SGC-7901 and BGC-823 and their respective resistance cells were purchased from Shanghai Institute of Cell Biology (Shanghai, China). All cell lines were cultured in RPMI 1640 (GIBCO, Rockville, MD, USA) medium supplemented with 10% fetal bovine serum (FBS), 100 U/ml penicillin and 100 mg/ml streptomycin in humidified air at 37°C with 5% CO2. miR-223 mimic or inhibitor or siRNA-FBXW7 and their negative controls were obtained from GenePharma (Shanghai, China). The open reading frame of FBXW7 that was generated by PCR was then inserted into the pcDNA 3.1 expression vector, which was named pcDNA-FBXW7. The recombinant vector was confirmed by the digestion analysis of restriction endonuclease and DNA sequencing. For ectopic expression of miR-223, miR-223 mimic or miR-NC vectors were purchased from GenePharm. The transfection was performed using Lipofectamine 2000 (Invitrogen, Carlsbad, CA, USA) according to the instructions.
Mir 223 mimic
Manufactured by GenePharma
Sourced in China
MiR-223 mimic is a synthetic RNA molecule designed to mimic the function of the naturally occurring microRNA, miR-223. MicroRNAs are small, non-coding RNA molecules that play a crucial role in gene expression regulation. The MiR-223 mimic can be used in research and experimental settings to study the biological functions and effects of miR-223 in various cellular and molecular processes.
Automatically generated - may contain errors
Lab products found in correlation
Mir 223 inhibitor, by GenePharma (3 mentions)
Rpmi 1640, by Thermo Fisher Scientific (1 mentions)
Lipofectamine 2000, by Thermo Fisher Scientific (1 mentions)
Bgc 823, by Shanghai Institute of Biochemistry and Cell Biology (1 mentions)
Sgc 7901, by Shanghai Institute of Biochemistry and Cell Biology (1 mentions)
Lipofectamine 2000 reagent, by Thermo Fisher Scientific (1 mentions)
Pgl3 promoter plasmid, by Promega (1 mentions)
Prl tk vector, by Promega (1 mentions)
Dual luciferase assay kit, by Promega (1 mentions)
Kod plus mutagenesis kit, by Toyobo (1 mentions)
Dual luciferase reporter assay, by Promega (1 mentions)
Mimic nc, by GenePharma (1 mentions)
Interferin, by Polyplus Transfection (1 mentions)
Lipofectamine 3000, by Thermo Fisher Scientific (1 mentions)
Inhibitor nc, by GenePharma (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (1 mentions)
Lipofectamine 2000 transfection reagent, by Thermo Fisher Scientific (1 mentions)
Hct116, by American Type Culture Collection (1 mentions)
Sw480, by American Type Culture Collection (1 mentions)
9 protocols using mir 223 mimic
1
Regulation of Gastric Cancer Cell Proliferation
2
miRNA mimic and inhibitor transfection
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The miR-223 mimic, miR-223 inhibitor or control mimic, control inhibitor used in this study was purchased from GenePharma (Shanghai, China). OSCC cells were transfected with miRNA mimic or miRNA inhibitor using the Lipofectamine 2000 reagent (Invitrogen) following the manufacturer’s instructions. Then, the cells were incubated at 37 C under 5% CO2 for 48 h.
3
Luciferase Reporter Assay for miR-223 Targets
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The 3′-UTR fragments of CDK2 and CUL1 containing the predicted miR-223-binding sites were amplified by PCR using the primers (Supplementary Table S3 ) and cloned into the pGL3-Promoter plasmid (Promega) downstream of the promote, with the resulting fragments named pGL3-WT-CDK2 and pGL3-WT-CUL1. Site-directed mutant vectors (base substitutions) were generated by using the KOD Plus Mutagenesis Kit (TOYOBO, Osaka, Japan) following the manufacturer’s instructions and named pGL3-MUT-CDK2 and pGL3-MUT-CUL1. For the luciferase reporter assays, Hepa1-6 cells with 70% confluence were grown in 48-well plates and transfected with 100 ng of WT or mutant vectors (pGL3-MUT-CDK2, pGL3-MUT-CUL1, pGL3-MUT-CDK2 and pGL3-MUT-CUL1) and 100 nM miR-223 mimic (Genepharma) or negative control using Lipofectamine 3000. As an internal control, 10 ng pRL-TK vector (Promega) was co-transfected for normalization of the transfection efficiency. Cells were lysed and the luciferase reporter activity was assayed 48 h after transfection using the dual luciferase assay kit (Promega) following the manufacturer’s protocol.
4
Validating miRNA-mRNA Targeting in NLRP3
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
TargetScan (http://www.targetscan.org/vert_61/ ), online bioinformatics software, concluded that NLRP3 is a downstream target gene of miR-223. The luciferase-3′-UTR reporter constructs were generated by introducing the 3′-UTRs of NLRP3 carrying the putative miR-223 binding site into a luciferase reporter vector. A plasmid vector containing the wild-type loci of miR-223 binding NLRP3 (NLRP3 wild) and a plasmid vector containing the mutant loci of miR-223 binding NLRP3 (NLRP3 mut) were designed and constructed. NLRP3-3′-UTR primer sequences synthesized by Shanghai GenePharma Co., Ltd. (Shanghai, China) are as follows (Table 2 ). After the sequence was verified by sequencing, HEK293 cells (ATCC) were co-transfected with 80 ng NLRP3 wild plasmid or NLRP3 mut plasmid, along with 40 ng dual fluorescence plasmid carrying firefly and Renilla luciferase reporters, and the miR-223 mimic, miR-223 inhibitor, or the control (Shanghai GenePharma Co., Ltd., Shanghai, China). After HEK293 cells were incubated at 37℃ for 24 h, proteins were extracted and analyzed by the dual-Luciferase reporter assay (Promega, Madison, WI, USA).
