RAS Pathway Knock-In Cell Lines
We offer a large selection of prebuilt CRISPR knock-in cell lines within the RAS-RAF pathway utilizing the HiBiT tag. Cell backgrounds including LgBiT allow live-cell kinetic analysis of the HiBiT-tagged protein.
| KI Cell Line | Background |
|---|---|
| HiBiT-KRAS(G12C) | MIA PaCa-2 |
| HiBiT-KRAS(G12D) | AsPc-1 |
| HiBiT-KRAS(G12D) | SW1990 |
| HiBiT-KRAS(G12D) | HEK293 |
| HiBiT-KRAS(G12S) | A549 |
| HiBiT-KRAS(G12V) | HEK293 |
| HiBiT-KRAS(G12V) | SW620 |
| HiBiT-KRAS(G13D) | NCI-H647 |
| HiBiT-KRAS(WT) | HEK293 |
| HiBiT-KRAS(WT) | NCl-H1299 |
| HiBiT-NRAS(Q61R) | SW1271 |
| HiBiT-NRAS(WT) | HEK293 |
| HiBiT-ARAF | A549 |
| HiBiT-BRAF | A549 |
| HiBiT-BRAF(V600E) | A375 |
| RAF1-HiBiT | A549 |
| RAF1-HiBiT | Hs766T |
| RAF1-HiBiT | PANC 08.13 |
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HiBiT Knock-In Cell Lines for RAS Pathway Targets
CRISPR-engineered HiBiT Knock-in Cell Lines offer a powerful tool to study the abundance and degradation of endogenous proteins involved in critical signaling pathways, such as the RAS-RAF-MEK-ERK cascade. This pathway begins with the activation of RAS, a family of small GTPases, which includes KRAS, HRAS and NRAS. Active, GTP-bound RAS activates the RAF family of serine/threonine kinases (c-Raf, B-Raf, A-Raf). This activation sequence leads to the phosphorylation of MEK and ultimately ERK, which are essential for controlling various cellular processes.
An overview of the RAS-RAF-MEK-ERK pathway.
HiBiT is a small, 11-amino-acid epitope tag that enables precise measurement of protein expression through bioluminescent signal when bound to its complementation partner, LgBiT. The bound complex exhibits high luciferase activity, producing a bright luminescent signal in the presence of added substrate. The small size of the HiBiT tag makes it ideal for endogenous tagging using CRISPR/Cas9 editing.
When inserted into key proteins within this pathway using CRISPR technology, the HiBiT tag enables precise monitoring of protein dynamics in real-time. Upon binding with LgBiT, the HiBiT tag generates a luminescent signal, which decreases as the tagged protein undergoes degradation. This method allows researchers to quantitatively assess protein degradation rates, Dmax and DC50 values, making it a powerful tool for studying protein stability and for high-throughput screening of compounds that influence the RAS-RAF-MEK-ERK signaling pathway.
A.
B.
Live-cell-degradation kinetics of endogenous HiBiT-KRasG12C following PROTAC treatment. Panel A. HiBiT was inserted via CRISPR/Cas9 at the N-terminal endogenous KRasG12C loci in the MIA-PaCa2 cell line. Following LgBiT expression, dose-response kinetic degradation experiments were performed using the VHL-based KRasG12C PROTAC, LC-2, in CO2-independent medium containing Nano-Glo® Endurazine™ Substrate. Fractional RLU is plotted relative to the DMSO control. Panel B. Similar live-cell luminescent studies of HiBiT-KRasG12C in MIA-Paca2 cells using the parent inhibitor, MRTX849, which does not show loss of KRasG12C over the same dose-response treatments.
Interested in our other CRISPR knock-in cell lines? View our full library of ready-to-use cells.
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