The CRISPR tool kit for genome editing and beyond

Abstract

CRISPR is becoming an indispensable tool in biological research. Once known as the bacterial immune system against invading viruses, the programmable capacity of the Cas9 enzyme is now revolutionizing diverse fields of medical research, biotechnology, and agriculture. CRISPR-Cas9 is no longer just a gene-editing tool; the application areas of catalytically impaired inactive Cas9, including gene regulation, epigenetic editing, chromatin engineering, and imaging, now exceed the gene-editing functionality of WT Cas9. Here, we will present a brief history of gene-editing tools and describe the wide range of CRISPR-based genome-targeting tools. We will conclude with future directions and the broader impact of CRISPR technologies.

Applying the latest advances in genomics and phenomics for trait discovery in polyploid wheat

“…Gene‐editing systems such as TALENs and CRISPR‐Cas9 have now been demonstrated to function in wheat including the production of transgene‐free edited lines (Wang et al ., ; Zhang et al ., ; Rey et al ., ). Beyond the most common use of CRISPR‐Cas9, to induce small deletions that lead to frame‐shifts and knock‐outs, this may involve more complex editing such as targeted gene insertion or specific base editing (for review, see Adli, ). In theory, the use of guide‐mediated gene editing could produce triple homoeolog knock‐outs in a single transgenic event, although efficiencies remain low (Zhang et al ., ).…”

Design of genomic signatures for pathogen identification and characterization

“…Another technological advance is seen in the recent breakthroughs in gene and genome synthesis (Gibson et al, 2008) and editing techniques such as CRISPR/Cas9 (Adli, 2018). Not only do we need to deal with emerging natural viruses from every remote corner of the planet, but now we also need to deal with the fact that for rapidly decreasing amounts of money, it is possible to synthesize combinatorial versions of any DNA one might wish to (re) create.…”

Guidelines for optimized gene knockout using CRISPR/Cas9

“…With the same aim of reducing Cas9 off-target activity, several Cas9 variants whose editing activity can be irreversibly or reversibly programmed are now also available. Finally, engineered Cas9 variants with novel PAM specificities enlarge the edition spectrum to previously inaccessible sites [69,70]. …”