CRISPR+101+ebook_20190829.pdf - CRISPR 101 Your Guide to...

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CRISPR 101 Your Guide to Understanding CRISPR
2 CRISPR 101: YOUR GUIDE TO UNDERSTANDING CRISPR SYNTHEGO.COM/CONTACT | (888) 611-6883 20190829 Introduction to Genome Editing Genome editing involves the deletion, insertion, or modification of specific DNA sequences in the genome. For many years, researchers had been trying to develop easy and cost-effective genome editing tools to address problems across a wide spectrum of fields. For instance, gene therapy in humans could progress rapidly if one could simply eliminate the gene responsible for a certain genetic disorder. In agriculture, manipulating plant DNA could be used to optimize crop yields 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, a robust molecular tool that can edit DNA at virtually any locus. CRISPR technology is igniting a revolution across the life sciences and is quickly becoming a standard tool in many labs. Given its ease-of-use and versatility, CRISPR is already being used 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 the fundamentals of CRISPR.
3 CRISPR 101: YOUR GUIDE TO UNDERSTANDING CRISPR SYNTHEGO.COM/CONTACT | (888) 611-6883 20190829 Genome Editing Tools Before CRISPR Although CRISPR has now become synonymous with gene editing, it is not the first technology developed to edit DNA. Genome editing techniques initially emerged with the discovery of restriction enzymes and meganucleases. The possibility of precision gene editing was made clearer with the discovery of zinc-finger nucleases (ZFNs). The ZFN method involves engineering an enzyme with both a zinc finger DNA-binding domain and a restriction endonuclease domain. The zinc finger domain is composed of 3-base pair site on DNA designed to target and bind to specific sequences of DNA, and the nuclease domain cleaves the DNA at the desired site. Although ZFN editing represented the first breakthrough in site-specific genome engineering, they have several limitations. In addition to exhibiting off-target effects, ZFNs are expensive and time-consuming to engineer. Furthermore, their inefficiency limits their practical application to only one genomic edit at a time. Many years after ZFNs made their debut, a similar method known as transcription activator-like effector nucleases (TALENs) was developed. The TALENs method utilizes engineered enzymes containing a DNA binding domain and a separate DNA-cleaving domain, similar to the ZFNs method. However, TALENs have an advantage over ZFNs because they are more flexible; their DNA-binding domains can target a wider range of sequences. Although they are easier to design than ZFNs, TALENs are expensive to produce.

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