The HEK293, Malme-3M, Malme-3 and A375 cell lines were obtained from American Type Culture Collection, and SK-Mel-19 was kindly provided by Dr. Neal Rosen (Memorial Sloan-Kettering Cancer Center). The cancer cell lines derived from melanoma (SK-Mel-19, A375 and Malme-3M) were maintained in RPMI-1640 medium (Gibco) supplemented with 10% heat-inactivated fetal bovine serum (Biological Industries), 50 U/ml penicillin, and 50 μg/ml streptomycin at 37°C with 5% CO2. The Malme-3 cells, which were derived from skin fibroblasts, were grown in McCoy's 5a Medium (Gibco) supplemented with 20% heat-inactivated FBS, 50 U/ml penicillin, and 50 μg/ml streptomycin at 37°C with 5% CO2. The human embryonic kidney cell line HEK293 was maintained in Dulbecco's modified Eagle's medium (Gibco) supplemented with 5% heat-inactivated fetal bovine serum, 50 U/ml penicillin, and 50 μg/ml streptomycin at 37°C with 5% CO2. The MEK inhibitors, PD325901, PLX4032 (Selleck) and U0126 (Sigma-Aldrich), and actinomycin D (Enzo) were dissolved in DMSO as stock solutions and stored at −20°C.
Pd325901
Manufactured by Selleck Chemicals
PD325901 is a chemical compound used in research laboratory settings. It functions as a potent and selective inhibitor of the MEK1/2 protein kinases. PD325901 is commonly utilized in cell-based assays and in vitro experiments to study the role of the ERK signaling pathway in various biological processes.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions)
Rpmi 1640 medium, by Thermo Fisher Scientific (2 mentions)
Plx4032, by Selleck Chemicals (1 mentions)
Fetal bovine serum (fbs), by Sartorius (1 mentions)
Actinomycin d, by Enzo Life Sciences (1 mentions)
Hek293, by Thermo Fisher Scientific (1 mentions)
Mccoy s 5a medium, by Thermo Fisher Scientific (1 mentions)
Dulbecco s modified eagle s medium, by Thermo Fisher Scientific (1 mentions)
U0126, by Merck Group (1 mentions)
Hek293, by American Type Culture Collection (1 mentions)
Sk mel 19, by American Type Culture Collection (1 mentions)
Malme 3, by American Type Culture Collection (1 mentions)
Phospho creb, by Cell Signaling Technology (1 mentions)
Kn 93, by Selleck Chemicals (1 mentions)
Sb203580, by Selleck Chemicals (1 mentions)
Sotrastaurin, by Selleck Chemicals (1 mentions)
Ly294002, by Merck Group (1 mentions)
Dabrafenib, by Selleck Chemicals (1 mentions)
Trametinib, by Selleck Chemicals (1 mentions)
Tet system approved fbs, by Takara Bio (1 mentions)
4 protocols using pd325901
1
Cell Lines and Inhibitors for Melanoma Research
2
Detailed ChIP-PCR, Mutagenesis, and Reporter Assays
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The primers used for ChIP-PCR, mutagenesis, constructing P27p-PGL3 luciferase reporters and CREB genotyping primers were listed in Supplementary Table 1. The CREB gene was cloned into the BglII sites of pRVKM plasmid using primers listed in Supplementary Table 1. The real-time RT-PCR primers used to quantify Il17, Il17f, Il21, RORα, RORγt, IkBζ, Batf, AHR and β-actin, and the Il17p-PGL3, CNS2-Il17p-PGL3 luciferase reporter constructs were described previously (Chang et al., 2011 (link), Yang et al., 2008 (link)). Most of the protein kinase inhibitors were purchased from Selleck Chemicals LLC, including H89 (Cat# S1582) for PKA and MSK1/2, PD325901 (Cat#S1036) for ERK, SB203580 (Cat#S1076) for p38, sotrastaurin (Cat#S2791) for PKC-ϴ and KN-93 (Cat#S7423) for CamKIV. The PI3K inhibitor LY294002 (Cat#L9908) was purchased from Sigma. The CREB and p27 antibodies were purchased from Cell Signaling: CREB (Cat#9197, Clone#48H2) & phospho-CREB (Cat#9198, Clone#87G3) for ChIP assays, phospho-CREB (Cat#9187, Clone#87G3) for flowcytometry assays and p27 antibody (Cat#3688, Clone#D37H1) for western blotting.
3
Melanoma Cell Line Viability Assay
4
Therapeutic Screening of Cancer Drugs
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Erlotinib and dasatinib were from LC laboratories. Docetaxel and doxorubicin were from Sigma. PD325901 and BEZ235, were from Selleck Chemicals. GDC-0941 was synthesized at Genentech. Gemcitabine was from Toronto Research. TGF-β1 was from R&D Systems Inc. Additional drugs used in the screen are described in Table S3 .
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