Abstract
This transformation procedure generates, with high efficiency (70–90%), hairy roots in cultivars, landraces and accessions of Phaseolus vulgaris (common bean) and other Phaseolus spp. Hairy roots rapidly develop after wounding young plantlets with Agrobacterium rhizogenes, at the cotyledon node, and keeping the plants in high-humidity conditions. Callogenesis always precedes hairy-root formation, and after 15 days, when roots develop at wounded sites, the stem with the normal root is cleaved below the hairy root zone. Transgenic roots and nodules co-transformed with a binary vector can be easily identified using a reporter gene. This procedure, in addition to inducing robust transgenic hairy roots that are susceptible to being nodulated by rhizobia and to fixing nitrogen efficiently, sets the foundation for a high-throughput functional genomics approach on the study of root biology and root–microbe interactions. This protocol can be completed within 30 days.
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Acknowledgements
This research was partially supported by CONACYT 42562-Q and by Dirección General de Asuntos del Personal Académico IN-215805-2 grants. We thank Dr. José Luis Reyes and Oswaldo Valdés L. for pTdTRNAi vector construction and Olivia Santana for technical assistance.
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Estrada-Navarrete, G., Alvarado-Affantranger, X., Olivares, JE. et al. Fast, efficient and reproducible genetic transformation of Phaseolus spp. by Agrobacterium rhizogenes. Nat Protoc 2, 1819–1824 (2007). https://doi.org/10.1038/nprot.2007.259
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DOI: https://doi.org/10.1038/nprot.2007.259
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