The company is a leading supplier of genetically defined cell lines, reporter gene assay
kits, genomic reference standards, and contract research services.
Bioinformatics analysis confirmed the existence of miR375 target sites in the 3' UTR of RLF and HIGD1A, and the dual luciferase reporter gene assay
further validated that RLF and HIGD1A were targeted by miR-375.
Dual-Luciferase Reporter Gene Assay
. NOX4 promoter (-2000-0) luciferase reporter plasmid was constructed adopting the GV238-REPORT vector (GENE, Shanghai, China).
Dual-luciferase reporter gene assay
. Renilla luciferase gene activity was reduced by half in 293T cells transfected with miR-181b mimics and the wild-type FAMLF 5' UTR sequence (FAMLF-WT/miR-181b), while miR-181b negative controls (fAMLF-MT/NC) and the mutated FAMLF 5' UTR sequence (FMALF-MT/miR-181b) caused no statistical changes in luciferase gene expression.
Cell Transfection for Reporter Gene Assay
and Gene Expression Analysis.
ADCC Reporter Gene Assay
. For assays using cells in culture, Jurkat effector cells were harvested and resuspended in RPMI 1640 medium with 10% heat-inactivated FBS and 1.2 x [10.sup.5] cells in 20 [micro]l were added to each well of a whitesided microtiter plate (Perkin Elmer, Waltham, MA).
Transient Transfection and Luciferase Reporter Gene Assay
. Hek-293 cells were grown to 60% confluence in 24well plates for 24 h.
The luciferase reporter gene assay
was performed using pGL3-[(CAGA).sub.12], a transcriptional reporter that can be driven by TGF-beta]/Smad3/Smad4 pathway (Chen et al., 2011).
Most of these are primarily binding-based assays, including solid-phase ELISA methods (8, 45, 49) and liquid-phase, HPLC-based homogenous mobility shift assay (HMSA) (50); in addition, a functional cell-based reporter gene assay
(RGA) has been developed (51 ) and is currently available for clinical use (52).
For example, the PPAR response element used in the Attagene cis-PPRE reporter gene assay
will bind PPAR[delta] and PPAR[alpha], RXR[alpha],[beta],[gamma], COUPTF[alpha],[beta], and HNF4 (Nakshatri and Bhat-Nakshatri 1998) at a minimum.
We then showed that Ngn3 could bind to the promoter of Sohlh1andincrease the transcription of Sohlh1 by ChIP and luciferase reporter gene assay
. Lastly, we demonstrated that the protein of Ngn3 and Sohlh1 could bind together in wild type testis by immune precipitation.
The possible lycopene-induced modulation of transcriptional response to cytokine was tested using reporter gene assay
. Due to the induced high mortality in the transfected THP1 cells after LPS treatment, we used TNF[alpha] and PMA as stimuli to enhance gene reporter expression in transfected THP1 cells since these proinflammatory molecules did not induce apoptosis in the transfected cells (data not shown).