EIF5B Knockout HEK293T Polyclonal Cells constitute a CRISPR/Cas9-edited polyclonal population derived from the HEK293T human embryonic kidney cell line, engineered to disrupt the EIF5B gene. This product provides a heterogeneous knockout model in which the target gene is inactivated through CRISPR/Cas9-mediated gene disruption, enabling functional studies of EIF5B-dependent translation initiation. The polyclonal nature allows researchers to assess population-level effects without clonal isolation, capturing a spectrum of genetic alterations that collectively ablate EIF5B function.
The parental HEK293T cell line originated from human embryonic kidney epithelial cells transformed with adenovirus 5 DNA and stably expresses the SV40 large T antigen. This feature permits episomal replication of plasmids containing the SV40 origin of replication, making HEK293T cells a premier host for high-level transient protein expression, lentiviral and retroviral production, and large-scale transfection-based experiments. Their robust growth characteristics and ease of transfection have established HEK293T as a foundational tool in cell and molecular biology.
EIF5B is a eukaryotic translation initiation factor and GTPase that catalyzes a critical step in protein synthesis: the joining of the 60S large ribosomal subunit to the 40S small subunit preinitiation complex, culminating in 80S ribosome formation. This activity is essential for cap-dependent and particularly cap-independent translation initiation, including internal ribosome entry site (IRES)-mediated translation and translation under stress conditions. EIF5B functions downstream of mTOR kinase and eIF2 complexes, integrating signals from nutrient availability, growth factors, and cellular stress. Upon GTP binding, EIF5B interacts with eIF1A, eIF5, and the 40S subunit, facilitating the recruitment and stable assembly of the 60S subunit, thereby licensing the ribosome for translation elongation. Its role is indispensable for viral IRES-dependent translation and cellular adaptation during the integrated stress response.
In the HEK293T background, disruption of EIF5B generates a powerful tool for dissecting the regulatory mechanisms of translation initiation. This polyclonal knockout model is particularly suited for investigating cap-independent translation pathways frequently exploited by RNA viruses, as well as the cellular response to mTOR inhibition or metabolic stress. Given HEK293T’s widespread use in viral vector production, EIF5B knockout cells enable researchers to examine the reliance of viral replication on host translation machinery. Furthermore, because EIF5B activity is often upregulated in cancer cells with high protein synthesis demands, this model supports studies on tumor cell growth dependencies and the validation of EIF5B as a potential therapeutic target.
Researchers can employ these polyclonal knockout cells in a variety of experimental workflows, including polysome profiling and ribosome profiling to assess global translation, puromycin incorporation assays to measure nascent protein synthesis, and dual luciferase IRES reporter assays for quantifying cap-independent translation. Virus replication assays and cell viability assays further extend applications to virology and anticancer drug screening. By combining EIF5B knockout with HEK293T’s transfection efficiency, scientists can dissect signaling networks linking mTOR, eIF2, and downstream translational control. For additional product details or technical assistance, please contact Ascent Research.
