The EFNB3 Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal cell population derived from the near-haploid HAP1 line, featuring targeted disruption of the EFNB3 gene. This product provides a mixed pool of edited cells, enabling immediate loss-of-function studies without clonal isolation.
HAP1 is a human chronic myeloid leukemia-derived adherent cell line with a near-haploid karyotype, offering a simplified genetic background that facilitates unambiguous gene disruption. Its robust growth and suitability for high-throughput screening make it a preferred platform for genetic interaction screens and functional genomics.
EFNB3 encodes ephrin-B3, a transmembrane ligand for EphB receptors (EphB1/B2/B3). Ligand-receptor engagement triggers bidirectional signaling: forward through EphB and reverse through ephrin-B3. Reverse signaling recruits Src kinases (e.g., Fyn) and phosphorylates FAK, modulating Rho GTPases (Rac1, RhoA) to remodel the actin cytoskeleton and regulate adhesion and migration. Ephrin-B3 also binds PDZ-domain proteins (GRIP1, syntenin) that organize signaling complexes. This pathway governs axon guidance, cell migration, and tissue boundary formation, with links to congenital cranial dysinnervation disorders, cancer, and Hirschsprung disease.
In HAP1, disruption of the single EFNB3 allele abolishes ephrin-B3 protein production, creating a clean loss-of-function model. The polyclonal population captures diverse editing events yet collectively yields potent phenotypic effects, making it ideal for pooled assays such as drug sensitivity screens or migration studies, where the haploid background amplifies the impact of gene loss.
Researchers can employ these cells in migration and invasion assays to dissect ephrin-B3??s role in cell motility, as well as adhesion assays to examine integrin-mediated attachment. Immunofluorescence for F-actin visualizes cytoskeletal changes, flow cytometry confirms loss of surface ephrin-B3, and co-immunoprecipitation of EphB receptors assesses ligand-receptor interactions. RT-qPCR and Western blotting monitor downstream effectors such as FAK and Rho GTPases. These applications support investigations into axon guidance, cancer metastasis, and drug target validation. For additional information, please contact Ascent Research.
