Transient transformation in Cas9-OE A. thaliana with CLCrV-sgRNA
This protocol is extracted from research article:
Heritable gene editing using FT mobile guide RNAs and DNA viruses
Plant Methods, Feb 17, 2021; DOI: 10.1186/s13007-021-00719-4

For transient expression and virus inoculation, all CLCrV-AtU6-26::sgRNA vectors and CLCrV-B were introduced into A. tumefaciens strain GV3101. The Agrobacterium cultures were inoculated in a 5 mL LB medium (containing 50 μg/mL kanamycin and 50 μg/mL rifampicin) and grown overnight in a 28 °C shaker. Agrobacterium cultures were harvested and resuspended in infiltration buffer (10 mM MgCl2, 10 mM MES and 200 μM acetosyringone), adjusted to an optical density at 600 nm of 1.0, and incubated at room temperature for 3 to 4 h. For virus inoculation, Agrobacterium cultures containing CLCrV-B and CLCrV-A or their derivatives were mixed at a 1:1 ratio. The mixed Agrobacterium solutions were infiltrated into A. thaliana leaves with a 1 ml syringe.

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