When to express Cas9 from the same vector as your sgRNA

A typical CRISPR knockout experiment require two components. First, a Cas9 enzyme that cuts the DNA. Second, a guide RNA (gRNA) that dictates where that cut should be made. Horizon provides an sgRNA All-In-One Vector that expresses the Cas9 and gRNA together. Alternatively, we also sell individual Cas9 and gRNA reagents that can be co-delivered or delivered successively.

So, which is format is best for you? These four questions will help you decide.

Are your cells difficult to transfect, transduce, or slow/difficult to culture?
  1. If the answer is “yes”, the sgRNA All-In-One Vector may be best for you. Since the Cas9 and sgRNA are expressed in one single reagent, you only do one transfection or transduction into your cells. In addition, with the All-In-One vector, you only do antibiotic selection with one resistance marker. If you use separate Cas9 and gRNA vectors, you’ll have to perform two antibiotic selections.
Are you following up on hits from a CRISPR screen that Horizon performed for you?
  1. If the answer is “yes”, you may prefer to proceed with the sgRNA All-In-One Vector. This product allows for an easy transition from our screening service to your follow-up analysis. For further investigation, you can order the same sequences we used for screening.
Do you already have cells that express Cas9?
  1. If you already have Cas9-expressing cells, then you may prefer to order gRNA reagents without Cas9 co-expressed. Since you already have the Cas9 expressed, you can order individual gRNAs to pair with your cells.
Are you worried about long-term expression of Cas9?
  1. If you prefer to use a DNA-free Cas9, you can pair either Cas9 mRNA or Cas9 protein with a separate gRNA reagent. This approach is for short-term expression of Cas9 and ensures that the Cas9 will not integrate into the genome. The sgRNA All-In-One Vector is provided as bacterial glycerol stock or viral particles, which can provide to long-term expression
Additional Resources
Edit-R All-in-one Lentiviral sgRNA
  • Genome-wide combined Cas9 and sgRNAs for efficient gene knockout & unparalleled specificity
CRISPR-Cas9 Gene Editing Products
  • Optimized tools for high-confidence genome engineering