The DNAAF2 Knockout HT29 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal cell population that carries targeted disruption of the DNAAF2 gene in the HT29 human colorectal adenocarcinoma cell line. This loss-of-function model provides a population-level knockout suitable for functional studies without requiring clonal isolation, making it ideal for bulk biochemical analyses and screening assays.
HT29 is a colorectal adenocarcinoma-derived epithelial cell line that harbors well-characterized oncogenic mutations, including inactivating mutations in APC and TP53. These genetic alterations confer tumorigenic properties and enable the line to serve as a versatile model of colorectal epithelium, capable of undergoing mucin-producing differentiation under appropriate conditions.
DNAAF2 encodes a cytoplasmic dynein axonemal assembly factor essential for the preassembly of outer and inner dynein arms within cilia and flagella. Its expression is transcriptionally regulated by FOXJ1 and RFX transcription factors such as RFX3. DNAAF2 functions in a multi-protein complex with DNAAF1, LRRC6, and ZMYND10 to facilitate the incorporation of dynein motors including the heavy chain DNAH5 and intermediate chain DNAI1 into the axoneme. Disruption of DNAAF2 therefore leads to defective dynein arm assembly, resulting in immotile cilia and impaired mucociliary clearance, a hallmark of primary ciliary dyskinesia.
In the context of HT29 cells, DNAAF2 knockout provides a unique platform to examine the interplay between ciliary dysfunction and oncogenic signaling. Although HT29 cells are not typically highly ciliated, they can express primary cilia and ciliary markers such as IFT88 and ARL13B. Loss of DNAAF2 may perturb ciliary-mediated signaling pathways, affecting differentiation, barrier function, and invasive properties, thereby offering insights into how ciliary defects influence colorectal cancer biology.
Typical research applications for these polyclonal knockout cells include western blotting and RT-qPCR for DNAAF2 and ciliary markers (FOXJ1, DNAH5, ARL13B), immunofluorescence staining for acetylated tubulin and ARL13B to visualize cilia, ciliary beat frequency analysis, RNA-sequencing-based transcriptomic profiling, mucin secretion assays, and cell migration/invasion studies. The model is also well-suited for drug screening targeting ciliopathies or ciliary signaling in cancer. For further technical inquiries, please contact Ascent Research.
