Log In Sign up
The largest database of trusted experimental protocols

Pmirglo luciferase reporter vector

Manufactured by Promega
Visit Supplier
Sourced in United States, China

The PmirGLO Luciferase Reporter Vector is a plasmid designed for the study of miRNA regulation. It contains a firefly luciferase reporter gene that can be used to monitor miRNA activity by measuring luciferase expression levels.

Automatically generated - may contain errors

Lab products found in correlation

Dual luciferase reporter assay system, by Promega (91 mentions) Lipofectamine 2000, by Thermo Fisher Scientific (21 mentions) Lipofectamine 3000, by Thermo Fisher Scientific (15 mentions) Dual luciferase reporter assay kit, by Promega (14 mentions) Dual luciferase assay system, by Promega (11 mentions) Dual luciferase assay kit, by Promega (8 mentions) Dual luciferase reporter system, by Promega (6 mentions) Dual luciferase kit, by Promega (5 mentions) Luciferase reporter assay kit, by Promega (4 mentions) Lipofectamine 2000 reagent, by Thermo Fisher Scientific (4 mentions) Cignal finder reporter array, by Qiagen (3 mentions) Endotoxin free plasmid dna miniprep kit, by Tiangen Biotech (3 mentions) Pcdna3.1 vector, by Thermo Fisher Scientific (3 mentions) Mut express 2 fast mutagenesis kit v2, by Vazyme (3 mentions) Lysis buffer, by Promega (2 mentions) Veritas microplate luminometer td 20 20 luminometer, by Turner Designs (2 mentions) Luciferase reporter assay system, by Promega (2 mentions) Glomax 20 20 luminometer, by Promega (2 mentions) Dual glo luciferase assay system, by Promega (2 mentions) Luciferase reporter system, by Promega (2 mentions)

193 protocols using pmirglo luciferase reporter vector

1

Constructing Luciferase Reporter Vectors for CCND1 3'UTR Analysis

Check if the same lab product or an alternative is used in the 5 most similar protocols
For wildtype luciferase reporter vector construction, the 3' untranslated region of CCND1 (from 1900 nt to 2635 nt) was ampli ed by PCR and ligated into the pMir-GLO luciferase reporter vector (Promega, USA).
For mutant luciferase reporter vector construction, the mutant CCND1 3'UTR (replaced TGTTACA with AAAAACA) were cloned into the MCS region of pMir-GLO luciferase reporter vector with Sac I and Xba I.
The primers for wildtype and mutant CCND1 3'UTR were as follows, F: 5'-gggagctcCTGTCCCACTC CTACGATAC-3', R1 (wildtype): 5'-tctctagaTGTAACATCAAAGGCAGAAGG-3', R2 (Mutant): 5'-tctctagaTGTTTTTTCAAAGGCAGAAGGTTTGTGT-3'. The luciferase reporter assay was conducted as we previously described before [14] . Brie y, the BGC823 cells were seeded into 12-well-tissue plates 24h before transfection, and then co-transfected with 5 nM siRNA and 1mg plasmid using the Lipofectamine 2000 Reagent (Invitrogen), according to the manufacturer's instructions. After another 48h, cells were assayed using the Dual-Luciferase reporter assay system kit (GeneCopoeia, USA). All experiments were performed in triplicate and data were pooled from three independent experiments.
Li D., Wang J., Zhang M., Hu X., She J., Cai M., Yang L., Liu W., Hu Q., Qiu X., & Qin S. (2020). Global miRNA Expression Analysis Identifies miR-194, miR-100, miR-125b and miR-199a as Promising Biomarkers for Gastric Cancer and miR-194 Inhibits Gastric Cancer Cell Growth by Targeting CCND1.
+ Open protocol
+ Expand
2

Dual-Luciferase Assay for miRNA-3662 Targeting

Cited 2 times
Check if the same lab product or an alternative is used in the 5 most similar protocols
The pmirGLO luciferase reporter vectors (Promega, Madison, WI) were used to construct DNA fragments from the 3’-UTR of HBP1 (Transcript: HBP1–201 ENST00000222574.4) by Pmel and Xbal (Promega) digestion according to the manufacturer’s protocol. The sequences for DNA construction and mutagenesis are listed in Supplementary Table S4. Luciferase activity was measured as described previously 46 (link)–48 (link). Briefly, cells were plated at a density of 1× 104 per well into 96-well plates and then transiently co-transfected using Lipofectamine 3000 with luciferase gene reporter vectors (pmirGLO-HBP1–3’-UTR) and miR-3662 mimic (50 nmol/L) or inhibitor (100 nmol/L) according to the manufacturer’s protocol. After transient transfections for 48 hours, cells were washed twice with ice-cold PBS and were lysed in 1 x lysis buffer (Promega) for 15 min on a shaker. The luciferase activity was assessed with a Veritas Microplate Luminometer (TD-20/20 Luminometer, Turner Designs) using a Dual-Luciferase Assay System (Promega).
Yi B., Wang S., Wang X., Liu Z., Zhang C., Li M., Gao S., Wei S., Bae S., Stringer-Reasor E., Wang L, & Liu R. (2021). CRISPR interference and activation of the microRNA-3662-HBP1 axis control progression of triple-negative breast cancer. Oncogene, 41(2), 268-279.
+ Open protocol
+ Expand
3

