Sequences of Fgf9ECR1a, Fgf9ECR1b, Fgf10hypso+, and Fgf10hypso− were inserted into the KpnI and HindIII sites in the pGL3-enhancer reporter vector (Promega). Mouse ameloblast-derived LS8 cells were cultured in DMEM with 10% FBS at 37°C and 5% CO2. Cells (1×105) were plated in 35mm plates. Each transfection was standardized for 0.5 μg of expression vector, 1 μg luciferase reporter vector, and 1 μg pTK-Renilla vector. Fugene (3 μl) and plasmids were mixed in 97 μl DMEM (Invitrogen). The mixture was incubated for 20 min at room temperature and added to the cells. The cells were cultured for 48 h, lysed and measured for luciferase activity by Dual-Glo Luciferase Assay System (Promega) standard protocol in a Glomax Luminometer (Promega). The Renilla luciferase vector was used for normalization. Assays were carried out to achieve n=3. Differences in luminescence level and standard deviation were calculated and significance was evaluated by student’s t-test (Sherf, 1996 ).
Pgl3 enhancer reporter vector
Manufactured by Promega
The PGL3-enhancer reporter vector is a plasmid used for gene expression studies. It contains a multiple cloning site upstream of the firefly luciferase reporter gene, allowing the insertion of DNA sequences to be tested for enhancer activity.
Automatically generated - may contain errors
Lab products found in correlation
2 protocols using pgl3 enhancer reporter vector
1
Investigating Fgf Enhancer Activities
2
Regulation of BIRC6 3'-UTR by miR-181a
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Based on the human BIRC6 mRNA sequence deposited in the GenBank database (accession no. NM_016252), the 3′-UTR of BIRC6 (1010 nt, 14,708–15,718) was amplified by PCR and subcloned in the psiCHECK-2 luciferase vector (Promega, Madison, WI, USA). The promoters of Bax containing the core promoter elements and p53 binding site were PCR amplified from genomic DNA and cloned in pGL3 enhancer reporter vector (Promega). Mesangial cells, HCT116 p53+/+ cells, or HCT116 p53−/− cells were transfected with corresponding luciferase reporter and a control vector expressing β-gal using lipofectamine 2000. After 24 h, miR-181a mimics, miRNA negative control, miR-181a inhibitor, or miRNA inhibitor negative control was transfected into cells for 48 h. The cell lysates were assayed for luciferase activity using the Luciferase Assay System (Promega) with a luminescence counter (Centro SX3 LB 960, Berthold Technologies, Bad Wildbad, Germany). Luciferase activity was normalized by the corresponding β-gal luciferase activity.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!