plasmid containing the renilla luciferase gene behind the simian virus 40 early promoter (prl)  (Promega)

Journal: Virology

Article Title: Identification of two auto-cleavage products of nonstructural protein 1 (nsp1) in porcine reproductive and respiratory syndrome virus infected cells: nsp1 function as interferon antagonist

doi: 10.1016/j.virol.2009.11.033

Figure Lengend Snippet: PRRSV nsp1 proteins inhibit IFN-β production. (A, B) HEK293T cells cultured in 24-well plates were cotransfected with a plasmid expressing nsp1 proteins, a plasmid expressing influenza virus NS1, or pCAGGS empty vector (P), pRL-SV40, and a luciferase reporter plasmid p125-Luc (A) or pCIB-55-Luc (B). At 20 h post transfection, cells were infected with Sendai virus (SeV) for 16 h to stimulate the production of interferon. (C–H, J) HEK293T cells in 24-well plates were cotransfected with the plasmid pEFneo-RIG-I (C), pEFneo-MDA5 (D), pEGFP-IPS-1 (E), pEFneo-TBK1 (F), pEFneo-IKKɛ (G), or pCAGGS-IRF3 (H), or pcDNA3-TRIF (J), along with pRL-SV40, pCAGGS expressing nsp1 proteins, and pCIB-55 plasmid for 20–24 h. (I) HEK293T cells were cotransfected with pNF-kB-luc, pcDNA3-TRIF, pRL-SV40 and pCAGGS expressing nsp1 proteins or pCAGGS empty vector (P) for 20 h. Cells were harvested and measured for firefly and Renilla luciferase activities. Relative luciferase activity is defined as a ratio of firefly luciferase reporter activity to Renilla luciferase activity. Each data point shown represents a mean value from three experiments. Error bars show standard deviations of the normalized data.

Article Snippet: The pRL-SV40 plasmid that expresses a Renilla luciferase under the control of a simian virus (SV) 40 promoter was purchased from Promega (Madison, WI).

Techniques: Cell Culture, Plasmid Preparation, Expressing, Luciferase, Transfection, Infection, Activity Assay

Journal: Virology

Article Title: Identification of two auto-cleavage products of nonstructural protein 1 (nsp1) in porcine reproductive and respiratory syndrome virus infected cells: nsp1 function as interferon antagonist

doi: 10.1016/j.virol.2009.11.033

Figure Lengend Snippet: PRRSV nsp1 proteins inhibit expression from an ISRE promoter. HEK293T cells were cotransfected with pISRE-luc, pRL-SV40 and pCAGGS expressing nsp1 proteins or pCAGGS empty vector (P) for 20 h. Cells were then infected with Sendai virus (A) or treated with IFN-α (B) and IFN-β (C) for 20 h. The cells were harvested and measured for firefly and Renilla luciferase activities. Relative luciferase activity is defined as a ratio of firefly luciferase reporter activity to Renilla luciferase activity. Each data point shown represents a mean value from three experiments. Error bars show standard deviations of the normalized data.

Article Snippet: The pRL-SV40 plasmid that expresses a Renilla luciferase under the control of a simian virus (SV) 40 promoter was purchased from Promega (Madison, WI).

Techniques: Expressing, Plasmid Preparation, Infection, Luciferase, Activity Assay

Journal:

Article Title: Requirement of p53 targets in chemosensitization of colonic carcinoma to death ligand therapy

doi: 10.1073/pnas.2435285100

Figure Lengend Snippet: p53 is required for sensitizing Bax-/- cells to TRAIL. Bax-/- HCT116 cells were infected with Ad-E6 or Ad-LacZ for 16 h. The infected cells were treated as described in the Fig. 1 legend. (A) p53 protein was determined by Western blot and active caspase 3 was analyzed by flow cytometry. (B) Infected cells were transfected with PG13-Luc and pRL-SV40 plasmids. Luciferase activity was determined by Fisher luminometer (Lower). PG13-Luc and pRL-SV40 stably transfected Bax-/- HCT116 cells were treated as described above, images were obtained by using the cooled charge-coupled camera of the in vivo imaging system (Upper).

Article Snippet: The PG13-luc, a p53 reporter with a firefly luciferase gene under the control of 13 p53 response elements, has been described ( 11 ). pRL-SV40 (simian virus 40) vector was purchased from Promega, and this vector contains a cDNA encoding Renilla luciferase, which was originally cloned from the marine organism Renilla reniformis .

Techniques: Infection, Western Blot, Flow Cytometry, Transfection, Luciferase, Activity Assay, Stable Transfection, In Vivo Imaging

Journal:

Article Title: Requirement of p53 targets in chemosensitization of colonic carcinoma to death ligand therapy

doi: 10.1073/pnas.2435285100

Figure Lengend Snippet: Optical imaging of p53 activity in living mice. PG13-Luc and pRL-SV40 stably transfected Bax-/- HCT116 cells were infected with Ad-LacZ or Ad-E6 for 16 h. Cells (1 × 106) were s.c. implanted into left or right forearm. CPT11 (80 mg/kg) was administered by i.p. injection. After 0 or 16 h of treatment, coelenterazine was injected 3 min before imaging, 2 h apart, followed by d-luciferin injection (5 min before imaging). A whole body image was acquired by using the cooled charge-coupled device camera. Each image was acquired at the same time, relative to the injected substrate, and all of the images are shown at the same scale.

Article Snippet: The PG13-luc, a p53 reporter with a firefly luciferase gene under the control of 13 p53 response elements, has been described ( 11 ). pRL-SV40 (simian virus 40) vector was purchased from Promega, and this vector contains a cDNA encoding Renilla luciferase, which was originally cloned from the marine organism Renilla reniformis .

Techniques: Optical Imaging, Activity Assay, Stable Transfection, Transfection, Infection, Injection, Imaging