Hello,

I'm preparing to preform CRISPR/Cas KO by microinjection in the silkworm Bombyx mori. I'm ordering synthetic sgRNA and using SpCas9 nuclease for this experiment.

I've read that when delivering RNPs by microinjection it is important to always resuspend and dilute gRNA in microinjection buffer. However, little information from is available from companies regarding microinjection (compared to cell plates). Has anyone any advice on this in B. mori or other insect eggs? There are several suggested buffers available online (two below*) but I'm not sure if these are specific to the cell type (zebrafish and Xenopus eggs vs. insect eggs).

I haven't seen anything regarding resuspending Cas9 in anything other than it's provided buffer but was curious if microinjection buffer would be preferable?

Any advice?

*Generic microinjection buffer - 10 mM Tris-HCl, 0.1 mM EDTA, pH 8.0

*Tsubota and Sezutsu (2017) - 0.5 mM potassium phosphate buffer (pH 7.0), 5 mM KCl [Genome Editing of Silkworms]