The BRAT1 Knockout HEK293T Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population with targeted disruption of the BRAT1 gene in the HEK293T host line. This loss-of-function model, generated via CRISPR/Cas9-mediated gene disruption without single-cell cloning, provides a heterogeneous pool of edited cells ideal for pooled functional studies and high-throughput screens. The polyclonal format captures population-level responses, making it suitable for experiments requiring robust genetic perturbation across diverse cellular backgrounds. These cells serve as a versatile tool for dissecting BRAT1-dependent processes in a widely used human cellular system.
The HEK293T cell line, derived from human embryonic kidney epithelium, stably expresses the SV40 large T antigen, allowing episomal replication of plasmids containing the SV40 origin. This property facilitates high-efficiency transfection, robust protein expression, and viral production. HEK293T??s genetic tractability and well-characterized signaling pathways make it a preferred host for exploring DNA damage responses and cell cycle regulation. Combining HEK293T??s experimental advantages with BRAT1 disruption yields a powerful model for mechanistic and pharmacological investigations.
BRAT1 acts as an essential activator of ATM kinase in the DNA double-strand break response. It bridges the MRN complex (MRE11-RAD50-NBS1) to ATM, promoting ATM autophosphorylation and subsequent phosphorylation of downstream effectors CHK2, p53, and H2AX (??H2AX), which enforce cell cycle arrest, DNA repair, or apoptosis. BRAT1 also interacts with repair factors including 53BP1, SMC1, and DNA-PKcs, integrating signals for genome maintenance. Loss of BRAT1 cripples ATM activation, compromising checkpoint fidelity and increasing genomic instability.
In HEK293T cells, BRAT1 knockout ablates efficient ATM signaling, enabling studies of pathologies like lethal neonatal rigidity and multifocal seizure syndrome (RMFSL) and neurodevelopmental disorders. The model also illuminates cancer-relevant processes, such as impaired apoptosis and uncontrolled proliferation under genotoxic stress. By screening for genetic or small-molecule modifiers that restore ATM activity independently of BRAT1, researchers can identify novel therapeutic targets for DNA repair-deficient cancers and related syndromes.
These polyclonal knockout cells support diverse assays: Western blotting for phosphorylation of ATM, CHK2, and p53; immunofluorescence of ??H2AX foci to monitor DNA repair; flow-cytometric cell cycle and annexin V apoptosis analyses; and clonogenic survival or drug sensitivity tests. Co-immunoprecipitation maps interaction networks, while RNA-seq reveals transcriptional consequences of BRAT1 loss. For additional information or to order, contact Ascent Research.
