Dual-Luciferase Reporter Plasmids for Mammalian Cells: Introduction and Applications

1. Recommended Plasmids

pmirGLO

pmirGLO is a commonly used dual-luciferase reporter vector.

Reporter gene: Firefly luciferase (Fluc)

Internal control gene: Renilla luciferase (Rluc)

For detailed plasmid information, please visit the pmirGLO product page.

psiCHECK-2

psiCHECK-2 is another commonly used dual-luciferase reporter vector.

Reporter gene: Renilla luciferase (Rluc)

Internal control gene: Firefly luciferase (Fluc)

For detailed plasmid information, please visit the psiCHECK-2 product page.

2. Common Applications

1. Promoter Activity Analysis

The promoter region of interest can be divided into different fragments for serial deletion analysis, or specific sites can be mutated to identify functional regulatory elements.

These promoter fragments or mutant sequences are then cloned into a luciferase reporter vector, and reporter activity is measured to evaluate promoter activity and identify important regulatory regions.

2. Validation of Transcription Factor-Mediated Regulation

A regulatory DNA sequence of interest can be inserted into a luciferase reporter vector and co-transfected with a plasmid overexpressing the transcription factor of interest.

Changes in luciferase activity can then be analyzed to determine whether the transcription factor affects the activity of the regulatory sequence.

3. Validation of miRNA Target Sites

To investigate whether a specific sequence is a direct target of a microRNA (miRNA), the sequence of interest can be cloned into the 3′ UTR of a luciferase reporter gene and co-transfected with the corresponding miRNA or miRNA mimic.

If luciferase activity decreases significantly compared with the appropriate control, this provides evidence that the tested sequence may be targeted by the miRNA.

4. Analysis of Transcription Factors in Signaling Pathways

A downstream response element of a signaling pathway can be cloned into a luciferase reporter vector.

Luciferase activity can then be measured under different upstream stimulation or treatment conditions to evaluate the activity of the downstream signaling pathway and determine how the pathway responds to specific regulatory signals.