siRNA against 3-hydroxy-3-methyglutaryl-coA reductase (HMGCR) and scrambled control (ctrl) siRNA were purchased from Shanghai GenePharma Co., Ltd. (cat. no. A10001). miR-125a mimic (sense, 5′-UCCCUGAGACCCUUUAACCUGUGA-3′ and antisense, 5′-UCACAGGUUAAAGGGUCUCAGGGA-3′), miRNA mimic negative control (miR-NC, sense, 5′-UUUGUACUACACAAAAGUACUG-3′ and antisense, 5′-CAGUACUUUUGUGUAGUACAAA-3′), inhibitor (5′-UCACAGGUUAAAGGGUCUCAGGGA-3′) and miRNA inhibitor negative control (miR-NC, 5′-CAGUACUUUUGUGUAGUACAAA-3′) were purchased from Guangzhou RiboBio Co., Ltd. siRNA (100 nM) and plasmid (1 µg/ml) transfections were performed using Lipofectamine 3000 (Thermo Fisher Scientific, Inc.). miRNA mimic (50 nM) and miRNA inhibitor (50 nM) were transfected into VSMCs cultured under HG or normal conditions (5.6 nM glucose) by using Lipofectamine® RNAiMAX (Thermo Fisher Scientific, Inc.) according to the manufacturer's instructions. Western blotting and RT-qPCR analysis were used to detect the effects of gene silencing and miR-125a expression in VSMCs at 24 h after transfection (Fig. S1 ).
Mir nc
Manufactured by RiboBio
Sourced in China, United States, Puerto Rico
The MiR-NC is a laboratory equipment designed for the examination and analysis of microRNA (miRNA) expression. It serves as a tool for researchers to study the functional roles of miRNAs in various biological processes.
Automatically generated - may contain errors
Lab products found in correlation
Lipofectamine 2000, by Thermo Fisher Scientific (93 mentions)
Lipofectamine 3000, by Thermo Fisher Scientific (90 mentions)
Anti mir nc, by RiboBio (87 mentions)
Si nc, by RiboBio (77 mentions)
Lipofectamine 3000 reagent, by Thermo Fisher Scientific (43 mentions)
Lipofectamine 2000 reagent, by Thermo Fisher Scientific (39 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (30 mentions)
Sh nc, by RiboBio (23 mentions)
Anti nc, by RiboBio (19 mentions)
Si nc, by GenePharma (16 mentions)
Mir nc, by Thermo Fisher Scientific (14 mentions)
Pcdna3.1 vector, by Thermo Fisher Scientific (13 mentions)
In mir nc, by RiboBio (12 mentions)
Inhibitor nc, by RiboBio (12 mentions)
Lipofectamine rnaimax, by Thermo Fisher Scientific (11 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (11 mentions)
Rpmi 1640 medium, by Thermo Fisher Scientific (11 mentions)
Sh nc, by GenePharma (10 mentions)
Pcdna, by RiboBio (8 mentions)
Penicillin, by Thermo Fisher Scientific (8 mentions)
329 protocols using mir nc
1
Silencing HMGCR and Regulating miR-125a in VSMCs
2
Establishing CDDP-resistant Cell Line and Transfection
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CDDP-resistant cell line H1299 (H1299/CDDP) and the corresponding parental cells H1299 were purchased from the American Type Culture Collection (ATCC, Manassas, VA, USA). All cells were cultured in RPMI-1640 medium (Thermo Fisher Scientific, Inc., Waltham, MA, USA) supplemented with 10% fetal bovine serum (FBS; Gibco, Grand Island, NY, USA), and 1% penicillin/streptomycin at 37 °C in a humidified atmosphere containing 5% CO2. To maintain the drug-resistant phenotype, CDDP (Sigma, St. Louis, MO, USA) was added to the culture media at a final concentration of 1 μg ml−1 for H1299/CDDP cells.
miR-124 mimics (miR-124) and its matched negative control (miR-NC), miR-142 mimics (miR-142) and its corresponding negative control (miR-NC), small interfering RNA (si-RNA) against sirtuin 1 (SIRT1) (si-SIRT1) and control siRNA (si-NC) were purchased from RiboBio (Guangzhou, China). To overexpress SIRT1, the full-length of SITR1 was synthesized and inserted into pcDNA3.1-overexpressing vector (Invitrogen, Carlsbad, CA, USA) to produce pcDNA-SIRT1 (SIRT1), with pcDNA3.1 empty control (pcDNA) as a negative control. H1299/CDDP cells were seeded into 6-well plates at a density of 2 × 105 cells per well and transfected with these above oligonucleotides or plasmids using Lipofectamine 2000 (Invitrogen) when cells were grown to 70–80% confluence.
