Evaluating miR-455-3p Binding Sites in SOX2-OT and PTEN

Predicted SOX2-OT or PTEN sequences containing miR-455-3p binding sites were inserted into the pmirGLO (Promega, WI, USA) and amplified by PCR to construct recombinants (Wild-type SOX2-OT 3ʹUTR (SOX2OT-WT)/SOX2-OT 3ʹUTR Mutant (SOX2OT-MUT) and Wild-type PTEN 3ʹUTR (PTEN-WT)/PTEN 3ʹUTR Mutant (PTEN-MUT)). Co-transfection was conducted with the Lipofectamine^TM 3000 reagent (Invitrogen) for 48 h. Luciferase activity was recognized using a luciferase activity assay kit (Promega, WI, USA), and the mean luciferase activity of cells was normalized to renilla luciferase activity [11–13 (link)].

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Yi C., Gu T., Li Y, & Zhang Q. (2022). Depression of long non-coding RNA SOX2 overlapping transcript attenuates lipopolysaccharide-induced injury in bronchial epithelial cells via miR-455-3p/phosphatase and tensin homolog axis and phosphatidylinositol 3-kinase/protein kinase B pathway. Bioengineered, 13(5), 13643-13653.

Publication 2022

pmirGLO LipofectamineTM 3000 reagent luciferase activity assay kit

Assay Binding sites Luciferase Promega Pten Renilla luciferase Sox2 Transfection

Corresponding Organization : Nanjing Medical University

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Variable analysis

independent variables

SOX2-OT 3'UTR (Wild-type and Mutant)
PTEN 3'UTR (Wild-type and Mutant)

dependent variables

Luciferase activity

control variables

Renilla luciferase activity

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

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