Luciferase Assay for miR-188-5p Binding

Check if the same lab product or an alternative is used in the 5 most similar protocols
The wild-type (WT) DARS-AS1 (CAAGGGAU) and mutant (MT) DARS-AS1 (CCCCUUGU) fragments were cloned into the pmirGLO luciferase reporter vectors (Promega, Madison, WI, USA). Then, SiHa cells were co-transfected with WT-DARS-AS1 and miR-188-5p agomir or MT-DARS-AS1 and miR-188-5p agomir, respectively, using Lipofectamine 2000 for 48 h. In addition, WT-HMGB1 (AAGGGAU) and MT-HMGB1 (CCUUGGU) 3′-UTR fragments were cloned into the pmirGLO luciferase reporter vectors. Then, WT-HMGB1 or MT-HMGB1 plasmid was co-transfected with miR-188-5p agomir and agomir-NC respectively using Lipofectamine 2000 in SiHa cells for 48 h. Subsequently, Dual Luciferase Reporter Assay System (Promega, Madison, USA) was used to detect luciferase activity with renilla luciferase activity as endogenous control.
Zhu J, & Han S. (2020). DARS-AS1 Knockdown Inhibits the Growth of Cervical Cancer Cells via Downregulating HMGB1 via Sponging miR-188-5p. Technology in Cancer Research & Treatment, 19, 1533033820971669.
+ Open protocol
+ Expand
4

Luciferase reporter assay for circular RNA and target interactions

Check if the same lab product or an alternative is used in the 5 most similar protocols
The hsa_circ_0000069 (wild type) and its mutant sequences were inserted into pmirGLO luciferase reporter vectors (Promega, Madison, WI, USA). Then, wild type (WT)-hsa_circ_0000069 or mutant (MT)-hsa_circ_0000069 plasmid was co-transfected with miR-144 agomir and NC, respectively, using Lipofectamine 2000 in SW1990 cells for 48 h. The STIL (WT) and its mutant sequences were inserted into pmirGLO luciferase reporter vectors. Then, WT-STIL or MT-STIL plasmid was co-transfected with miR-144 agomir and NC, respectively, using Lipofectamine 2000 in SW1990 cells for 48 h. Later on, luciferase activities were measured by using the Dual-Luciferase Reporter Assay System (Promega, Madison, USA) with renilla luciferase activity as endogenous control.
Ye Z., Zhu Z., Xie J., Feng Z., Li Y., Xu X., Li W, & Chen W. (2020). Hsa_circ_0000069 Knockdown Inhibits Tumorigenesis and Exosomes with Downregulated hsa_circ_0000069 Suppress Malignant Transformation via Inhibition of STIL in Pancreatic Cancer. International Journal of Nanomedicine, 15, 9859-9873.
+ Open protocol
+ Expand
5

Luciferase Assay for lncRNA-miRNA Interaction

Cited 1 time
Check if the same lab product or an alternative is used in the 5 most similar protocols
The full length of lncRNA KCNMB2-AS1 was ligated into pmirGLO luciferase reporter vectors (Promega) to construct wild-type (Wt) pmirGLO-lncRNA. PmirGLO-lncRNA mutant (Mut) was also developed in which the binding sites of miR-3194-3p were mutated. The plasmids were synthesized by Invitrogen and then cotransfected with miR-3194-3p mimics or NC mimics into TE-1 and Eca109 cells using Lipofectamine 3000 (Invitrogen). After 48 h of transfection, the Dual-Luciferase Kit (Promega) was utilized to determine the luciferase activity [37 (link)].
Xu J., Liu X., Liu X, & Zhi Y. (2021). Long noncoding RNA KCNMB2-AS1 promotes the development of esophageal cancer by modulating the miR-3194-3p/PYGL axis. Bioengineered, 12(1), 6687-6702.
+ Open protocol
+ Expand
6

