pt3 myr akt ires luciferase  (Addgene inc)

Journal: Nucleic Acids Research

Article Title: Two redundant transcription factor binding sites in a single enhancer are essential for mammalian sex determination

doi: 10.1093/nar/gkae178

Figure Lengend Snippet: Luciferase Reporter Assays of the various mutations in TFBS in Enh13. (A–C) Luciferase reporter assays of HEK293T cells co-transfected with combinations expression plasmids of NR5A1, SOX9, SRY along with Luciferase construct containing the Enh13 WT, Enh13 SOX9*5bp - SRY*6bp ( A ) or Enh13 Ogawa containing the Poly G sequences ( B ). All values were normalized to the Enh13 WT luciferase activity with no expression plasmids added. Values were also normalized to the Renilla expression. ( C ) Luciferase reporter assays of HEK293T cells co-transfected with combinations expression plasmid of WT1 (-KTS) along with Luciferase construct containing the Enh13 WT and Enh13 Ogawa containing the Poly G sequences. All values were normalized to the Enh13 WT luciferase activity with no expression plasmids added. Means and SEM from at least three independent experiments are shown. Statistical analysis was done using two-way ANOVA followed by Tukey's/Dunnett's posttest. * P < 0.05, ** P < 0.01, *** P < 0.001 and **** P < 0.0001, ns- not significant. WT, wild type, n = number of biological replicates.

Article Snippet: The following expression plasmids for luciferase assays were used: pCMV3-hWT1-KTS-C-HA (SINO BIOLOGICAL, Ref: HG12282-CY), pcDNA3-Flag-hSOX9 (A gift from Gerd Scherer to FP, described in ( )), pcDNA3-2X-MYC-hSF1, and pcDNA3-mSRY-4X-Myc.

Techniques: Luciferase, Transfection, Expressing, Construct, Activity Assay, Plasmid Preparation