miR-124 mimics (miR-124) and its matched negative control (miR-NC), miR-142 mimics (miR-142) and its corresponding negative control (miR-NC), small interfering RNA (si-RNA) against sirtuin 1 (SIRT1) (si-SIRT1) and control siRNA (si-NC) were purchased from RiboBio (Guangzhou, China). To overexpress SIRT1, the full-length of SITR1 was synthesized and inserted into pcDNA3.1-overexpressing vector (Invitrogen, Carlsbad, CA, USA) to produce pcDNA-SIRT1 (SIRT1), with pcDNA3.1 empty control (pcDNA) as a negative control. H1299/CDDP cells were seeded into 6-well plates at a density of 2 × 105 cells per well and transfected with these above oligonucleotides or plasmids using Lipofectamine 2000 (Invitrogen) when cells were grown to 70–80% confluence.
3
Investigating miR-182 Inhibitors and FOXN3 Overexpression in Gallbladder Cancer
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miR-182 inhibitors (5′-TTCTACCATTGCCAA-3′) and miRNA negative control (miR-NC; 5′-ACGTCTATACGCCCA-3′) were purchased from Guangzhou RiboBio Co., Ltd. To the best of our knowledge, no effects of miR-NC on cell viability have been confirmed. GBC-SD and SGC-996 cells were seeded onto 6-well plates at a density of 1×105 cells/well. Once they reached 60% confluence, the cells were transfected with 50 nM miR-182 inhibitors and miR-NC using Lipofectamine 3000 reagent (Thermo Fisher Scientific, Inc.) for 24 h at 37°C, according to the manufacturer's protocol.
The pcDNA3.1-FOXN3 vector (Guangzhou RiboBio Co., Ltd) was prepared by cloning the open reading frame of FOXN3 into the vector. GBC-SD and SGC-996 cells were transfected with the pcDNA3.1-FOXN3 vectors for 24 h at 37°C, using Lipofectamine 3000 reagent.
The pcDNA3.1-FOXN3 vector (Guangzhou RiboBio Co., Ltd) was prepared by cloning the open reading frame of FOXN3 into the vector. GBC-SD and SGC-996 cells were transfected with the pcDNA3.1-FOXN3 vectors for 24 h at 37°C, using Lipofectamine 3000 reagent.
4
Knockdown of hsa_circ_0004396 in NSCLC
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For stable knocking down of hsa_circ_0004396, short hairpin RNA against hsa_circ_0004396 was applied. Both sh‐hsa_circ_0004396 and sh‐NC (GenePharma) were constructed into the lentiviral vector (Invitrogen), followed by introduction into A549 cells and cultured in cell medium with 2 μg/ml puromycin.
Meanwhile, based on the producer’s direction of Lipofectamine 3000 (Invitrogen), miR‐615‐5p mimics/inhibitor and its control (miR‐NC/anti‐miR‐NC), empty vector (pcDNA3.1), overexpression vectors of hsa_circ_0004396 (pcDNA3.1‐hsa_circ_0004396), and PAK1 (pcDNA3.1‐PAK1) from Ribobio were transfected into 2 × 105 NSCLC cells in six‐well plates.
Meanwhile, based on the producer’s direction of Lipofectamine 3000 (Invitrogen), miR‐615‐5p mimics/inhibitor and its control (miR‐NC/anti‐miR‐NC), empty vector (pcDNA3.1), overexpression vectors of hsa_circ_0004396 (pcDNA3.1‐hsa_circ_0004396), and PAK1 (pcDNA3.1‐PAK1) from Ribobio were transfected into 2 × 105 NSCLC cells in six‐well plates.
5
Regulation of HAX-1 by miR-223
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MiR-223 mimics, negative controls (miR-NCs), control siRNA (siRNA-NC) and HAX-1 siRNA were synthesized by RiboBio Co. Ltd. (China). The HAX-1 eukaryotic expression vector was generated by cloning the open reading frame of the HAX-1 gene into a pcDNA3.1 vector (Life Technologies, USA). For miR-223 overexpression, 50 pmol/ml miR-223 mimics were transfected using Lipofectamine 2000 (Invitrogen), according to the manufacturer’s instructions. For gain- and loss-of-function studies involving the HAX-1 gene, 2 μg/ml HAX-1 vector and 50 pmol/ml HAX-1 siRNA were transfected using Lipofectamine 2000.
6
Characterizing PCDHB17P in Breast Cancer
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According to the manufacturers' protocol, MDA-MB-231 and MCF-7 cells were severally transfected with each of the following by utilizing Lipotransfectamine 3000(Invitrogen, California, USA). PCDHB17Pspeci c short hairpin RNA (sh-PCDHB17P-1, sh-PCDHB17P-2) and negative control (sh-NC) were produced by RiboBio (Guangzhou, China), along with pcDNA3.1 vector containing PCDHB17P and empty vectors, were all from GenePharma (GenePharma, Shanghai, China). MiR-145-3p mimics, miR-145-3p inhibitors and their corresponding miR-NCs were synthesized by RiboBio (Guangzhou, China).
