Author: Hu, Zhiqiang; Xing, Yaling; Qian, Yuanyu; Chen, Xiaojuan; Tu, Jian; Ren, Lening; Wang, Kai; Chen, Zhongbin
Title: Anti-radiation damage effect of polyethylenimine as a toll-like receptor 5 targeted agonist Document date: 2012_10_26
ID: 6fp8nly0_10
Snippet: Luciferase reporter gene assay was performed as previously described [15] . Briefly, HEK 293T cells were placed in a 96-well plate at a density of 8 × 10 3 for 24 h. The pcDNA3.1-V5/HisB-hTLR5 was transiently co-transfected with the NF-κB-Luc and TK-PRL (an internal control for luciferase gene expression) by using Lipofectamine TM 2000. Twenty-four hours after transfection, the cells were treated with PEI or flagellin (a known TLR5 agonist), as.....
Document: Luciferase reporter gene assay was performed as previously described [15] . Briefly, HEK 293T cells were placed in a 96-well plate at a density of 8 × 10 3 for 24 h. The pcDNA3.1-V5/HisB-hTLR5 was transiently co-transfected with the NF-κB-Luc and TK-PRL (an internal control for luciferase gene expression) by using Lipofectamine TM 2000. Twenty-four hours after transfection, the cells were treated with PEI or flagellin (a known TLR5 agonist), as a positive control, for 6 h, then washed and lysed. Luciferase activity was measured using a dual-luciferase assay kit according to the manufacturer's instructions. The increased amount of promoter activity was expressed as the ratio of induction, which was the relative luciferase activity of the stimulated cells divided by the relative luciferase activity of the mock cells. The assays were repeated three times independently.
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