CD320 Knockout HT29 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population derived from the HT29 human colorectal adenocarcinoma cell line. This product introduces loss-of-function mutations in the CD320 gene via CRISPR/Cas9-mediated gene disruption, generating a heterogeneous cell pool with targeted ablation of the CD320-encoded transcobalamin II receptor. As a polyclonal knockout, it avoids the limitations of single-cell clones and retains population-level diversity, making it suitable for applications requiring robust representation of knockout phenotypes without clonal artifacts. The cells are provided as a ready-to-use polyclonal population optimized for downstream cellular and molecular assays in biomedical research.
The HT29 host cell line is an established epithelial cell model derived from primary colorectal adenocarcinoma of a 44-year-old Caucasian female. These cells exhibit adherent growth and intestinal epithelial characteristics, widely employed in colorectal cancer biology, drug absorption studies, and epithelial barrier function assays. HT29 cells retain key metabolic pathways relevant to gastrointestinal physiology and cancer metabolism, including active one-carbon metabolism and folate cycling, providing a physiologically relevant background for studying cobalamin-dependent processes in the context of colorectal adenocarcinoma.
CD320 encodes the high-affinity receptor for transcobalamin II (TCN2), responsible for cellular uptake of cobalamin (vitamin B12). Upon binding TCN2-cobalamin complexes, CD320 mediates endocytosis and lysosomal release of B12, which serves as a cofactor for two critical enzymes: methionine synthase (MTR) in the methionine cycle and methylmalonyl-CoA mutase (MUT) in succinyl-CoA synthesis. Disruption of CD320 impairs intracellular B12 supply, leading to decreased MTR activity and homocysteine remethylation, as well as reduced MUT function, resulting in accumulation of homocysteine and methylmalonic acid. This knockout therefore models the metabolic consequences of vitamin B12 deficiency, linking cobalamin transport to one-carbon metabolism, S-adenosylmethionine synthesis, and folate cycle regulation.
In the context of HT29 colorectal adenocarcinoma cells, CD320 knockout provides a powerful tool to investigate the intersection of vitamin B12 metabolism and cancer cell physiology. Colorectal cancer cells often exhibit heightened dependence on one-carbon metabolism for nucleotide synthesis and methylation reactions, and B12 deficiency can compromise these pathways, potentially sensitizing cells to metabolic stress or altering drug responses. This model enables dissection of how impaired cobalamin uptake affects methionine cycle flux, genomic methylation patterns, and redox balance in epithelial cancer cells, offering insights into the metabolic vulnerabilities associated with B12 depletion that are relevant to pathological conditions like methylmalonic aciduria and homocystinuria, as well as nutritional deficiencies in cancer patients.
Researchers can employ this knockout model in a wide array of experimental workflows, including cobalamin uptake assays using labeled B12, intracellular B12 quantification by ELISA, homocysteine and methylmalonic acid profiling via LC-MS, methionine synthase activity measurements, and proliferation or colony formation assays under defined B12 conditions. Additional applications include RNA-seq transcriptome analysis to identify B12-responsive genes, apoptosis assessment by flow cytometry, and drug sensitivity screens to evaluate interactions between chemotherapeutic agents and B12 metabolic status. The polyclonal nature supports bulk population studies, RNAi validation, and complementation experiments with wild-type CD320. For technical inquiries or to explore custom assay development, please contact Ascent Research.
