A benchmark of computational CRISPR-Cas9 guide design methods

PLoS Comput Biol. 2019 Aug 29;15(8):e1007274. doi: 10.1371/journal.pcbi.1007274. eCollection 2019 Aug.

Abstract

The popularity of CRISPR-based gene editing has resulted in an abundance of tools to design CRISPR-Cas9 guides. This is also driven by the fact that designing highly specific and efficient guides is a crucial, but not trivial, task in using CRISPR for gene editing. Here, we thoroughly analyse the performance of 18 design tools. They are evaluated based on runtime performance, compute requirements, and guides generated. To achieve this, we implemented a method for auditing system resources while a given tool executes, and tested each tool on datasets of increasing size, derived from the mouse genome. We found that only five tools had a computational performance that would allow them to analyse an entire genome in a reasonable time, and without exhausting computing resources. There was wide variation in the guides identified, with some tools reporting every possible guide while others filtered for predicted efficiency. Some tools also failed to exclude guides that would target multiple positions in the genome. We also considered two collections with over a thousand guides each, for which experimental data is available. There is a lot of variation in performance between the datasets, but the relative order of the tools is partially conserved. Importantly, the most striking result is a lack of consensus between the tools. Our results show that CRISPR-Cas9 guide design tools need further work in order to achieve rapid whole-genome analysis and that improvements in guide design will likely require combining multiple approaches.

Publication types

  • Evaluation Study

MeSH terms

  • Animals
  • Benchmarking / methods
  • Benchmarking / statistics & numerical data
  • CRISPR-Cas Systems*
  • Computational Biology
  • Databases, Nucleic Acid / statistics & numerical data
  • Gene Editing / methods*
  • Gene Editing / standards
  • Gene Editing / statistics & numerical data
  • Mice
  • RNA, Guide, CRISPR-Cas Systems / genetics*
  • Software

Substances

  • RNA, Guide, CRISPR-Cas Systems

Grants and funding

The authors received no specific funding for this work.