A TaqMan real-time PCR assay targeting the cytochrome o ubiquinol oxidase subunit II gene for detection of several pathovars of <i>Pseudomonas syringae</i>
Bibliographic record
Abstract
Abstract The identification and detection of eight pathovars of Pseudomonas syringae, bacterial pathogens of several important agricultural plants, was achieved by TaqMan real-time polymerase chain reaction of a specific DNA fragment of the cytochrome o ubiquinol oxidase gene. Under optimal real-time PCR conditions, the selected primers and probe were specific for the detection of pathovars syringae, tomato, maculicola, tabaci, atropurpurea, phaseolicola, pisi and glycinea. Two pathovars (coriandricola and morsprunorum) tested could be differentiated from the other eight due to a single nucleotide mismatch. Thirty other Pseudomonas strains and 20 non-Pseudomonas strains were negative. The real-time PCR assay detected 100 fg of DNA and 4.5 × 103 P. syringae colony forming units per millilitre (four cells per reaction). In growth chamber experiments, tomato plants were inoculated using strain DC3000 and assayed by TaqMan real-time PCR. Serial dilution of leaf extracts spiked with lambda DNA and processed by real-time PCR indicated the presence of inhibitors. A 1:10 dilution of the crude extract reduced threshold cycles to those of milliQ water spiked with the same amount of lambda DNA. The TaqMan real-time assay consistently detected the pathogen in inoculated tomato leaves after a 1:10 dilution of crude extracts. TaqMan real-time results were validated by dilution-plating of leaf extracts and conventional PCR using the same primer set. This assay offers real-time monitoring of the targeted amplicon with high specificity and sensitivity, with no post-amplification analysis needed. This reduces opportunity for contamination of the reaction mixtures with target DNA, making this real-time PCR assay more reliable than conventional PCR. However, for routine diagnosis of the detected pathovars under greenhouse or field conditions, optimization of the assay might be required.
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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 | 0.001 | 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".