The AFF2 Knockout HEK293T Polyclonal Cells are a CRISPR/Cas9-mediated gene disruption model designed to eliminate AFF2 expression in the HEK293T cell line. This polyclonal knockout cell population provides a heterogeneous loss-of-function system for studying AFF2-dependent transcriptional regulation, without clonal selection, thereby preserving a broader representation of genetic backgrounds common in pooled screening applications.
HEK293T is a human embryonic kidney epithelial cell line derived from HEK293 cells, which were originally transformed with sheared adenovirus 5 DNA. It stably expresses the SV40 large T antigen, enabling episomal replication of plasmids containing the SV40 origin of replication. This feature, combined with high transfectability and robust protein expression capacity, has made HEK293T a workhorse for viral production, recombinant protein expression, and cell-based functional assays.
AFF2 encodes a transcriptional activator belonging to the AF4/FMR2 family and serves as a core scaffold protein of the super elongation complex (SEC). Within the SEC, AFF2 directly interacts with elongation factors ELL and ELL2, histone methyltransferase MLL (KMT2A), and the P-TEFb components CDK9 and Cyclin T1. This complex is recruited to chromatin via BRD4 binding to acetylated histones, where CDK9 phosphorylates RNA polymerase II to release promoter-proximal pausing and drive productive elongation. AFF2 activity is modulated by the Wnt/??-catenin pathway, as ??-catenin/TCF complexes can direct SEC to target promoters. Key downstream effectors include immediate early genes such as MYC, FOS, and JUN, as well as neurodevelopmental genes like BDNF and GRIN2B.
In the HEK293T cellular context, disruption of AFF2 impairs SEC integrity and attenuates transcriptional elongation of its target genes, leading to altered cell proliferation and gene expression profiles. While HEK293T is not a neuronal lineage, its robust signaling pathways and high manipulability allow researchers to dissect the fundamental mechanisms of SEC function and cross-talk with Wnt/??-catenin signaling. The polyclonal nature permits assessment of heterogeneous gene-editing outcomes, making it suitable for pooled functional genomics screens and drug sensitivity assays.
This product is applicable to a wide array of research areas, including the investigation of transcriptional elongation mechanisms, the study of AFF2 dysfunction in neurodevelopmental disorders such as Fragile XE mental retardation (FRAXE) and autism spectrum disorder, and high-throughput screening for SEC modulators. Typical experimental workflows involve western blotting to confirm AFF2 depletion, RT-qPCR and RNA-seq to quantify changes in SEC target expression, co-immunoprecipitation to map altered protein interactions, and ChIP-qPCR to assess SEC occupancy at regulatory regions. For further technical specifications, validation data, and ordering information, please contact Ascent Research.
