The Starbasev3.0 (http://starbase.sysu.edu.cn/ ) and the TargetScanv7.2 (http://www.targetscan.org/vert_72/ ) predictive databases were used to predict the potential targets of miR-338-3p in humans. Following bioinformatics prediction and screening, potential binding sites were identified between miR-338-3p and NOVA1. Therefore, a dual-luciferase reporter assay was performed to verify that miR-338-3p directly binds to NOVA1. Briefly, wild-type (wt) and mutant (mut) 3′-UTR of NOVA1 were cloned into the pmirGLO luciferase reporter vector (Guangzhou RiboBio Co., Ltd.). Subsequently, the Y79 and WERI-Rb-1 cells were co-transfected with wt-NOVA1 or mut-NOVA1 and 100 nM agomir-338-3p (Guangzhou RiboBio Co., Ltd.) or 100 nM agomir-NC Shanghai GeneChem Co., Ltd. using Lipofectamine® 2000 (Invitrogen; Thermo Fisher Scientific, Inc.) at 37°C for 48 h. Finally, luciferase activity was determined using the dual-luciferase reporter assay system (Promega Corporation) and normalized to Renilla luciferase activity.
Agomir 338 3p
Manufactured by RiboBio
Sourced in China
Agomir-338-3p is a synthetic RNA molecule that mimics the mature form of microRNA-338-3p. microRNA-338-3p is a small, non-coding RNA that plays a regulatory role in gene expression.
Automatically generated - may contain errors
Lab products found in correlation
Lipofectamine 2000, by Thermo Fisher Scientific (2 mentions)
Pmirglo luciferase reporter vector, by RiboBio (1 mentions)
Dual luciferase reporter assay system, by Promega (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Mir 338 3p mimic, by RiboBio (1 mentions)
Agomir nc, by RiboBio (1 mentions)
2 protocols using agomir 338 3p
1
Verification of miR-338-3p Binding to NOVA1 3'UTR
2
Glioma Cell Transfection with miR-338-3p
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Human glioma cells (HS683, A172, and U251) and primary normal human astrocytes (NHA) were provided by KeyGEN Biotech (Nanjing, China). All cells were cultured in Dulbecco’s modified Eagles Medium containing 10% fetal bovine serum (Gibco, Carlsbad, CA) and then maintained at 37°C in an environment with 5% CO2.
RiboBio (Shanghai, China) designed and synthesized agomiR-338-3p, miR-338-3p mimic, agomiR negative control (agomiR-NC), and mimic-NC. THBS1 sequences were subcloned into a plasmid vector (pcDNA3.1) with an empty vector as the NC. Lipofectamine 2000 (Invitrogen, Carlsbad, CA, USA) was used to introduce the mimics and vectors into U251 and A172 cells. After 48 h of transfection, the efficiency of the transfections was determined by performing quantitative polymerase chain reaction (qPCR).
RiboBio (Shanghai, China) designed and synthesized agomiR-338-3p, miR-338-3p mimic, agomiR negative control (agomiR-NC), and mimic-NC. THBS1 sequences were subcloned into a plasmid vector (pcDNA3.1) with an empty vector as the NC. Lipofectamine 2000 (Invitrogen, Carlsbad, CA, USA) was used to introduce the mimics and vectors into U251 and A172 cells. After 48 h of transfection, the efficiency of the transfections was determined by performing quantitative polymerase chain reaction (qPCR).
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