Computational Tools and Resources Supporting CRISPR-Cas Experiments

Abstract

The CRISPR-Cas system has become a cutting-edge technology that revolutionized genome engineering. The use of Cas9 nuclease is currently the method of choice in most tasks requiring a specific DNA modification. The rapid development in the field of CRISPR-Cas is reflected by the constantly expanding ecosystem of computational tools aimed at facilitating experimental design and result analysis. The first group of CRISPR-Cas-related tools that we review is dedicated to aid in guide RNA design by prediction of their efficiency and specificity. The second, relatively new group of tools exploits the observed biases in repair outcomes to predict the results of CRISPR-Cas edits. The third class of tools is developed to assist in the evaluation of the editing outcomes by analysis of the sequencing data. These utilities are accompanied by relevant repositories and databases. Here we present a comprehensive and updated overview of the currently available CRISPR-Cas-related tools, from the perspective of a user who needs a convenient and reliable means to facilitate genome editing experiments at every step, from the guide RNA design to analysis of editing outcomes. Moreover, we discuss the current limitations and challenges that the field must overcome for further improvement in the CRISPR-Cas endeavor.

Rapid and highly efficient genomic engineering with a novel iEditing device for programming versatile extracellular electron transfer of electroactive bacteria

“…It is empowered by both the self‐cleavage module and the adopted Red or RecET recombination system. The induction of DSBs by the DNA self‐cleavage module stimulates the cellular DNA repair system (Sledzinski et al ., 2020). The adoption of the Red or RecET recombination system is critical to significantly improve the genome‐editing efficiency of the iEditing device by protecting the linear dsDNA and improving its recombineering efficiency.…”

CRISPR Screens in Synthetic Lethality and Combinatorial Therapies for Cancer

“…sgRNAs selection for the CRISPR library must be done by cutting efficiency (which varies among sgRNAs and sequence context) and target specificity (as several mismatches along the sgRNA can be tolerated causing off-target mutations). Recently, many computational models and resources offer further advances in efficiency and specificity prediction for the design and selection of CRISPR pooled libraries [ 32 , 33 , 34 , 35 ]. sgRNA libraries are composed of a set of oligonucleotides, ~20 nt sequences targeting genes or non-coding regions (promoter elements, putative regulatory sequences).…”

Locus-Specific Genomic DNA Purification Using the CRISPR System: Methods and Applications

“…The design of target-specific gRNAs can overcome these issues and various design tools are available. 43,44 Researchers can choose the appropriate gRNAs from the candidates listed from these design tools based on the following guidelines. First, to target promoter regions, gRNAs should be designed to bind several hundred base pairs upstream of the transcription start site to avoid interference with the recruitment of transcription factors and RNA polymerases by the CRISPR complex.…”