Agrobacterium tumefaciens-Mediated Genetic Transformation of the Ect-endomycorrhizal Fungus Terfezia boudieri
Abstract
:1. Introduction
2. Materials and Methods
2.1. Terfezia Boudieri Culture
2.2. Hygromycin and Geneticin Sensitivity Test
2.3. A. tumefaciens Culture and Plasmid Transfection/Electroporation
2.4. T. boudieri Infection and Co-Cultivation with A. tumefaciens
2.5. Selection of Fungal Transformants
2.6. Microscopic Analysis
2.7. Fungal Genomic DNA Extraction and PCR Analysis
2.8. Fungal Transgene Insertion and Stability
2.9. Inoculation of the Host Plant with Transformed Hyphae
2.10. Data Collection and Statistical Analysis
3. Results
3.1. Sensitivity of T. boudieri Mycelium to Antibiotics and the Effect of Acetosyringone on Transformation Efficiency
3.2. Effects of A. tumefaciens Cell Density and Infection Time on T. boudieri Transformation
3.3. The Effect of Pre-Selection Cultivation Time on Transformation Efficiency
3.4. The Effect of the Mycelia Developmental Stage on Transformation Efficiency
3.5. Molecular Verification of Putative Transformants
3.6. Genetic Stability of T. boudieri Transformants
3.7. Expression of Reporter Genes in Free-Living Mycelia and in Mycorrhizal Association
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Satish, L.; Kamle, M.; Keren, G.; Patil, C.D.; Yehezkel, G.; Barak, Z.; Kagan-Zur, V.; Kushmaro, A.; Sitrit, Y. Agrobacterium tumefaciens-Mediated Genetic Transformation of the Ect-endomycorrhizal Fungus Terfezia boudieri. Genes 2020, 11, 1293. https://doi.org/10.3390/genes11111293
Satish L, Kamle M, Keren G, Patil CD, Yehezkel G, Barak Z, Kagan-Zur V, Kushmaro A, Sitrit Y. Agrobacterium tumefaciens-Mediated Genetic Transformation of the Ect-endomycorrhizal Fungus Terfezia boudieri. Genes. 2020; 11(11):1293. https://doi.org/10.3390/genes11111293
Chicago/Turabian StyleSatish, Lakkakula, Madhu Kamle, Guy Keren, Chandrashekhar D. Patil, Galit Yehezkel, Ze’ev Barak, Varda Kagan-Zur, Ariel Kushmaro, and Yaron Sitrit. 2020. "Agrobacterium tumefaciens-Mediated Genetic Transformation of the Ect-endomycorrhizal Fungus Terfezia boudieri" Genes 11, no. 11: 1293. https://doi.org/10.3390/genes11111293
APA StyleSatish, L., Kamle, M., Keren, G., Patil, C. D., Yehezkel, G., Barak, Z., Kagan-Zur, V., Kushmaro, A., & Sitrit, Y. (2020). Agrobacterium tumefaciens-Mediated Genetic Transformation of the Ect-endomycorrhizal Fungus Terfezia boudieri. Genes, 11(11), 1293. https://doi.org/10.3390/genes11111293