Codon Optimization of saCas9 Gene Enhances Protein Expression in Human Kidney Cells to Compensate for Difficulties in Delivery Vectors to Boost CRISPR‐Cas9 in vivo Therapeutic Application

Abstract

The novel gene editing technology CRISPR‐Cas9 shows great promise in revolutionizing treatments of genetic disorders; however, currently the only practical treatments are ex vivo, limited to blood diseases. In vivo treatment can substantially extend the clinical reach of the technology, but is limited by a challenging delivery process. The large size of the Cas9 gene (~4500bp) inhibits delivery by the small viral vectors and lipid nanoparticles used. We hypothesize that a Cas9 gene codon optimized for human cells will support significantly higher protein expression than the presently used genes; this can compensate for the low delivery efficiency and has great potential to improve the viability of in vivo CRISPR‐Cas9 therapies.To investigate our hypothesis, we selected a commonly used parent saCas9 Addgene’s public plasmid database. We engineered a novel, codon‐optimized gene sequence from the parent using GenScript’s OptimumGene™ algorithm. We will compare protein expression via ELISA assay and Western Blot analysis. In this poster we will present the data gleaned from these assays.