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The Luc-Pair™ miR Luciferase Assay Kit provides an efficient system in a convenient 96-well plate format for measuring firefly and Renilla luciferases sequentially. Optimized for use with GeneCopoeia miRNA 3' UTR Target Sequence Expression Clones, Luc-Pair™ miR assays, enable easy, cost effective validation of miRNA activity.
Figure 1. The inhibitory effect of miRNA on a target sequence (3' UTR) expression clone can be measured with GeneCopoeia Luc-Pair™ miR Luciferase Assay Kit. HEK 293 cells were plated on a 6-well plate. On the second day, the cells were transfected with 1.0 µg of target sequence (3' UTR) expression clone (pEZX-MT01-Lin28 UTR-fLuc) and 1.4 µg of miRNA expression vector (or miRNA control vector) as indicated in the figure. The cells were transferred to a 96-well plate 18 hours after transfection and cultured for another 24 hours. Both firefly luciferase and Renilla luciferase activities were measured as described and data was recorded on Victor II machine. Firefly luciferase activity was then normalized with Renilla luciferase activities in the same well.
Figure 2a. Firefly Luciferase Activity
Figure 2b. Renilla Luciferase
Figure 2. Linear relationship between emitted light and amount of luciferase expression vector transfected in HEK293. HEK 293 cells were plated on a 6-well plate. On the second day, the cells were transfected with different amounts of GeneCopoeia pMRO or pEZX-MT01 miRNA 3' UTR Target Sequence Clone as indicated in the figure. The cells were transferred to a 96-well plate 18 hours after transfection and cultured for another 24 hours. Firefly luciferase assay was then performed on pMRO vector (Fig. 2a) and Renilla luciferase assay was performed on pEZX-MT01 vector (Fig. 2b).