The IDH3G Knockout HEK293T Polyclonal Cells represent a CRISPR/Cas9-edited polyclonal population carrying a targeted disruption of the IDH3G gene. This loss-of-function model is generated in HEK293T cells through CRISPR/Cas9-mediated gene disruption, resulting in a heterogeneous knockout cell pool suitable for studying IDH3G-dependent metabolic and regulatory networks. The polyclonal format provides a robust experimental system that captures diverse editing events without the selective pressure of clone isolation.
HEK293T cells are human embryonic kidney cells constitutively expressing the SV40 large T antigen, which facilitates high-level protein expression, viral vector production, and efficient transfection. Derived from the adherent epithelial HEK293 cell line, HEK293T cells serve as a versatile host for exogenous DNA replication and recombinant protein synthesis. Their well-characterized genetic background and metabolic activity make them an ideal platform for interrogating mitochondrial enzyme function, particularly in cancer biology and metabolic reprogramming studies.
IDH3G encodes the gamma subunit of mitochondrial NAD+-dependent isocitrate dehydrogenase (IDH3), which catalyzes the oxidative decarboxylation of isocitrate to ??-ketoglutarate, generating NADH. This key TCA cycle enzyme is regulated by PGC-1??, NRF1, and HIF-1??, and its activity supplies ??-ketoglutarate to downstream 2-oxoglutarate-dependent dioxygenases, including TET enzymes, prolyl hydroxylases, and JmjC histone demethylases. IDH3G associates with IDH3A and IDH3B to form the functional heterotetramer and cooperates with mitochondrial malate dehydrogenase and NADH salvage pathways to maintain redox balance. Disruption of IDH3G compromises the IDH3 complex, diminishing ??-ketoglutarate pools and impairing TCA cycle flux, which may trigger compensatory glutamine-dependent reductive carboxylation. This metabolic dysregulation can alter the epigenetic landscape by reducing substrate availability for ??-ketoglutarate-dependent dioxygenases.
In the HEK293T background, IDH3G knockout introduces a defined metabolic lesion that permits detailed functional analyses of mitochondrial bioenergetics and NAD+/NADH dynamics. The loss of IDH3G activity is expected to reduce electron transport chain efficiency and lower TCA cycle throughput, potentially shifting cellular metabolism towards reductive carboxylation to sustain biosynthesis under normoxic or hypoxic conditions. Because HEK293T cells retain robust proliferation and biosynthetic capacity, this model is particularly suited for dissecting the interplay between mitochondrial metabolism and epigenetic regulation, as well as for studying adaptive metabolic responses in a simplified cellular environment.
This polyclonal knockout cell population is a powerful tool for a spectrum of research applications, including TCA cycle dysfunction modeling, cancer cell metabolism investigations, mitochondrial disease mechanism studies, ??-ketoglutarate-dependent epigenetic regulation, and metabolic drug screening campaigns. Typical assays compatible with these cells encompass western blotting and RT-qPCR for confirming gene disruption, Seahorse metabolic flux analysis to assess oxygen consumption and glycolytic rates, LC-MS-based metabolite profiling of key intermediates (??-ketoglutarate, citrate, NAD+/NADH), IDH enzyme activity measurements, and cell proliferation or apoptosis assays. For further technical details or pricing inquiries, please contact Ascent Research.
