Keep up to date with the most relevant applications and developments for the Edit-R product line.
Are you or will you be doing gene editing using the CRISPR Cas9 system?
Browse our scientific posters on gene editing and learn about using CRISPR Cas9 in order to perform HDR, as well as how the design of your reagents and the promoter you choose to drive Cas9 expression can have a profound effect on gene editing efficiencies in eukaryotic cell lines.
Homology-directed repair with Dharmacon Edit-R CRISPR-Cas9 and single-strand DNA oligos
- In this poster, we demonstrate the use of synthetic single-strand DNA oligo donors in a novel gene editing (Dharmacon Edit-R) platform comprised of synthetic tracrRNA and crRNAs which program Cas9 nuclease to perform HDR, resulting in precise insertion of short DNA sequences. By carefully optimizing lipid-based transfection conditions, we how how we can utilize this platform to create knockins with efficiencies as high as 25%.
Increasing gene editing efficiencies in eukaryotic cell lines by selection of appropriate CRISPR-Cas9 reagents
- Genetic engineering of living cells is critical for understanding gene function in normal and diseased states. The CRISPR-Cas9 system is widely utilized because of its ease-of-use compared to other gene editing methods. This system requires a complex of Cas9 protein with tracrRNA and a gene-targeting crRNA to introduce double-strand DNA breaks at specific locations in the genome to disrupt protein translation and knockout gene function. In this poster, we show how to achieve high gene editing efficiencies, by choosing the best CRISPR-Cas9 reagents for delivery and expression in the cells of interest.
Design considerations for highly specific and efficient synthetic crRNA molecules
- In order to understand the parameters affecting CRISPR-Cas9 gene editing efficiency, we systematically transfected synthetic CRISPR RNA (crRNA) and trans-activating CRISPR RNA (tracrRNA) reagents targeting components of the proteasome into a reporter cell line in which knockout of proteasome function results in fluorescence of a ubiquitin-EGFP fusion protein that is normally degraded by the proteasome pathway. We then evaluated the functionality of > 1100 crRNA sequences in this system and demonstrate the results here.
Picking the best CRISPR-Cas9 targets for functional gene knockout: a machine learning algorithm based on both specificity and functionality
- Not all gene cleavage events result in functional knockout of the target protein. Here we share important advancements that have helped to achieve the goal of picking the very best crRNA targets that result in functional gene knockout, and not just formation of indels (insertions or deletions in the DNA).
Cas9 driven by an optimal promoter improves gene editing in eukaryotic cell lines when paired with synthetic crRNA and tracrRNA
- Presented here are results on the efficiency of using synthetic crRNA and tracrRNA to introduce gene editing events when co-transfected with a plasmid expressing Cas9. We explored the use of antibiotic and fluorescence activated cell sorting (FACS) methods for enrichment of cells that have undergone gene editing, and the use of multiple promoters to increase efficiency of gene editing with Cas9 and synthetic tracrRNA and crRNAs.