OvCa cells were maintained in standard DMEM supplemented with 10% FB-Essence (VWR Life Science SeraDigm, Radnor, PA) with 1% penicillin/streptomycin (Corning, Corning, NY), 1% MEM non-essential amino acids (Corning) and 1% MEM vitamins (Corning). All cell lines were genotyped to confirm their authenticity (IDEXX Bioresearch short tandem repeat marker profiling every 3 months), and were used between 2-12 passages from first thaw. All experiments were carried out in low glucose DMEM (VWR Life Science) supplemented as above. The pLX304 SPHK1-V5 lentiviral expression vector was purchased from DNASU (Tempe, AZ). The constitutively active HIF1α (P402A/P564A) and HIF2α (P405A/P531A) plasmids were a gift from William Kaelin (27 (link)) obtained through Addgene (Cambridge, MA). 5HRE-GFP reporter was a gift from Martin Brown and Thomas Foster obtained through Addgene (28 (link)). SPHK1 siRNA was obtained from GE Dharmacon (Lafayette, CO). Metformin, phenformin, CoCl2, and thiazolyl blue tetrazolium bromide were purchased from Sigma Aldrich (St. Louis, MO). S1P (Huzzah S1P) was purchased from Avanti Polar Lipids (Alabaster, AL). Hypoxia was induced through 1% O2/5% CO2 using Hypoxia Incubator Chamber by STEMCELL Technologies (Vancouver, BC, Canada).
Hif1α p402a p564a
Manufactured by Addgene
HIF1α (P402A/P564A) is a plasmid construct that expresses a mutant form of the Hypoxia-Inducible Factor 1-alpha (HIF1α) protein. The P402A and P564A mutations prevent the hydroxylation and subsequent degradation of HIF1α, leading to its stabilization under normoxic conditions.
Automatically generated - may contain errors
Lab products found in correlation
Hif2α p405a p531a, by Addgene (2 mentions)
Mem non essential amino acid, by Corning (1 mentions)
Penicillin streptomycin, by Corning (1 mentions)
Hypoxia incubator chamber, by STEMCELL (1 mentions)
Metformin, by Merck Group (1 mentions)
Thiazolyl blue tetrazolium bromide, by Merck Group (1 mentions)
Cocl2, by Merck Group (1 mentions)
Mem vitamins, by Corning (1 mentions)
Phenformin, by Merck Group (1 mentions)
Anti hif 1α antibody, by BD (1 mentions)
Dual glo luciferase assay, by Promega (1 mentions)
Anti hif 2α antibody, by Novus Biologicals (1 mentions)
E cadherin, by Cell Signaling Technology (1 mentions)
Anti α β tubulin, by Cell Signaling Technology (1 mentions)
2 protocols using hif1α p402a p564a
1
Hypoxia Regulation of Ovarian Cancer
2
Regulation of CEMIP Expression by HIF
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Oligo primers were synthesized by Operon. The anti-KIAA1199 (CEMIP) antibody was purchased from Abcam. The anti-HIF-1α antibody was purchased from BD Transduction Laboratories and the anti-HIF-2α antibody was purchased from Novus Biologicals. The anti-α/β-tubulin, -E-cadherin, and -Jarid1A antibodies were purchased from Cell Signaling Technology. HIF-1α P402A/P564A (plasmid 18955), HIF-2α P405A/P531A (plasmid 18956), and Jarid1A plasmids (plasmid 14800) were purchased from Addgene. Dual Glo-Luciferase assay was purchased from Promega. aiRNA against CEMIP was purchased from Boston Biomedical, Inc. [27 ].
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