Is your CRISPR workflow labor intensive and prolonged? Inefficient for your gene editing requirements? The new CRISPR-SNIPER technology and services delivered by REPROCELL could be the solution.
Conventional CRISPR-Cas technology, although useful, has limited scope for some gene editing applications including simultaneous multiple gene knock-out, knock-in of longer DNA fragments and single nucleotide modification. CRISPR- SNIPER technology, a timesaving CRISPR-Cas system with built-in optimization capacity and improved gene editing efficiency, overcomes these limitations of conventional systems, broadening application of gene editing technology and leading to improved levels of success.
The CRISPR-Cas experimental workflow consists of a three-stage process: design, delivery, and analysis. Challenges occur at each stage and often require intense and time-restrictive optimisation. Cas9 nuclease activity, target site selection, single-guide RNA design, delivery methods, off-target effects, and the incidence of homology-directed repair are all factors, that can affect the efﬁciency and speciﬁcity of the system. Developments in informatics and machine-learning help abrogate some factors and molecular modification of components can increase activity. However, even with these improvements, the complete workflow can take several months to complete, and the gene editing efficiency plus the number of positively edited clones may still be inadequate. Experimental failure is therefore only recognized after many weeks or months
of effort have been expended.
Advancing CRISPR-Cas technology with SNIPER screen
CRISPR-SNIPER technology addresses key areas of the three-stage process—the analytical stage and the optimization steps required for the design and delivery stages.
The analytical stage of CRISPR-Cas technology is generally the most onerous, labor-intensive, and costly stage, and it often ends in failure. Following delivery and transfection of the system components, a limiting dilution of the transfected cells is performed and individual clones are picked for analysis. Depending on the efficiency of the system, this process is repeated several times. On average, hundreds of clones selected and checked.
SNIPER (Specification of Newly Integrated Position and Exclusion of Random-integration) combines a checkerboard of culture conditions with highly sensitive digital-PCR to empirically prescreen and identify clones most likely to be positive for the desired gene edit prior to the limiting dilution step. As a result, fewer clones are selected for checking, more positive clones are identified, and the screening process is significantly reduced.
Additionally, and unique to CRISPR-SNIPER, component design, multiple component delivery, and transfection strategies can be assessed under different culture conditions within the checkerboard to optimize and select the best combination for a particular cell type, negating the need for multiple time-consuming optimization steps in the early stages of the process.
The addition of SNIPER to the CRISPER-Cas system thereby facilitates improved and quick optimization together with increased gene editing efficiency. Crucially, success or failure is identified at an early stage in the workflow, ensuring that time and costs are kept to a minimum for unsuccessful gene editing attempts.
REPROCELL CRISPR-SNIPER gene-editing services
Unlike comparable gene editing companies, REPROCELL offers a modular CRISPR-SNIPER approach that suits many different and complex gene editing needs.
The “Design and Feasibility” module enables quick optimization of conditions for a specific gene edit and supplies a bulk clone stock ready for clone selection. REPROCELL will only request the sequence and locus of your target gene—no additional methodology is required. REPROCELL will custom design the single-guide RNA and donor vectors for your experiment and will test up to 10 transfection conditions to identify the best design and delivery option.
The “Full Service” module offers a more comprehensive approach, incorporating not only the design and feasibility component but also subsequent clone selection, edit characterization, and clone expansion components to supply individual clones with the required edit.