The BRAF Knockout HAP1 Polyclonal Cells product provides a CRISPR/Cas9-edited polyclonal cell population carrying a targeted disruption of the BRAF gene in the HAP1 human near-haploid cell line. This knockout model serves as a loss-of-function tool for investigating BRAF-dependent signaling and tumor biology. The polyclonal format captures a heterogeneous pool of gene-edited cells, enabling robust population-level analyses without clonal selection biases. The product is designed for advanced research applications requiring dissection of the MAPK pathway and its roles in cancer.
HAP1 is a human near-haploid chronic myeloid leukemia cell line originally derived from KBM-7. Its near-haploid karyotype simplifies genetic manipulation and functional genomics studies, as single-copy gene disruption directly yields phenotypic consequences without confounding effects from second allele compensation. The HAP1 line is widely adopted for CRISPR screening, pathway dissection, and high-throughput drug profiling. This genetic background provides a controlled system for evaluating gene function in a disease-relevant context, particularly in hematopoietic malignancies and signal transduction research.
BRAF encodes a serine/threonine-protein kinase that functions as a critical effector downstream of RAS proteins (KRAS, HRAS, NRAS) in the Ras-Raf-MEK-ERK (MAPK) signaling cascade. Upon activation by upstream receptor tyrosine kinases and growth factors such as EGF and FGF, RAS recruits and activates BRAF, which in turn phosphorylates and activates MEK1 (MAP2K1) and MEK2 (MAP2K2). These kinases subsequently phosphorylate ERK1 (MAPK3) and ERK2 (MAPK1), promoting the transcription of immediate-early genes including FOS, JUN, and MYC through phosphorylation of transcription factors like ELK1. BRAF activity is modulated by interacting partners such as 14-3-3 proteins, the scaffold protein KSR1, SRC kinase, and the HSP90 chaperone, which regulate its localization, stability, and catalytic output. The BRAF-MEK-ERK axis governs fundamental cellular processes including proliferation, differentiation, and survival, and its dysregulation is a hallmark of numerous malignancies.
In the HAP1 background, disruption of BRAF creates a defined genetic model for interrogating MAPK pathway dependence and drug sensitivity. Given the near-haploid genome, each cell carries a single functional copy of the BRAF gene, allowing a complete loss-of-function at the protein level in the knockout population without the need for homozygous editing. This model is particularly relevant for studying BRAF-driven cancers such as melanoma, colorectal cancer, and non-small cell lung cancer, where oncogenic BRAF mutations drive constitutive pathway activation. Moreover, the knockout line enables investigation of upstream regulatory inputs and downstream effectors in a clean genetic context, facilitating the dissection of resistance mechanisms to BRAF inhibitors like vemurafenib and dabrafenib.
The BRAF knockout HAP1 polyclonal cells are suited for a broad range of biomedical research applications. Functional validation can be performed using western blotting to assess loss of phospho-ERK and total ERK levels, confirming pathway inactivation. Cell proliferation assays (MTS/MTT), colony formation assays, and flow cytometry-based cell cycle and apoptosis analyses allow quantitative assessment of growth and survival phenotypes. Drug sensitivity testing with targeted therapies, particularly BRAF and MEK inhibitors, can be conducted to map pathway dependencies and explore combination strategies. Additionally, the cells are amenable to high-throughput screening for synthetic lethal interactions and for testing novel compounds targeting the MAPK pathway. MAPK pathway reporter assays and co-culture experiments further expand the utility for studying cell signaling dynamics. For more information or technical support, please contact Ascent Research.