7
Transfection of circRbms1 and miR-742-3p
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For cell transfection, all oligonucleotides and vectors were synthesized from Ribobio (Guangzhou), including the circRbms1 small interference RNA (si-circRbms1) and its controls (si-NC), miR-742-3p mimic and inhibitor (miR-742-3p and anti-miR-742-3p) or their controls (miR-NC and anti-NC). The mimic and inhibitor of miR-742-3p were designed and synthesized by Sangon (Shanghai, China). The circRbms1 overexpression vector and FOXO1 overexpression vector was synthesized by subcloning a sequence of circRbms1 and FOXO1 into the pCD5-ciR vector and pcDNA3.1 vector, respectively. Lipofectamine 3000 (Invitrogen, Carlsbad, CA, USA) was used to transfect them into cells. The concentration of oligonucleotides was 50 nM, and the concentration of vectors was 4.0 µg. After transfection for 24 h, the cells were cultured under hypoxia for 24 h.
8
Targeting hsa_circ_0000735 in DTX-resistant PCa
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Small interference RNA targeting hsa_circ_0000735 (si-circ-1 and si-circ-2) and matching control (si-NC), as well as lentivirus-mediated sh-hsa_circ_0000735 (sh-circ) and corresponding control (sh-NC), were achieved from Genepharma (Shanghai, China). MiR-7 mimic and inhibitor (miR-7 and anti-miR-7), as well as their negative controls (miR-NC and anti-NC), were purchased from Ribobio (Guangzhou, China). The oligonucleotides were transfected into DTX-resistant PCa cells through using Lipofectamine 3000 reagent (Life Technologies, Carlsbad, CA, USA).
9
Investigating miR-497 and FGFR1 in Cells
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miR-NC and miR-497 were synthesized by RiboBio (China). Cells were seeded in 6-well plates and transfected with miR-NC and miR-497 using Lipofectamine 2000 (Invitrogen; Thermo Fisher Scientifc, Inc., USA) for 48 h at 37°C according to the manufacturer's protocol.
Short hairpin RNA (shRNA) was constructed to specifically target FGFR1 by shRNA design tools (http://rnaidesigner.thermofisher.com/rnaiexpress/ ). Using BLAST (http://blast.ncbi.nlm.nih.gov/Blast.cgi ), we verified that the designed shRNA targeted only the FGFR1. Sh-FGFR1 and sh-NC were synthesized by GenePharma (China).
A mammalian expression plasmid (pReceiver-M02-ERBB3) designed to specially express the full-length open reading frame of human FGFR1 without miR-497 responsive 3′-UTR was purchased from GeneCopoeia (USA). An empty plasmid served as a negative control. Overexpressed FGFR1 plasmid (vector-FGFR1) and control (vector-NC) were transfected into cells using Lipofectamine 2000 (Invitrogen; Thermo Fisher Scientifc, Inc.) for 48 h at 37°C according to the manufacturer's protocol.
Short hairpin RNA (shRNA) was constructed to specifically target FGFR1 by shRNA design tools (
A mammalian expression plasmid (pReceiver-M02-ERBB3) designed to specially express the full-length open reading frame of human FGFR1 without miR-497 responsive 3′-UTR was purchased from GeneCopoeia (USA). An empty plasmid served as a negative control. Overexpressed FGFR1 plasmid (vector-FGFR1) and control (vector-NC) were transfected into cells using Lipofectamine 2000 (Invitrogen; Thermo Fisher Scientifc, Inc.) for 48 h at 37°C according to the manufacturer's protocol.
10
Chordoma Cell Line Culture and Manipulation
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Chordoma cells (U-CH1 and JHC7) were purchased from American Type Culture Collection (ATCC; Manassas, VA, USA). U-CH1 cells were grown in a 4:1 mixture of IMDM-RPMI 1640 (ATCC) containing 10 % fetal bovine serum (10 %; Invitrogen, Carlsbad, CA, USA). JHC7 cells were grown in DMEM (ATCC) containing 10 % FBS. All the cells were cultured at 37℃ in humidified air with 5 % CO2.Short hairpin RNA (shRNA) targeting XIST (sh-XIST), miR-320d mimic and inhibitor (miR-320d and in-miR-320d), ARF6-overexpression plasmid (ARF6), and their controls (sh-NC, miR-NC, in-miR-NC, and pcDNA) were constructed by RiboBio (Guangzhou, China). The oligonucleotide and plasmid were introduced into chordoma cells (2 × 105 cells/well) by using Lipofectamine 3000 Reagent (Invitrogen).
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