I want to perform a bombardment using biolistics transfection methods for genome editing in wheat. We have Cas9 protein and sgRNA as Ribonucleoprotein complexes and I would like to coat this RNPs on gold nanoparticles for biolistics delivery in calli without using plasmid.
Does anyone have the protocols for coating proteins/RNPs in monocot plant? Please do share me your experience.
Thank you so much in advance and much appreciated.
Please also see this discussion on RG:
"Coating of gRNA and Cas9 protein with gold/tungsten for gene gun mediated transformation?"
https://www.researchgate.net/post/Coating_of_gRNA_and_Cas9_protein_with_gold_tungsten_for_gene_gun_mediated_transformation#view=58dd3fd63d7f4bfa2e5bb2b1
Hi Omar,
I think that it should has an isoelectric point because one part of the Ribonucleoprotein complex is protein (Cas9).
Hi Heikham Russiachand Singh,
When you said gold 'nanoparticles', did you mean the regular gold particles (micro-meter in diameter) or another kind of gold particles which is 'nano-size'? In plants, we routinely use micro-particles. Ex. 1.0 μm diameter gold particles for use with the PDS-1000/He and Helios gene gun systems from Bio-Rad 9 (see below picture).
But, people also use nano-particles for bombardment. For example, see attached paper:
"Nano-biolistics: a method of biolistic transfection of cells and tissues using a gene gun with novel nanometer-sized projectiles"
Hi Heikham Russiachand Singh,
Or, are you talking about something like Gold Mesoporous Silica Nanoparticle Mediated Protein and DNA Co-delivery to Plant Cells.
See attached paper.
Hi Yuan-Yeu, Yes I am talking about the regular gold particles 0.6 μm.