A Deep Learning-Based System (Microscan) for the Identification of Pollen Development Stages and Its Application to Obtaining Doubled Haploid Lines in Eggplant
Abstract
1. Introduction
2. Materials and Methods
2.1. Plant Material
2.2. Experimental Layout and Workflow
2.3. Phase 1: Model Training
2.3.1. Microspore and Pollen Isolation
2.3.2. Digital Data Image Acquisition
2.3.3. Image Labelling
2.3.4. Preprocessing
2.3.5. Predictive Model
2.4. Phase 2: In Vitro Androgenesis Induction Test Using the Anther Selection Software
2.4.1. E6 Protocol
2.4.2. Cb Protocol
2.4.3. Flow Cytometry
2.4.4. Single Primer Enrichment Technology (SPET) Genotyping
3. Results
3.1. Phase 1: Model Training
3.2. Phase 2: In Vitro Androgenesis Induction Test Using the Microscan
4. Discussion
5. Conclusions
Author Contributions
Funding
Conflicts of Interest
References
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Medium | MS + Vitamins (g/L) | Prepared C (g/L) | Prepared R (g/L) | Sucrose (g/L) | Zeatin Riboside (mg/L) | Kinetin (mg/L) | 2,4-D (mg/L) | Vitamin B12 (mg/L) | Gelrite (g/L) | Bacto-Agar (g/L) |
---|---|---|---|---|---|---|---|---|---|---|
E0 | 2.20 | - | - | 15.00 | - | - | - | - | 7.00 | - |
E6 | 2.20 | - | - | 15.00 | 2.00 | - | - | - | 7.00 | - |
Cb | - | 4.55 | - | 120.00 | - | 5.00 | 5.00 | 0.20 | - | 8.00 |
R | - | - | 4.55 | 30.00 | - | 0.10 | - | - | - | 8.00 |
Class | Total Cellular Events | Average Precision (%) |
---|---|---|
Tetrad | 290 | 87.40 |
Young Microspore | 641 | 86.28 |
Medium Microspore | 1893 | 81.97 |
Vacuolated Microspore | 1185 | 87.60 |
Young Pollen | 1876 | 82.32 |
Mature Pollen | 2186 | 92.19 |
mAP | 86.30 |
Protocol E6 | Protocol Cb | |||||
---|---|---|---|---|---|---|
Size Range/Genotype | Anthers (n) | Response (%) | Type of Response | Anthers (n) | Response (%) | Type of Response |
3.5–4 mm | ||||||
BC3 17-8 | 45 | 0.00 ± 0.00 | - | 45 | 0.00 ± 0.00 | - |
BC3 17-19 | 45 | 0.00 ± 0.00 | - | 45 | 0.00 ± 0.00 | - |
BC3 17-4 | 45 | 0.00 ± 0.00 | - | 45 | 0.00 ± 0.00 | - |
5.5–6 mm | ||||||
BC3 17-8 | 45 | 75.30 ± 0.04 | Embryo | 45 | 78.40 ± 0.04 | Callus |
BC3 17-19 | 45 | 60.40 ± 0.05 | Embryo | 45 | 80.50 ± 0.04 | Callus |
BC3 17-4 | 45 | 40.30 ± 0.05 | Embryo | 45 | 71.20 ± 0.05 | Callus |
>6 mm | ||||||
BC3 17-8 | 45 | 0.00 ± 0.00 | - | 45 | 3.20 ± 0.02 | Somatic callus |
BC3 17-19 | 45 | 0.00 ± 0.00 | - | 45 | 4.00 ± 0.02 | Somatic callus |
BC3 17-4 | 45 | 0.00 ± 0.00 | - | 45 | 4.30 ± 0.03 | Somatic callus |
Genotype | Embryos (n) | Acclimatized Plants (n) | n | n + 2n | 2n |
---|---|---|---|---|---|
BC3 17-8 | 42 | 12 | 9 | 3 | 0 |
BC3 17-19 | 26 | 12 | 8 | 1 | 3 |
BC3 17-4 | 9 | 7 | 6 | 1 | 0 |
Accession/Offspring | n | Missing SNPs | Heterozygosity (%) |
---|---|---|---|
ELE BC3 17-19 | 1 | 0.00 | 100.00 |
BC4 (ELE BC3 17-19 × MEL3) | 6 | 4.00 (0.00–6.00) | 55.07 (39.06–65.66) |
DH (ELE BC3 17-19 doubled haploids) | 10 | 5.00 (0.00–12.00) | 1.59 (0.19–3.95) |
ELE BC3 17-4 | 1 | 0.00 | 100.00 |
BC4 (ELE BC3 17-4 × MEL3) | 5 | 0.40 (0.00–2.00) | 53.87 (13.35–92.66) |
DH (ELE BC3 17-4 doubled haploids) | 7 | 1.14 (0.00–4.00) | 0.67 (0.19–1.70) |
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García-Fortea, E.; García-Pérez, A.; Gimeno-Páez, E.; Sánchez-Gimeno, A.; Vilanova, S.; Prohens, J.; Pastor-Calle, D. A Deep Learning-Based System (Microscan) for the Identification of Pollen Development Stages and Its Application to Obtaining Doubled Haploid Lines in Eggplant. Biology 2020, 9, 272. https://doi.org/10.3390/biology9090272
García-Fortea E, García-Pérez A, Gimeno-Páez E, Sánchez-Gimeno A, Vilanova S, Prohens J, Pastor-Calle D. A Deep Learning-Based System (Microscan) for the Identification of Pollen Development Stages and Its Application to Obtaining Doubled Haploid Lines in Eggplant. Biology. 2020; 9(9):272. https://doi.org/10.3390/biology9090272
Chicago/Turabian StyleGarcía-Fortea, Edgar, Ana García-Pérez, Esther Gimeno-Páez, Alfredo Sánchez-Gimeno, Santiago Vilanova, Jaime Prohens, and David Pastor-Calle. 2020. "A Deep Learning-Based System (Microscan) for the Identification of Pollen Development Stages and Its Application to Obtaining Doubled Haploid Lines in Eggplant" Biology 9, no. 9: 272. https://doi.org/10.3390/biology9090272
APA StyleGarcía-Fortea, E., García-Pérez, A., Gimeno-Páez, E., Sánchez-Gimeno, A., Vilanova, S., Prohens, J., & Pastor-Calle, D. (2020). A Deep Learning-Based System (Microscan) for the Identification of Pollen Development Stages and Its Application to Obtaining Doubled Haploid Lines in Eggplant. Biology, 9(9), 272. https://doi.org/10.3390/biology9090272