JoVE Encyclopedia of Experiments
Microbiology
0 views • 3:03 min • February 2nd, 2026
Begin with an Agrobacterium tumefaciens culture grown in medium supplemented with two antibiotics to prevent contamination and ensure plasmid retention.
The bacteria contain a plant expression vector carrying a gene of interest and an antibiotic-resistance gene.
They also contain a helper plasmid with a region of virulence genes required for delivering the gene of interest into plant cells.
Transfer a portion into a tube containing a buffered medium supplemented with acetosyringone, a plant-derived signal molecule, and the antibiotics. Incubate with shaking.
Acetosyringone diffuses into the cell, triggers a signaling pathway, and activates expression of virulence genes required for the transfer of the gene of interest.
Transfer the culture, centrifuge to collect the bacteria, and discard the supernatant.
Add a salt buffer and wash the cells repeatedly to remove the added antibiotics.
Resuspend the bacteria in the buffer with acetosyringone and incubate without shaking to maintain virulence gene expression.
Use the culture immediately for efficient transformation.
Use a tip to pick a positive colony from the LB plate and inoculate the cells into a glass tube containing 5 milliliters of LB me
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