A simplified protocol for Agrobacterium-mediated transformation of cell suspension cultures of the model species Medicago truncatula A17

This manuscript describes a unique protocol for the rapid transformation of Medicago truncatula A17 cell suspension cultures mediated by Agrobacterium tumefaciens. Medicago cells were collected on day 7 of the growth curve, which corresponded to the beginning of the exponential phase. They were then co-cultured with Agrobacterium for 3 days before being spread onto a petri dish with appropriate antibiotic selection. The Receptor Binding Domain of the Spike protein of SARS-CoV-2 was used as a model to develop this protocol. The presence of the transgene was assessed using PCR, and the integrity of the product was evaluated by SDS-PAGE and Western-blotting. Key messageWe have developed an effective and simple protocol for the transformation of Medicago truncatula A17. The success of this protocol relies on proper handling of the cultures and is based solely on co-cultivating plant cells with Agrobacterium. This method does not require experienced professionals or specialized equipment, making it faster, easier, and more affordable than other available protocols. Additionally, there is no need for protoplast preparation or vacuum application.