CNE-1 cells were obtained from Institute of Virology, Chinese Academy of Preventive Medicine; NP-69 (the SV40 large T immortalized nasopharyngeal epithelial cell line) and CNE-2 cells were purchased from the Shanghai Institute of Cell Biology (Shanghai, P.R. China). The EBV+ C666-1 cells were from the cell bank of Xiangya Central Laboratory (Central South University, Changsha, P.R. China). Cells were maintained in RPMI-1640 or DMEM/F12 (1:1) medium (Gibco, Thermo Fisher Scientific, Waltham, MA, USA) containing streptomycin, penicillin, and supplemented with 10% fetal bovine serum (Gibco, Thermo Fisher Scientific) and cultured at 37°C in a humidified incubator with 5% CO2. For functional analysis, C666-1 cells grown to 80%–90% confluence were transfected with the miR-206 mimic, the miR-206 inhibitor, or scrambled miRNA as a control (Genepharma Company, Suzhou, P.R. China) using Lipofectamine RNAimax (Invitrogen, Thermo Fisher Scientific, Waltham, MA, USA). Cells were harvested 48–72 h after transfection for further experiments. Lentiviral transduction of C666-1 cells was carried out with lentivirus carrying miR-206 precursor and its control according to the manufacturer’s protocol (Genechem Company, Shanghai, P.R. China). Stable miR-206-overexpressing cells and the control cells were then selected using puromycin.
Scrambled mirna
Manufactured by GenePharma
Sourced in China
Scrambled miRNA is a laboratory tool used to generate random, non-targeting sequences of microRNA (miRNA) molecules. It serves as a control for experiments involving miRNA studies, allowing researchers to differentiate the effects of specific miRNA targets from potential off-target or background influences.
Automatically generated - may contain errors
Lab products found in correlation
Opti mem medium, by Thermo Fisher Scientific (2 mentions)
Mir 128 mimic, by GenePharma (2 mentions)
Oligonucleotides, by GenePharma (2 mentions)
Lipofectamine rnaimax, by Thermo Fisher Scientific (1 mentions)
Dmem f12 1 1 medium, by Thermo Fisher Scientific (1 mentions)
Rpmi 1640, by Thermo Fisher Scientific (1 mentions)
Mir 206 inhibitor, by GenePharma (1 mentions)
Lentivirus, by Genechem (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Lipofectamine rnaimax transfection, by Thermo Fisher Scientific (1 mentions)
Mir 10a mimic, by GenePharma (1 mentions)
Lipofectamine rnaimax transfection reagent, by Thermo Fisher Scientific (1 mentions)
Plvthm, by Addgene (1 mentions)
Lipofectamine 2000, by Thermo Fisher Scientific (1 mentions)
Cont mir, by Sangon (1 mentions)
Mir 379 mimics, by GenePharma (1 mentions)
Pcdna3, by RiboBio (1 mentions)
Fugene 6 transfection reagent, by Promega (1 mentions)
Lipofectamine 2000 reagent, by Thermo Fisher Scientific (1 mentions)
Mir 128 aso, by GenePharma (1 mentions)
7 protocols using scrambled mirna
1
Modulating miR-206 Expression in Nasopharyngeal Carcinoma Cells
2
MicroRNA Transfection and Analysis
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All of the cells were transfected with 20 nM of either miR-10a or miR-10b mimic (GenePharma) using the Lipofectamine RNAiMAX transfection reagent and Opti-MEM medium (Life Technologies) according to the manufacturer’s instructions. The control cells were transfected with 20 nM scrambled miRNA (GenePharma). Cells were incubated for 48 hours before being subjected to subsequent analysis unless otherwise mentioned.
3
Overexpression of miR-10a in Cells
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All cells were transfected with 20 nM of miR-10a mimics (GenePharma), using Lipofectamine RNAiMAX transfection reagent and Opti-MEMmedium (Life Technologies), according to the manufacturer’s instructions. Control cells were transfected with 20 nM scrambled miRNA (GenePharma). Cells were incubated for 48 h before being subjected to subsequent analysis, unless otherwise mentioned.
