Comparison of injecting DNA vs RNA / pronucleus vs. cytoplasm

 The CRISPR guide RNA and Cas9 mRNA can be directly injected, or, transiently expressed from DNA plasmids.  Here's a head-to-head comparison of the resulting efficiencies when injecting DNAs vs. RNAs into mouse embryos, and whether to go into the pronucleus or the cytoplasm.

Bottom line:  RNA injection into the cytoplasm is the most efficient way for generating both efficient mutagenesis and numbers of live pups.   However, plasmid DNA works too.   For me, the main concern with RNA is the reliability of the RNA synthesis to provide consistency across injections.

Sci Rep. 2014 Mar 28;4:4513.  

Validation of microinjection methods for generating knockout mice by CRISPR/Cas-mediated genome engineering.

Horii T, Arai Y, Yamazaki M, Morita S, Kimura M, Itoh M, Abe Y, Hatada I.