex labels and 10,348 direct Gemma labels. Candidate is the union of thresholded teacher heads; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels or direct frontier model labels.
Codex and Gemma teacher scores by category
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.000 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one teacher head, not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".