Cloud-Based Design of Short Guide RNA (sgRNA) Libraries for CRISPR Experiments

CRISPR/Cas-based genome editing in any biological application requires the evaluation of suitable genomic target sites to design efficient reagents. Considerations for the design of short guide (sg) RNAs include the assessment of possible off-target activities, the prediction of on-target efficacies and mutational outcome. Manual design of sgRNAs taking into account these parameters, however, remains a difficult task. Thus, computational tools to design sgRNA reagents from small scale to genome-wide libraries have been developed that assist during all steps of the design process. Here, we will describe practical guidance for the sgRNA design process using the web-based tool E-CRISP used in the design of individual sgRNAs. E-CRISP (www.e-crisp.org) has been the first web-based sgRNA design tool and uniquely features simple, yet efficient, scoring schemes in combination with fast evaluation and simple usage. We will also discuss the installation of a dockerized version of CRISPR Library Designer (CLD) that can be deployed locally or in the cloud to support the end-to-end design of sgRNA libraries for more than 50 different organisms. CLD was built upon E-CRISP to further increase the scope of sgRNA design to more experimental modalities (CRISPRa/i, Cas12a, all possible protospacer adjacency motifs) offering the same flexibility as E-CRISP, plus the scalability through local and cloud installation. Together, these tools facilities the design of small and large-scale CRISPR/Cas experiments.