The toolset for carrying out precision genome engineering with S. pyogenes CRISPR-Cas9 has expanded rapidly since its demonstrated use in mammalian cells. For a closer look at synthetic guide RNA that employs a two-RNA system, GE scientists have authored a July 2016 review article that covers many aspects of application and design of synthetic guide RNAs.
Chemical synthesis and modifications
Because solid-phase synthesis chemistry is used to generate synthetic guide RNAs, rather than enzymatic generation, there is greater flexibility in length and yield, and higher accuracy in the resulting RNAs. Whether using a two-RNA approach (with a crRNA and tracrRNA to program Cas9) or a single guide RNA (sgRNA; a long, chimeric guide RNA), synthesis also permits the addition of various modification patterns, which are primarily of interest to improve RNA stability in applications where exposure to nucleases (i.e., in vivo, electroporation) can cause degradation of the guide RNA and result in reduced gene editing efficiency.
Flexibility and ease-of-use
Synthetic guide RNAs arrive ready-to-use and are much easier to deliver to cells with high efficiency than DNA plasmids; so synthetic guide RNAs are easier to adapt to workflows ranging from single-gene knockout to HDR to high-throughput arrayed screening. Synthetic guides are also amenable to co-delivery with any type of transient Cas9 nuclease expression system (plasmid, mRNA, or protein) and demonstrate very high knockout efficiency when delivered to cells with stable Cas9 expression .
If you want to learn more, take a look at the open access paper.
Versatility of chemically synthesized guide RNAs for CRISPR-Cas9 genome editing »
Additional Resources
- Learn more about the Edit-R algorithm for highly functional and specific guide RNAs
CRISPR-Cas9 gene editing products - Learn more about the Edit-R algorithm for highly functional and specific guide RNAs
An algorithm for selecting highly functional and specific guide RNAs for CRISPR-Cas9 gene knockout - poster