Bioinformatics tools (microRNA.org ) were used to predict the miR-143 binding sites of PVT1. Human GBC-SD cells were transfected with 150 ng of empty pmirGLO-NC, pmirGLO-PVT1-wt or pmirGLO-PVT1-mut (GenePharma). Two nanograms of pRL-TK (Promega, WI, USA) were cotransfected with the miR-143 mimic or miR-NC into GBC-SD cells by using Lipofectamine 2000 (Invitrogen) following the manufacturer’s procedures. The relative luciferase activity was normalized to Renilla luciferase activity 48 h after transfection.
Pmirglo nc
Manufactured by GenePharma
Sourced in China
The PmirGLO-NC is a laboratory equipment product designed for gene expression analysis. It serves as a reporter vector for monitoring microRNA (miRNA) activity in cells. The core function of this product is to facilitate the study of miRNA regulation and its impact on gene expression.
Automatically generated - may contain errors
4 protocols using pmirglo nc
1
Predicting miR-143 Binding Sites in PVT1
2
Regulation of miR-519b-3p in Colorectal Cancer
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Bioinformatics tools TargetScanHuman 7.1 were employed to predict the Duxap8 binding sites of miR-519b-3p and the miR-519b-3p binding sites of ZNF277. Human colorectal cancer cells were transfected with 150 ng of empty pmirGLO-NC, pmirGLO-Duxap8-WT or pmirGLO-Duxap8-Mut (GenePharma) and the miR-519b-3p mimics or mimics NC were co-transfected into cells by using Lipofectamine 2000 (Invitrogen) following the manufacturer's procedures. The relative luciferase activity was measured after transfection. To verify the targeting relationship between miR-519b-3p and ZNF277, the principle is the same as above.
3
Dual-luciferase Assay of miR-27b and JMJD3
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Dual-luciferase reporter gene assay was performed to study the interaction between miR-27b and JMJD3. The potential target genes of miR-27b were predicted via bioinformatics analysis using Starbase (http://starbase.sysu.edu.cn/ ). The wild-type (wt) 3′untranslated region (UTR) of JMJD3 mRNA sequence containing the predicted target sites of miR-27b was synthesized. The reporter vectors containing JMJD3 wt (pmirGLO-Pygo2-wt) and JMJD3 mutant miR-27b-binding sequence (pmirGLO-JMJD3-mut) or the NC sequence (pmirGLO-NC) (GenePharma, Shanghai, China) were co-transfected into BMDMs with miR-27b-mimic or NC-mimic. After 24-h transfection, activities of firefly luciferase and Renilla luciferase were detected according to the manufacturer’s instruction for the dual-luciferase reporter assay system (Promega, Madison, WI, USA).
4
NLRP3 3'UTR Luciferase Assay
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The reporter plasmids, pmirGLO-NLRP3-3′-UTR, pmirGLO-NLRP3-3′-UTR-MUT and pmirGLO-NC were purchased from Shanghai GenePharma Co., Ltd. THP-1 cells were co-transfected with the plasmids and hsa-miR-223-3p mimic/inhibitor using Lipofectamine 3000 reagent (Invitrogen; Thermo Fisher Scientific, Inc.). Following incubation at 37°C for 24 h, luciferase activities were detected using a Dual-Luciferase Reporter assay system (Promega Corporation), and results were compared with Renilla luciferase activity.
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