The 3′‐UTR of MCU was synthesized and inserted into the psiCHECK‐2 vector (Ambion). The possible miR-129-3p binding sites in the 3′-UTR (MCU: AAGGGCT) were replaced with TCACTGA and validated via sequencing. The two reporter plasmids named MCU-WT and MCU-MUT were co-transfected into rat primary hippocampal neuronal cells with miR-129-3p mimic or NC mimic, respectively. The Dual Luciferase Reporter Assay kit (Promega, USA) with renilla luciferase as an internal control was used to perform luciferase assays, and The dual-luciferase activity was detected by Promega’s Digital-Luciferase Reporter Assay System (Promega, USA).
Digital luciferase reporter assay system
Manufactured by Promega
Sourced in United States
The Digital-Luciferase Reporter Assay System is a laboratory equipment designed for the detection and quantification of luciferase-based reporter gene activity. The system utilizes digital technology to provide precise and sensitive measurements of luciferase reporter signals.
Automatically generated - may contain errors
2 protocols using digital luciferase reporter assay system
1
miR-129-3p Regulation of MCU
2
Validating miR-129-1-3p binding to MCU
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The 3′-untranslated region (3′-UTR ) of MCU was synthesized and inserted into the psiCHECK2.0 vector. The 3′-UTR of MCU (AAGGGCT), the possible miR-129-1-3p binding sites, was substituted with TCACTGA and validated via sequencing. The MCU-WT and MCU-MUT reporter plasmids were cotransfected into chicken granulosa cells with miR-129-1-3p mimic or NC mimic, respectively. The Dual Luciferase Reporter Assay kit (Yeasen, Shanghai, China) was used to perform luciferase assays. The Promega's Digital-Luciferase Reporter Assay System (Promega, Madison, WI) was used to detect dual-luciferase activity.
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