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CRISPR 101Your Guide to Understanding CRISPR
2CRISPR 101: YOUR GUIDE TO UNDERSTANDING CRISPRSYNTHEGO.COM/CONTACT | (888) 611-688320190829Introduction to Genome EditingGenome editing involves the deletion, insertion, or modification of specificDNA sequences in the genome. For many years, researchers had been trying todevelop easy and cost-effective genome editing tools to address problems acrossa wide spectrum of fields. For instance, gene therapy in humans could progressrapidly if one could simply eliminate the gene responsible for a certain geneticdisorder. In agriculture, manipulating plant DNA could be used to optimize cropyields and control plant diseases. Similarly, bacterial genomes could be fine-tuned to increase their product yields in several industrial applications.Finally, the efforts of researchers paid off with the development of CRISPR, arobust molecular tool that can edit DNA at virtually any locus. CRISPR technologyis igniting a revolution across the life sciences and is quickly becoming a standardtool in many labs. Given its ease-of-use and versatility, CRISPR is already beingused for a variety of applications and holds a lot of promise for the future.Read on for a crash course in everything you need to know about thefundamentals of CRISPR.
3CRISPR 101: YOUR GUIDE TO UNDERSTANDING CRISPRSYNTHEGO.COM/CONTACT | (888) 611-688320190829Genome Editing Tools Before CRISPRAlthough CRISPR has now become synonymous with gene editing,it is not the first technology developed to edit DNA. Genome editingtechniques initially emerged with the discovery of restriction enzymesand meganucleases. The possibility of precision gene editing was madeclearer with the discovery of zinc-finger nucleases (ZFNs).The ZFN method involves engineering an enzyme with both a zinc fingerDNA-binding domain and a restriction endonuclease domain. The zincfinger domain is composed of 3-base pair site on DNA designed to targetand bind to specific sequences of DNA, and the nuclease domain cleavesthe DNA at the desired site. Although ZFN editing represented the firstbreakthrough in site-specific genome engineering, they have severallimitations. In addition to exhibiting off-target effects, ZFNs are expensiveand time-consuming to engineer. Furthermore, their inefficiency limitstheir practical application to only one genomic edit at a time.Many years after ZFNs made their debut, a similar method known astranscription activator-like effector nucleases (TALENs) was developed.The TALENs method utilizes engineered enzymes containing a DNAbinding domain and a separate DNA-cleaving domain, similar to the ZFNsmethod. However, TALENs have an advantage over ZFNs because theyare more flexible; their DNA-binding domains can target a wider rangeof sequences. Although they are easier to design than ZFNs, TALENs areexpensive to produce.