A Novel DNA Purification Technology Allows for Efficient and Faster Assembly of Expression Clones for CRISPR/Cas9-Mediated Genome Editing

The CRISPR/Cas9 technology allows for rapid and efficient genomic modifications in a variety of cellular contexts. However, the construction of expression clones can be monotonous, requiring multiple rounds of cloning and/or PCR. Due to the necessity of replicates and repeat experiments to obtain even a single desired construct, multiple rounds and hours of nucleic acid purifications are performed from post-enzymatic reactions and bacteria. Here, we describe the assembly of gRNA expression cassettes into vectors via Golden Gate and Gibson cloning technologies using two distinct DNA purification technologies in parallel. We demonstrate that the utility of a novel PCR and plasmid purification method provides significant time-savings, increased performance, and ease of use compared to traditional spin-columns. Furthermore, the efficiency of this novel system was verified by Next Generation Sequencing. The system reported here provides a much more efficient and faster process to construct expression clones for CRISPR/Cas9-mediated genome editing.