CRISPR 101: Mammalian Expression Systems and Delivery Methods
Type
Blog Post
Published
Sept. 24, 2015, 2:30 p.m.
...vivo viral delivery.
Cas9 mRNA and gRNA
Plasmids containing gRNA and Cas9 are used in in vitro transcription....
Cas9-gRNA ribonucleoprotein complexes
Purified Cas9 protein and in vitro transcribed gRNA are combined...or HDR.
EGFP validation of sgRNAs: Can evaluate component delivery and sgRNA efficacy by cloning in genome...can help you make precise point mutations near the gRNA target site.
Choose an appropriate expression system...you will need to deliver. Minimally, one or more sgRNAs and Cas9 are required for any application. If you... or inducible. U6 promoter is typically used for gRNA. May contain reporter gene (e.g. GFP) to identify...lines.
Transient or stable expression of Cas9 and/or gRNA in a mammalian cell line that can be transfected...