CRISPR

Clustered regularly interspaced short palindromic repeats (CRISPR) is quickly becoming the most popular tool in the genome editing arena. CRISPR has been adapted for use in mammalian cells from bacterial anti-bacteriophage immune systems.

The TriLink CRISPR tool kit includes two components: the first is a CRISPR nuclease such as Cas9 mRNA or Cas12a mRNA and the second is a guide RNA. In nature, the Cas9 guide consists of a CRISPR RNA (crRNA), as well as a trans-acting CRISPR RNA (tracrRNA). For simplicity and increased potency, researchers fused these two RNAs into a single RNA of ~100 nucleotides, which they called a single guide RNA (sgRNA).

In contrast to Cas9, Cas12a utilizes a single guide RNA of ~42 nucleotides. When Cas9 or Cas12a mRNA and a sgRNA are co-transfected into cells, the guide strand directs the Cas9 or Cas12a protein to a specific location in the genome, where Cas9 then creates a double stranded break. Cas9 Nickase mRNA encodes Cas9 with a D10A mutation, which causes a single stranded nick instead of a double stranded break. This may result in fewer off-target effects.

TriLink offers several mRNAs for CRISPR that are unmodified or modified with 5-methoxyuridine (5moU) to reduce innate immune responses. These have been sequence optimized with the help of leading researchers in the field of optimal expression. When you are ready to move your research to the clinic, TriLink offers cGMP synthesis of guide strands and nuclease mRNAs in our purpose built cGMP facility.