HeLa cell transfection was performed in a 96-well plate at 80% confluency. The vector containing the HIF-1α response element pGL4.42[luc2P/HRE/Hygro] (Promega) was co-transfected with the control vector phRL-TK (Promega) as a transfection efficiency control. Forty-eight hours later, luciferase activity was measured using the Dual-Glo Luciferase Assay System (Promega). Luciferase activity was normalized to that of Renilla luciferase activity.
Pgl4.42 luc2p hre hygro
Manufactured by Promega
Sourced in United States
The PGL4.42[luc2P/HRE/Hygro] is a reporter vector that contains the firefly luciferase gene (luc2P) under the control of a hypoxia response element (HRE) promoter and a hygromycin resistance gene for selection in mammalian cells. This vector is designed to measure hypoxia-inducible gene expression.
Automatically generated - may contain errors
Lab products found in correlation
Dual glo luciferase assay system, by Promega (1 mentions)
Phrl tk, by Promega (1 mentions)
Prl tk, by Promega (1 mentions)
Effectene transfection reagent, by Qiagen (1 mentions)
Passive lysis buffer (plb), by Promega (1 mentions)
Dual luciferase reporter assay system, by Biotium (1 mentions)
Glomax multi plus plate reader, by Promega (1 mentions)
2 protocols using pgl4.42 luc2p hre hygro
1
HIF-1α Transcriptional Activity Assay
2
Hypoxia-Responsive Dual Luciferase Assay
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Cells were seeded at a density of 70,000 cells/well in a 12-well plate, grown overnight, and co-transfected with 187.5 ng pGL4.42[luc2P/HRE/Hygro] (Promega, Madison, WI, USA, E4001) and 12.5 ng pRL-TK (Promega, Madison, WI, USA, E2241) luciferase vectors by Effectene Transfection Reagent (Qiagen, Qiagen, MD, USA, 301425). Next day, cells were kept under various temperatures and oxygen concentrations for 4 and 24 h. Cells were lysed in 1x Passive Lysis Buffer (Promega, Madison, WI, USA, E1941) with gentle shaking at RT for 15 min. The activities of firefly and renilla luciferase were measured using the Dual Luciferase Reporter Assay System (Biotium, Fremont, CA, USA, 30081) according to manufacturer’s protocols. Luminescence was measured on a Promega GloMax Multiplus Plate Reader using a 10-s pre-read delay, followed by a 5-s measurement period.
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