Applications

PRCISR™ CRISPR libraries in action

Applications

PRCISR™ CRISPR libraries in action

Explore the full range of applications enabled by PRCISR^TM CRISPR libraries

Monogenetic

Knockout: drug sensitizing and resistance, context-dependency, reporter screens
Inhibition or overexpression: cancer driver mutations, target discovery
Target validation: focused set of genes or families in various contexts, isogenic screens
Others: Epigenetic regulation, chromatin purification, etc.

Fixed-pair

Enhanced gene editing rates: monogenetic screens with gRNA pairs (e.g. dual exon targets, DNA excision)
Validation of combinatorial hits: reduced scale by targeted approach
Gene or protein structure characterization: gene, exon or domain tiling
Target non-coding genome: DNA excision (e.g. lncRNAs, miPs, smORFs)
Variant screening: editing reporter systems, protein engineering, mutational scanning or saturation screens (mutate all possible / relevant positions of a gene)
Barcoded libraries: track single cells or cell populations

Dual-multiplexing

Combinatorial effects between gene families of interest: genetic interactions, synthetic lethality, paralog buffering (all by all, few by all, etc.)
Orthogonal combinatorial applications: disease modeling and vulnerability testing, functional buffering, target discovery, context-independent isogenic screens (e.g. knockout, inactivation, activation in one system)

Synthetic lethality screening

Genetic interactions occur when the combination of two mutations lead to a phenotype that is more severe than either mutation alone. Synthetic lethality is an extreme example of a negative genetic interactions, in which the combination of mutations leads to cellular death.

Synthetic lethality exploits the genetic vulnerabilities of diseased cells by identifying combinations of genetic alterations, where the presence of one mutation sensitizes the diseased cell to the inhibition of another gene or pathway. When these vulnerabilities are targeted, diseased cells can be selectively killed while sparing healthy cells, leading to more effective.

PRCISR™ CRISPR libraries enable synthetic lethality screens that test all possible gene combinations in a single experiment, maximizing hit retention rates while minimizing false negatives, and allows experimental downscaling (up to 10-fold).

Case studies:

Few-by-all: Combinatorial screens uncover genetic interactions in autophagy.

Diehl V, Wegner M, Grumati P, Husnjak K, Schaubeck S, Gubas A, Shah VJ, Polat IH, Langschied F, Prieto-Garcia C, Müller K, Kalousi A, Ebersberger I, Brandts CH, Dikic I, Kaulich M. Minimized combinatorial CRISPR screens identify genetic interactions in autophagy. Nucleic Acids Res. 2021; 49(10): 5684-5704. doi: 10.1093/nar/gkab309.

All-by-all: Combinatorial cells uncover metabolic rewiring in acute myeloid leukemia (AML)
Baker F, Polat IH, Abou-El-Ardat K, Alshamleh I, Thoelken M, Hymon D, Gubas A, Koschade SE, Vischedyk JB, Kaulich M, Schwalbe H, Shaid S, Brandts CH. Metabolic Rewiring Is Essential for AML Cell Survival to Overcome Autophagy Inhibition by Loss of ATG3. Cancers (Basel). 2021; 13(23):6142. doi: 10.3390/cancers13236142.

Parallel screening and experimental downscaling

Vivlion’s uniform PRCISR™ CRISPR libraries enable downscaling of experiments without impeding phenotypic separation or hit calling, thereby facilitating parallel screening in multiple contexts and treatments. In addition, these screens can be used for applications that are beyond the reach of conventional PCR libraries – e.g. target identification in more than 1 M genes etc.

Dual-multiplexing in DNA excision & exon tiling

Vivlion’s PRCISR™ CRISPR dual-multiplex libraries enable DNA excision and domain tiling. When target sequences are in close proximity, the interspanning DNA is excised, enabling the unbiased investigation of coding and non-coding sequence elements. These libraries can be used to explore: