Depletion of Microbiome-Derived Molecules in the Host Using Clostridium Genetics

The rapid expansion of research using CRISPR-Cas requires a flexible software solution to guide precise genome editing beyond model organisms and simple applications. Here we present CASPER (CRISPR Associated Software for Pathway Engineering and Research), an integrated software platform that provides end-users the ability to design CRISPR tools to manipulate non-traditional biological systems beyond model organisms with a wide-variety of newly discovered and engineered Cas endonucleases. CASPER contains algorithms for on- and off-target analysis and multitargeting that enable target design of user-provided genomes and populations/metagenomes. Using CASPER, we were able to rapidly construct a library of 125 targets for a S. aureus infection in a skin microbiome, limiting predicted off-targets across the microbial community and cross-compatibility with both the endogenous CRISPR-Cas system of S. aureus and the ubiquitous S. pyogenes Cas machinery. Tight integration with NCBI and KEGG databases expands the bioinformatic capabilities of CASPER, including the ability to rapidly generate guide-RNA libraries from database provided annotations.