The ACADVL Knockout HEK293T Polyclonal Cells comprise a CRISPR/Cas9-edited polyclonal population derived from HEK293T cells, featuring targeted disruption of the ACADVL gene. This gene encodes very long-chain acyl-CoA dehydrogenase (VLCAD), a mitochondrial enzyme critical for the initial step of long-chain fatty acid ??-oxidation. The resulting polyclonal knockout cell pool provides a reliable loss-of-function model for investigating fatty acid metabolism and associated disorders.
The HEK293T host cell line is a widely utilized human embryonic kidney epithelial derivative that stably expresses the SV40 large T antigen, enabling episomal replication of plasmids containing the SV40 origin of replication. This feature, combined with its ease of transfection and rapid growth, makes HEK293T a preferred cell line for transient protein expression and lentiviral production. In the context of metabolic gene knockout studies, HEK293T provides a tractable and scalable platform, allowing researchers to dissect mitochondrial functions without the complexities of primary cell culture.
The ACADVL protein (VLCAD) catalyzes the dehydrogenation of very long-chain acyl-CoA esters (C14?CC20) within the mitochondrial matrix, representing the first and rate-limiting step of long-chain fatty acid ??-oxidation. Its activity is transcriptionally regulated by PPAR?? and PGC-1??, and it is functionally coupled to electron transfer flavoprotein (ETF) and ETF-ubiquinone oxidoreductase to feed electrons into the respiratory chain. Downstream products include acetyl-CoA, NADH, and FADH2, which support ATP synthesis and ketone body production, particularly during fasting or high-fat diet challenges.
Although HEK293T cells are not a classical metabolic cell type, they retain functional mitochondrial ??-oxidation machinery and express key regulators of fatty acid metabolism, making them extensively employed for pathway dissection. Disruption of ACADVL in this background creates a versatile model to study VLCAD deficiency-related metabolic defects, mitochondrial dysfunction, and compensatory metabolic adaptations, such as shifts in substrate utilization or upregulation of alternative oxidases. This model is particularly valuable for high-throughput screening and mechanistic studies where genetic tractability is paramount.
This polyclonal knockout pool is suitable for a wide range of research applications, including metabolic profiling of long-chain acylcarnitines via LC-MS, assessment of mitochondrial respiration using Seahorse analyzers with palmitate as substrate, and evaluation of PPAR?? target gene expression by RT-qPCR. It also supports immunofluorescence studies of mitochondrial morphology and investigations into cardiac metabolism and drug screening for VLCAD deficiency. This model serves as a reliable tool for studying very long-chain acyl-CoA dehydrogenase deficiency, enabling mechanistic insights into cardiomyopathy, skeletal myopathy, and hypoketotic hypoglycemia. For additional details or technical support, please contact Ascent Research.
