| Autor: |
Tanner M. Cook, Eva Biswas, Somak Dutta, Siddique I. Aboobucker, Sara Hazinia, Thomas Lübberstedt |
| Jazyk: |
angličtina |
| Rok vydání: |
2024 |
| Předmět: |
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| Zdroj: |
Plant Methods, Vol 20, Iss 1, Pp 1-14 (2024) |
| Druh dokumentu: |
article |
| ISSN: |
1746-4811 |
| DOI: |
10.1186/s13007-023-01132-9 |
| Popis: |
Abstract Background Strategies to understand meiotic processes have relied on cytogenetic and mutant analysis. However, thus far in vitro meiosis induction is a bottleneck to laboratory-based plant breeding as factor(s) that switch cells in crops species from mitotic to meiotic divisions are unknown. A high-throughput system that allows researchers to screen multiple candidates for their meiotic induction role using low-cost microfluidic devices has the potential to facilitate the identification of factors with the ability to induce haploid cells that have undergone recombination (artificial gametes) in cell cultures. Results A data analysis pipeline and a detailed protocol are presented to screen for plant meiosis induction factors in a quantifiable and efficient manner. We assessed three data analysis techniques using spiked-in protoplast samples (simulated gametes mixed into somatic protoplast populations) of flow cytometry data. Polygonal gating, which was considered the “gold standard”, was compared to two thresholding methods using open-source analysis software. Both thresholding techniques were able to identify significant differences with low spike-in concentrations while also being comparable to polygonal gating. Conclusion Our study provides details to test and analyze candidate meiosis induction factors using available biological resources and open-source programs for thresholding. RFP (PE.CF594.A) and GFP (FITC.A) were the only channels required to make informed decisions on meiosis-like induction and resulted in detection of cell population changes as low as 0.3%, thus enabling this system to be scaled using microfluidic devices at low costs. |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
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