GenePharma provided (Shanghai, China) miR-20b-5p mimic, miR-125a-5p mimic, miR-20b-5p inhibitor, miR-125a-5p inhibitor and their corresponding negative controls. Lipofectamine 2000® (Invitrogen, Carlsbad, CA, USA) was used for cell transfection following the manufacturer’s instructions.
Mir 20b 5p mimic
Manufactured by GenePharma
Sourced in China
MiR-20b-5p mimic is a synthetic RNA molecule designed to mimic the function of the endogenous miR-20b-5p microRNA. MicroRNAs (miRNAs) are small, non-coding RNA molecules that play a crucial role in the regulation of gene expression.
Automatically generated - may contain errors
Lab products found in correlation
Lipofectamine 2000 reagent, by Thermo Fisher Scientific (3 mentions)
Lipofectamine 2000, by Thermo Fisher Scientific (1 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (1 mentions)
Arpe 19, by Thermo Fisher Scientific (1 mentions)
Arpe 19, by Merck Group (1 mentions)
Pmir report plasmid, by Thermo Fisher Scientific (1 mentions)
Luciferase assay system, by Promega (1 mentions)
4 protocols using mir 20b 5p mimic
1
Cellular Transfection with miRNA Modulators
2
Glucose-induced Diabetic Retinopathy Model in ARPE-19 Cells
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Human RPE cell line ARPE‐19 (ATCC, USA) was cultured in DMEM (Gibco, USA) supplemented with 10% fetal bovine serum (FBS) and maintained in an incubator (5% CO2) at 37°C. The ARPE‐19 cells were exposed to 5 mM [normal glucose (NG)], 10 mM, 15 mM, 20 mM, or 25 mM (HG) of glucose (Sigma‐Aldrich, USA) for 72 h. The exposure of cells to high glucose (25 mM) was used to establish the cell model of diabetic retinopathy25 , 26 , 27 . Short hairpin RNA (shRNA) for circZNF532 (sh‐ZNF532), circZNF532 expression vector (oe‐ZNF532), miR‐20b‐5p mimic, miR‐20b‐5p inhibitor, STAT3 expression vector (oe‐STAT3), and their corresponding negative control (NC) (all by GenePharma, China) were introduced into ARPE‐19 cells alone or in combination using lipofectamine 2000 reagents (Invitrogen, USA).
3
Luciferase Reporter Assay for miRNA Targets
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MEG3 sequence or CREB1 3ʹ-UTR containing miR-20b-5p binding site was inserted into the pMIR-REPORT plasmid (Thermo Fisher Scientific, Waltham, MA, USA) to construct the wild-type plasmids of MEG3 and CREB1 (MEG3 WT and CREB1 WT), respectively. Subsequently, the MEG3 fragment of the mutant sequence or CREB1 3ʹ-UTR was inserted into the plasmid respectively to construct the mutant-type plasmids (MEG3 MUT and CREB1 MUT). The constructed luciferase reporter plasmids were then co-transfected with miR-20b-5p mimic or its NC (GenePharma) into 293 T cells (ATCC) using the Lipofectamine 2000 reagent (Invitrogen). After 48 h, a luciferase assay system (Promega, Madison, WI, USA) was employed for luciferase activity analysis [24 (link)].
4
Inflammatory Colon Cell Response to EVs
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Conditionally immortalized murine colon epithelial cell line was procured from Crisprbio (#CE19668; Beijing, China), and then cultured in RPMI-1640 medium comprising of penicillin-streptomycin-glutamine and 5% FBS at 37°C with 5% CO2 in air [22 (link)]. Young adult mouse colon (YAMC) cells were treated with 1 μg/mL LPS for 24 h to simulate the inflammatory environment of UC in vitro [28 (link)]. After 30 min of pre-stimulation, YAMC cells were subjected to treatment with 100 μg/mL M2-EVs, with the conditioned medium of the GW group serving as the control. miR-20b-5p mimic and its NC were provided by GenePharma and transfected into YAMC cells using the Lipofectamine 2000 reagent (Invitrogen Inc., Carlsbad, CA, USA).
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