![]()
NLRP3-3′-UTR primer sequences.
| Primers | Sequences |
|---|---|
| Forward | 5′-ACCTCAACAGTCGCTACACG-3′ |
| Reverse | 5′-TAGACTCCTTGGCGTCCTGA-3′ |
5
Overexpressing miR-223 in 3T3L1 Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
To induce miR-223 overexpression, synthetic 100 nM miRNA mimics were transfected into 3T3L1 cells (2×105/well) using 3 µl INTERFERin® small INTERFERing RNA transfection reagent (Polyplus-transfection SA, Illkirch, France) on a 24-well plate at 37°C for 24 h. The miR-223 mimic and NC mimic (Shanghai GenePharma Co., Ltd., Shanghai, China) sequences were as follows: miR-223 mimic forward, 5′-UGUCAGUUUGUCAAAUACCCA-3′ and reverse, 5′-GGGUAUUUGACAAACUGACAUU-3′; NC mimics forward, 5′-UUCUCCGAACGUGUCACGUTT-3′ and reverse, 5′-ACGUGACACGUUCGGAGAATT-3′.
6
Modulating miR-223 and FOXO1 in Leukemia Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The miR-223 mimic, negative control (NC)-mimic, miR-223 inhibitor and NC-inhibitor were bought from GenePharma (Shanghai, China). FOXO1 siRNA-1 (si-FOXO1-1), si-FOXO1-1 and si-FOXO1 NC (si-NC) were obtained from RIBO Bio (Guangzhou, China). CCRF-CEM and NALM-6 cells grown to 80% confluence were transfected or co-transfected with these above agents using Lipofectamine 3000 (Invitrogen). The CCRF-CEM and NALM-6 cells were randomly divided into NC-mimic group (transfected with NC-mimic), miR-223 mimic group (transfected with miR-223 mimic), NC-inhibitor group (transfected with NC-inhibitor) and miR-223 inhibitor group (transfected with miR-223 inhibitor). In addition, the NALM-6 cells were randomly divided into si-NC group (transfected with si-NC), si-FOXO1-1 group (transfected with si-FOXO1-1), si-FOXO1-2 group (transfected with si-FOXO1-2), si-NC + NC-inhibitor group (co-transfected with si-NC and NC-inhibitor), si-FOXO1 + NC-inhibitor group (co-transfected with si-FOXO1 and NC-inhibitor) and si-FOXO1 + miR-223 inhibitor group (co-transfected with si-FOXO1 and miR-223 inhibitor). Cells without transfection were considered as the BLANK (BLANK group). All cells were cultured in 37°C incubator for 48 h.
7
Colorectal Cancer Cell Culture and Transfection
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The human CRC SW480, LoVo, DLD-1, SW620, HCT-116, and normal colonic epithelial cell line (NCM460) were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA). The cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM, Invitrogen, Carlsbad, CA, USA) containing with 10% dialyzed fetal bovine serum (FBS, Gibco Life Technologies), and with 1% penicillin and streptomycin at 37°C in the presence of 5% carbon dioxide.
The miR-223 mimic and corresponding mimic negative control (miR-NC) were designed by GenePharma (Shanghai, China). The pcDNA (Vector) and pcDNA-LPP overexpression (LPP-OE) plasmids were obtained from GenePharma (Shanghai, China). The target cells were transfected with the mimic, plasmids, and miR-NC using the Lipofectamine 2000 transfection reagent (Invitrogen) according to the manufacturer’s instruction.
The miR-223 mimic and corresponding mimic negative control (miR-NC) were designed by GenePharma (Shanghai, China). The pcDNA (Vector) and pcDNA-LPP overexpression (LPP-OE) plasmids were obtained from GenePharma (Shanghai, China). The target cells were transfected with the mimic, plasmids, and miR-NC using the Lipofectamine 2000 transfection reagent (Invitrogen) according to the manufacturer’s instruction.
8
miRNA Mimics and Inhibitors Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The miR-223 mimics, negative control (NC), miR-223 inhibitors, miRNA inhibitor NC, siRNA against chicken IGF2, ZEB1 and MYOD were all purchased from GenePharma (GenePharma, Shanghai, China).
9
MiR-223 Inhibitor Transfection Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The cells were seeded in six-well plates and transfected with miR-223 inhibitor or the nonspecific control or miR-223 mimics (GenePharma, Shanghai, China) using DharmaFect Transfection Reagent (Dharmacon, Lafayette, CO, USA) following the manufacturer’s protocol.39 (link) The miR-223 inhibitor was: 5′-UGG GGU AUU UGA CAA ACU GAC A-3′. The cells were subjected to further analysis as described in the Results section.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!