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AAT Bioquest

What are the main components of CRISPR-Cas9 technique?

Posted June 22, 2020


Answer

CRISPR-Cas9 genome editing includes two key components: a single-guide RNA (gRNA) and a CRISPR-associated endonuclease (Cas). The sgRNA and Cas9 are combined into a ribonucleoprotein complex when they are used in CRISPR experiments.

  • Cas9: It is an endonuclease enzyme which functions as a pair of molecular scissors to cut the target DNA sequence.
  • sgRNA: Single-guide RNA is the combination of tracrRNA and crRNA. crRNA contains two main parts, the spacer sequence that directs the complex to the target DNA and a region that binds to tracrRNA. When tracrRNA binds to crRNA, a functional guide RNA is formed for Cas9 to recognize. Multiple crRNAs can be incorporated into a crRNA array and then packaged with tracrRNA to form a sgRNA.
Additional resources

Helixyte™ Green *10,000X Aqueous PCR Solution*

6-ROX glycine *25 uM fluorescence reference solution for PCR reactions*

Doudna, J. A., & Charpentier, E. (2014). The new frontier of genome engineering with CRISPR-Cas9. Science, 346(6213), 1258096.

Ran, F. A., Hsu, P. D., Wright, J., Agarwala, V., Scott, D. A., & Zhang, F. (2013). Genome engineering using the CRISPR-Cas9 system. Nature protocols, 8(11), 2281.