Exploration of New Nucleic Acid Factors to Improve the Development of Synthetic Biosensing Switches
Bibliographic record
Abstract
In biosensor development, synthetic nucleic acid or protein responsive switches are critical for improving disease diagnostic performance. My thesis investigates several topics related to the exploration of new nucleic acid factors to improve the performance of current strand displacement switches and to discover RNA/ssDNA-responsive prokaryotic promoters. Incorporating a secondary looped structural element (flex-loop) into a switching domain allows the acceleration of toehold-mediated strand displacement in various constructs, including toehold-exchange, associative toehold, remote toehold, and allosteric toehold systems. The flex-loop switch has the ability to boost the i) efficiency of DNA-based computing, ii) analytical performance of an allosteric toehold technique for DNA sensing, and iii) performance of artificial gene circuits that regulate gene expression. Additionally, the flex-loop regulatory element enabled protein responsive proximity-strand displacement switch was investigated. The proximity flex-loop switch-functionalized antibodies allowed on-cell multi-receptor detection. This strategy, combined with the droplet microfluidic technique, enables a fluorescent-droplet cytometry assay that allows the characterization and profiling of live cancer cells with the single-cell resolution based on surface expression phenotype. Using this approach, we achieve live-cell phenotype profiling of multi-surface markers acquired with small (< 40 cells) collections of cells. We also develop an in vitro, non-traditional promoter structure selection strategy, in which a cell-free lacZ gene expression screening method carries on a new type of promoter structure acting as transcription factors. These T7 promoter structures performed as cis-acting transcription factors under the control of prokaryotic RNA polymerase. A rationally designed sequence responsive T7 promoter, termed Omega promoter, was developed to control gene expression via a toehold-mediated promoter rearrangement principal. Transcription-only Omega promoter-based sensors demonstrate engineered reporting activity through the use of RNA aptamers or control of Cas12a cleavage. We also discover that Omega promoter sensors could be operated from a circular DNA template for sequence-specific detection of ssDNA and RNA. The Omega promoter design can be directly adapted to the control of SP6 RNAP using prescribed sequences of ssDNA/RNA complexes. A single-stranded Omega promoter enables DNA-dependent RNA polymerase to accomplish the simplest in vitro RNA replication. Together, these studies explore nucleic acid factors to improve synthetic biosensing switches development in disease diagnostic applications.
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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) | 0.001 | 0.000 |
| Bibliometrics | 0.000 | 0.001 |
| 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".