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

Ana Clara Ferreira, Bárbara A. Rebelo, Rita Abranches

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Abstract

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.

Original languageEnglish
Pages (from-to)669-675
Number of pages7
JournalPlant Cell, Tissue and Organ Culture
Volume153
Issue number3
DOIs
Publication statusPublished - Jun 2023

Keywords

  • Agrobacterium
  • Cultured cells
  • Medicago
  • Model legume
  • Plant-based system
  • RBD
  • SARS-CoV-2
  • Stable transformation

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