KC-3522

293T-UAS-Luc2-GAL4BD-ELK1 Cell Line

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Background of 293T-UAS-Luc2-GAL4BD-ELK1 Cell Line

ELK1 is transcription factor that binds to purine-rich DNA sequences. It forms a ternary complex with SRF and the ETS and SRF motifs of the serum response element (SRE) on the promoter region of immediate early genes such as FOS and IER2. ELK1 induces target gene transcription upon JNK-signaling pathway stimulation.The biological function of Elk-1 relies essentially on the interaction with other proteins. Elk-1 binds to SRF and generates a functional ternary complex that is required to activate SRE-mediated gene transcription. Elk-1 is kept in an inactive state under basal conditions via binding of a SUMO-histone deacetylase complex.

Specifications

Catalog Number	KC-3522
Cell Line Name	293T-UAS-Luc2-GAL4BD-ELK1 Cell Line
Host Cell Line	293T-UAS-Luc2
Description	Stable 293T cell line expressing exogenous luciferase gene under the control of ELK1 signaling pathway.
Quantity	Two vials of frozen cells (≥2-10⁶/vial)
Stability	Stable in culture over a minimum of 10 passages
Application	Drug screening and biological assays
Freezing Medium	70% DMEM+20% FBS+10% DMSO
Propagation Medium	DMEM+10% FBS +75μg/ml Hygromycin B+0.5μg/ml puromycin
Selection Marker	Puromycin, Hygromycin B
Morphology	Epithelial
Subculture	Split saturated culture 1:4-1:8 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL
Incubation	37 °C with 5% CO₂
Storage	Liquid nitrogen immediately upon receiving
Doubling Time	Approximately 30 hours
Mycoplasma Status	Negative

Cell Line Generation

Split saturated culture 1:4-1:8 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL

Characterization

Figure 1. 293T-UAS-Luc2-GAL4BD-ELK1 cells were seed into the 96-well plate, and treated with IGF1 for 16 h, then readout with Bright-Glo system.

Cell Resuscitation

Prewarm culture medium (DMEM supplemented with 10% FBS, 75μg/ml Hygromycin and 0.5μg/ml Puromycin)in a 37°C water bath.
Thaw the frozen vial in a 37°C water bath for 1-2 minutes.
Transfer the vial into biosafety cabinet, and wipe the surface with 70% ethanol.
Unscrew the top of the vial and transfer the cell suspension gently into a sterile centrifuge tube containing 9.0mL complete culture medium.
Spin at ~ 125 × g for 5-7 minutes at room temperature, and discard the supernatant without disturbing the pellet.
Resuspend cell pellet with the appropriate volume of complete medium and transfer the cell suspension into a T25 culture flask.
Incubate the flask at 37°C, 5% CO₂ incubator.
Split saturated culture 1:4-1:8 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

Prepare the freezing medium (70% DMEM + 20% FBS + 10% DMSO) fresh immediately before use.
Keep the freezing medium on ice and label cryovials.
Transfer cells to a sterile, conical centrifuge tube, and count the cells.
Centrifuge the cells at 250×g for 5 minutes at room temperature and carefully aspirate off the medium.
Resuspend the cells at a density of at least 3×10⁶ cells/mL in chilled freezing medium.
Aliquot 1 mL of the cell suspension into each cryovial.
Freeze cells in the CoolCell freezing container overnight in a -80°C freezer.
Transfer vials to liquid nitrogen for long-term storage

References

1.Thiel G, Backes TM, Guethlein LA, Rössler OG. Critical Protein-Protein Interactions Determine the Biological Activity of Elk-1, a Master Regulator of Stimulus-Induced Gene Transcription. Molecules. 2021 Oct 11;26(20):6125. doi: 10.3390/molecules26206125. PMID: 34684708; PMCID: PMC8541449.
2.Janknecht R, Nordheim A. Elk-1 protein domains required for direct and SRF-assisted DNA-binding. Nucleic Acids Res. 1992 Jul 11;20(13):3317-24. doi: 10.1093/nar/20.13.3317. PMID: 1630903; PMCID: PMC312483.