Re:Confused about how this works (Score 2)

CRISPER works via the connection between the CRISPER and the enzyme Cas9 (that's why it's actually called CRISPER-Cas). I will simply talk about the synthetic bio version, not the naturally occurring. So let's say you want to do a targeted genome edit at a specific point in the genome. To do this, you need to make your CRISPER RNA as follows: you have the "Cas section" (crRNA) and "localization" section (tracrRNA). The way it works is that you design the localization section to be complimentary to the DNA you wish to cut (people have worked out how to make RNA strands bind to the DNA in specific points). You then inject some of the Cas9 enzyme which binds to the "Cas section" (or in bio terms you tend to say the CRISPER promotes the enzyme) and holds it steady. This enzyme, when held near the DNA, will cause it to cut. So now you have a tool for cutting. Let's say you want to cut out a gene. You can just make two CRISPERs, one before the gene and one after. Inject the CRISPERs, inject the Cas9, and that gene is gone. But I forgot to mention the important part: you have to fix the break! You can then do genome editing by pairing this with something called Homology Directed Repair to put any sequence you want in there. There are two questions to address: why is this better than what we had before and how good is it? Before the main tools were restriction enzymes and TALENs. Restriction enzymes had a specific sequence they could cut (for example: BamHI cuts GGATCC). You can use this with some ligation (gluing back together) tools to attempt to do this stuff, but it's quite hard if you cannot specifically target an area. TALENs I haven't really used (I'm too new, my lab just started by telling me about CRISPERs) but from what I know people say they weren't reliable and were hard to make. However, CRISPERs are just an RNA sequence. There are online tools that help you design the CRISPERs as long as you know the DNA you want to target (which you hopefully do thanks to the magic of genome sequences). Making this RNA is a simple technique and once you get it, you basically have an infinite amount of the CRISPERs in case you mess up. You just take that, inject it with the Cas9 enzyme that you buy, and it works surprisingly well. Thus it has taken over in genome editing for many species. I know that in zebrafish it has become the tool to use, and from my reading it sounds like it's being used a lot in mouse, plants, and flies as well. It's just easy and reliable. Will this mean that you will use this to change your baby's eye color? I don't know, but from what I hear about its effectiveness in the zebrafish lab, although you get it working a good amount of the time, it's still not all of the time. People are okay if it doesn't correctly re-glue the DNA in 10% of your fish embryos, but would you be okay with it failing 10% on your baby? I'm not sure.