MiR-10a precursor sequence and flank sequences were cloned into MDH1-PGK-GFP plasmids (addgene no. 11375) and retrovirus were packaged in HEK293T cells. PTEN shRNAs were designed using the on-line design program from MIT (http://sirna.wi.mit.edu/home.php ). The 19-nucleotide hairpin-type shRNAs with a 9-nucleotide loop were cloned into pLVTHM (Addgene, Cambridge, USA). The lentivirus was packaged in HEK293T cell. Green fluorescent protein (GFP) sorting was used to isolate successfully transfected cells. qRT-PCR and Western blot was used for validation of knockdown effect.
MiR-10a precursor sequence and flank sequences were cloned into MDH1-PGK-GFP plasmids (addgene no. 11375) and retrovirus were packaged in HEK293T cells. PTEN shRNAs were designed using the on-line design program from MIT (
4
miR-128 Mimics and Transfection Protocol
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miR-128 mimics and cont-miR were synthesized by Sangon Biotechnology (Sangon, Shanghai, China), and cotransfections were performed with Lipofectamine 2000 (Invitrogen). The oligonucleotides (GenePharma, Shanghai, China) were as follows: miR-128 mimics, 5′-UCACAGUGAACCGGUCUCUUU-3′(sense) and 5′-AGAGACCGGUUCACGGUGAUU-3′ (anti-sense); scrambled miRNA (NC), 5′-UUCUCCGAA CGUGUCACGUTT-3′(sense) and 5′-ACGUGACACGU UCGGAGAATT-3′ (anti-sense).
5
Modulating miR-103 in Rat MCAO Model
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The rats were divided into 4 groups with 10 in each group. (1 (link)) Sham group: Sham-operated rats were with separated blood vessel but without thread insertion; (2 (link)) MCAO group: Rats were given miR-103 negative control (NC) scramble by intracerebroventricular injection about 5 min after MCAO; (3 (link)) MCAO+miR-103 mimic group; (4 (link)) MCAO+miR-103 inhibitor group: Rats were given 10 μg of miR-103 mimic, and miR-103 inhibitor by intracerebroventricular injection about 5 min after MCAO respectively. Scrambled miRNA, and miR-103 mimic/inhibitor were purchased from Shanghai GenePharma Co, Ltd, which were wrapped using PEG liposomes before administration. At the 24 hr after MCAO, the brain tissues of rats were taken out immediately after decapitation and frozen in liquid nitrogen at -80°C for later use.
6
miRNA Mimics and Inhibitors Synthesis
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miR-379 mimics, miR-379 inhibitors, and scrambled miRNA were purchased from GenePharma (Shanghai, China). The empty vectors pcDNA3.1 and pcDNA3.1-IGF-1 were synthesized by Ribobio (Guangzhou, China).
7
Investigating miR-128 and SNAI1 in Cells
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The oligonucleotides (GenePharma, Shanghai, China) were as follows: miR-128 mimics 5′-UCACAGUGAACCGGUCUCUUU-3′ (sense), 5′-AGAGACCGGUUCACGGUGAUU-3′ (anti-sense); scrambled miRNA (NC) 5′-UUCUCCGAACGUGUCACGUTT-3′ (sense), 5′-ACGUGACACGUUCGGAGAATT-3′ (anti-sense); miR-128 ASO 5′-AAAGAGACCGGUUCACUGUGA-3′ and miR-128 ASO (NC) 5′-CAGUACUUUUGUGUAGUACAA-3′. The SNAI1 shRNA and control shRNA plasmids were obtained from GenePharma (Shanghai, China). SNAI1 plasmid was purchased from ADDGENE (catalogue # 16225), and p-EGFP C2 empty plasmid was from Invitrogen. According to the product specification, the oligonucleotides were transfected into cells with Lipofectamine 2000 reagent (Invitrogen) and FuGENE 6 Transfection Reagent (Promega) were used for plasmid transfection.
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