I'm new to using CRISPR, and all the protocols I'm reading are just telling me to order the oligos for the primers for gRNA synthesis. Is this what everyone does, or is there a way to feasibly make your own oligos?
Hello Drake,
I think you can easily order the gRNA oligos same way you order it for Primer sequences.
Hey Drake,
It is a very easy process to produce your own gRNA with the proper equipment and protocols in place. Some time back in the genome engineering lab that I was working in, we produced them ourselves. However, it is a somewhat tedious process that takes time and mostly just pays off if you have to produce a lot of them on a regular basis. Also, working with RNA requires some caution.
So, it is "easy" to produce gRNAs yourself. However, I would recommend just ordering them from ITD for example. It is uncomplicated and saves a lot of time that you can spend otherwise in the lab.
All the best!
Hey Glen Drake
Since you are beginner to CRISPR, if your plan is to edit your gene of interest in an easily transfected cell line, I would suggest you to order the sgRNA oligos from IDT, anneal and clone in sgRNA plasmid, confirm the sequence fidelity with sanger sequence and co-transfect with Cas9 plasmid. While if your plan to generate a mouse model you can either use the same spacer "you have tested previously in vitro" ,clone it in under a T7 promoter and synthesize your own sgRNA, or you can directly ordered the synthesized shRNA as RNA from Synthon.
Hopefully this help!