An Integrated Approach to Functional Genomics: Construction of a Novel Reporter Gene Fusion Library for<i>Sinorhizobium meliloti</i>
Bibliographic record
Abstract
As a means of investigating gene function, we developed a robust transcription fusion reporter vector to measure gene expression in bacteria. The vector, pTH1522, was used to construct a random insert library for the Sinorhizobium meliloti genome. pTH1522 replicates in Escherichia coli and can be transferred to, but cannot replicate in, S. meliloti. Homologous recombination of the DNA fragments cloned in pTH1522 into the S. meliloti genome generates transcriptional fusions to either the reporter genes gfp(+) and lacZ or gusA and rfp, depending on the orientation of the cloned fragment. Over 12,000 fusion junctions in 6,298 clones were identified by DNA sequence analysis, and the plasmid clones were recombined into S. meliloti. Reporter enzyme activities following growth of these recombinants in complex medium (LBmc) and in minimal medium with glucose or succinate as the sole carbon source allowed the identification of genes highly expressed under one or more growth condition and those expressed at very low to background levels. In addition to generating reporter gene fusions, the vector allows Flp recombinase-directed deletion formation and gene disruption, depending on the nature of the cloned fragment. We report the identification of genes essential for growth on complex medium as deduced from an inability to recover recombinants from pTH1522 clones that carried fragments internal to gene or operon transcripts. A database containing all the gene expression activities together with a web interface showing the precise locations of reporter fusion junctions has been constructed (www.sinorhizobium.org).
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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".