Initial Colony Morphology-Based Selection for iPS Cells Derived from Adult Fibroblasts Is Substantially Improved by Temporary UTF1-Based Selection
Bibliographic record
Abstract
BACKGROUND: Somatic cells can be reprogrammed into induced pluripotent stem (iPS) cells. Recently, selection of fully reprogrammed cells was achieved based on colony morphology reminiscent of embryonic stem (ES) cells. The maintenance of pluripotency was analysed. METHODOLOGY/PRINCIPAL FINDINGS: Clonal murine iPS cell line TiB7-4, which was derived from adult fibroblasts, was analysed for maintenance of pluripotency. Colony morphology, expression of pluripotency factors and stage specific embryonic antigen 1 (SSEA1) were analysed by real time PCR and flow cytometry. We found the iPS cell line TiB7-4 and its subclones to be rather diverse and exhibiting a tendency towards spontaneous differentiation and loss of pluripotency independent of their initial colony morphology. In contrast an undifferentiated transcription factor 1 (UTF1) promoter-driven G418 (Neo) resistance significantly improved the quality of these iPS cells. After selection with UTF-Neo for two weeks iPS subclones could be stably maintained for at least 40 passages in culture and differentiate into all three germ layers. As control, a construct expressing G418 resistance under the control of the ubiquitously active SV40 early promoter formed subclones with different colony morphology. Some of these subclones could be cultured for at least 12 passages without loosing their pluripotency, but loss of pluripotency eventually occured in an unpredictable manner and was independent of the subclones' initial morphology and SSEA1 expression. A UTF-Neo-based selection of a whole population of TiB7-4 without further subcloning resulted in the generation of cultures with up to 99% SSEA1 positive cells under stringent selection conditions. CONCLUSIONS: Our data indicate that temporary selection using a genetic UTF1-based system can generate homogenous pluripotent iPS cells that can be maintained without permanent selection pressure.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.001 | 0.000 |
| Meta-epidemiology (narrow) | 0.001 | 0.000 |
| Meta-epidemiology (broad) | 0.001 | 0.001 |
| Bibliometrics | 0.001 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.001 | 0.000 |
| Open science | 0.001 | 0.001 |
| Research integrity | 0.001 | 0.001 |
| Insufficient payload (model declined to judge) | 0.004 | 0.004 |
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 source (direct Gemma or distilled Codex), 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".