mDP cells were derived from mouse dental mesenchymal cells40 (link),52 (link), and maintained in Dulbecco’s modified Eagle’s medium (DMEM)/F-12, supplemented with 10% fetal bovine serum (Gibco/Life Technologies), and 1% penicillin/streptomycin (Gibco/life Technologies) at 37 °C in a humidified atmosphere of 5% CO2. For western blotting and scratch assays, cells were cultured with 10 ng/ml of mouse recombinant protein PDGF-AA or PDGF-BB (#315-17-2UG, #315-18-2UG, PeproTech) for 24 hours. Constructed vectors, or mimic miRNA (miR875-5p, MSY0004937, Qiagen) or control miRNA (YM00479902, Qiagen) were transfected using a Neon Transfection System (Thermo Fisher Scientific), according to the manufacturer’s protocol.
Ym00479902
Manufactured by Qiagen
Sourced in Germany
The YM00479902 is a laboratory equipment product manufactured by Qiagen. It is a compact and versatile instrument designed for DNA/RNA purification and sample preparation. The core function of this product is to provide reliable and efficient nucleic acid extraction and purification from a variety of sample types.
Automatically generated - may contain errors
Lab products found in correlation
Lipofectamine 2000, by Thermo Fisher Scientific (2 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions)
Neon transfection system, by Thermo Fisher Scientific (1 mentions)
Yi00199006, by Qiagen (1 mentions)
Mircury lna mirna inhibitor, by Qiagen (1 mentions)
Hiperfect transfection reagent, by Qiagen (1 mentions)
Mircury lna mirna mimic, by Qiagen (1 mentions)
Opti mem reduced serum medium, by Thermo Fisher Scientific (1 mentions)
3 protocols using ym00479902
1
mDP Cell Culture and Manipulation
2
Transfection of miRNA mimics and inhibitors in hESCs
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Hsa-miR mimics (339173, miRCURY LNA miRNA Mimic) and inhibitors (339121, miRCURY LNA miRNA Inhibitors) were ordered from Qiagen, Hilden, Germany. Additionally, negative control mimics (YM00479902, Negative Control miRCURY LNA miRNA Mimic) and negative control inhibitors (YI00199006, Negative control A), which both have no homology to any known miRNA or mRNA in human, were ordered from Qiagen, Hilden, Germany. The following miRNAs were labeled with a fluorescent 5′-carboxyfluorescein (FAM) tag: hsa-miR-19a mimic + inhibitor, hsa-miR-19b mimic + inhibitor, negative control A mimic, negative control inhibitor. Thirty thousand decidualized or non-decidualized hESCs were cultured in single wells of a 48 wells plate and transfected with 25 nM miRNA mimic or negative control mimic. Alternatively, 30 000 hESCs were transfected with 150 nM miRNA inhibitor or negative control inhibitor. All transfections were performed in D- or C-medium by using HiPerFect Transfection Reagent according to manufacturer’s instructions (301704, Qiagen, Hilden, Germany). After transfection for 24 h, hESCs were washed with PBS and transfection efficiency was determined by calculating the percentage of fluorescently labeled hESCs from the total number of hESCs per image.
3
Dual-Luciferase Assay for miRNA Targeting
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0.5 × 106 HEK293 cells were plated on a 6-well plate and incubated overnight. The next day, the HEK293 cells were transfected by using Lipofectamine 2000 (Thermo Fisher Scientific, Waltham, MA, USA). In short, 2 ug of pMiR-REPORT-KLF7-3′UTR and 1 ug of pGL4.73[hRluc/SV40] (renilla) vector (Promega, Madison, WI, USA) together with 5 nM of hsa-miR-132-3p miRCURY LNA miRNA mimic (YM00472088, Qiagen, Hilden, Germany) or negative control miRCURY LNA miRNA mimic (YM00479902, Qiagen, Hilden, Germany) was added in a tube containing 250 µL Opti-MEM Reduced Serum Medium (Gibco, Carlsbad, CA, USA). Another tube was prepared containing a mastermix of 250 µL Opti-MEM and 5 µL of Lipofectamine 2000 per transfection. The tubes were mixed at RT for 5 min before combining the Lipofectamine 2000 mixture with the DNA mixture. The tubes were mixed again before incubating at RT for 20 min. The transfection complexes were added drop-wise to the cells and incubated overnight. The next day, the medium was changed. The cells were collected 72 h after transfection.
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