Dual-Luciferase Assay for miRNA-3662 Targeting

Cited 2 times
Check if the same lab product or an alternative is used in the 5 most similar protocols
The pmirGLO luciferase reporter vectors (Promega, Madison, WI) were used to construct DNA fragments from the 3’-UTR of HBP1 (Transcript: HBP1–201 ENST00000222574.4) by Pmel and Xbal (Promega) digestion according to the manufacturer’s protocol. The sequences for DNA construction and mutagenesis are listed in Supplementary Table S4. Luciferase activity was measured as described previously 46 (link)–48 (link). Briefly, cells were plated at a density of 1× 104 per well into 96-well plates and then transiently co-transfected using Lipofectamine 3000 with luciferase gene reporter vectors (pmirGLO-HBP1–3’-UTR) and miR-3662 mimic (50 nmol/L) or inhibitor (100 nmol/L) according to the manufacturer’s protocol. After transient transfections for 48 hours, cells were washed twice with ice-cold PBS and were lysed in 1 x lysis buffer (Promega) for 15 min on a shaker. The luciferase activity was assessed with a Veritas Microplate Luminometer (TD-20/20 Luminometer, Turner Designs) using a Dual-Luciferase Assay System (Promega).
Yi B., Wang S., Wang X., Liu Z., Zhang C., Li M., Gao S., Wei S., Bae S., Stringer-Reasor E., Wang L, & Liu R. (2021). CRISPR interference and activation of the microRNA-3662-HBP1 axis control progression of triple-negative breast cancer. Oncogene, 41(2), 268-279.
+ Open protocol
+ Expand
7

Luciferase Reporter Assays for Signaling Pathways

Check if the same lab product or an alternative is used in the 5 most similar protocols
The MLE-12/LPS cells were seeded in 96-well plates and transfected with the indicated luciferase reporter plasmids to detect the luciferase activities of Notch pathway, Wnt pathway, JAK/STAT3 pathway, NF-κB pathway, PI3K/AKT pathway, NRF2 pathway, Hedgehog pathway, MAPK/JNK pathway and MAPK/ERK pathway, respectively. For gene promoter analyses, cells were co-transfected with indicated transfection plasmids and the pGL3-basic reporter vectors (Promega, Madison, WI, USA) containing Ikbkb promoter, Usp5 promoter or SHH promoter. In addition, the Ikbkb fragment covering miR-182-5p wild-type or mutant binding sites was inserted into pmirGLO luciferase reporter vectors (Promega), and then co-transfected with miR-182-5p mimics or NC mimics into MLE-12/LPS cells and HEK-293T cells (ATCC; Manassas, VA, USA). The pmirGLO vectors which contained Usp5 fragment covering miR-23a-3p wild-type or mutant binding sites, were co-transfected with miR-23a-3p mimics or NC mimics into MLE-12/LPS and HEK-293T cells. At 48 h post transfection, all luciferase activities were examined with a luciferase reporter assay system (Promega).
Xiao K., He W., Guan W., Hou F., Yan P., Xu J., Zhou T., Liu Y, & Xie L. (2020). Mesenchymal stem cells reverse EMT process through blocking the activation of NF-κB and Hedgehog pathways in LPS-induced acute lung injury. Cell Death & Disease, 11(10), 863.
+ Open protocol
+ Expand
8

Luciferase Assay for miR-9 and Target Genes

Check if the same lab product or an alternative is used in the 5 most similar protocols
The luciferase reporter assay was used to determine the association between miR-9 and MIR22HG, and CPEB3. Wild-type (WT) and mutant (MUT) MIR22HG or CPEB3 3′-untranslated region sequences were inserted into the pmirGLO luciferase reporter vectors (Promega Corporation) to construct a luciferase reporter vector. In total, 50 ng WT and MUT luciferase vectors or 50 nM miR-9 mimic were transfected into U87 or U251 cells using Lipofectamine® 2000 (Invitrogen; Thermo Fisher Scientific, Inc.) for 20 min at room temperature. After 48 h, the luciferase activities were detected using a dual-luciferase reporter assay kit (Promega Corporation). The firefly luciferase was normalized using Renilla luciferase as the internal control.
He Y., Nan H., Yan L., Ma T., Man M., Tian B., Guo S, & Zhang X. (2020). Long non-coding RNA MIR22HG inhibits glioma progression by downregulating microRNA-9/CPEB3. Oncology Letters, 21(2), 157.
+ Open protocol
+ Expand
9

Signaling Pathways Regulated by RP1-59D14.5

Check if the same lab product or an alternative is used in the 5 most similar protocols
Cells were planted into 96-well plates and transfected with the various luciferase reporter vectors to detect the luciferase activities of Myc pathway, PI3K/AKT pathway, NF-κB pathway, Oct4 pathway, Nanog pathway, Notch pathway, Hippo pathway, Wnt pathway and Hedgehog pathway, respectively. The Cignal Finder Reporter Array (336841, QIAGEN, Germany) was used to identify the signaling pathways regulated by RP1-59D14.5. RP1-59D14.5 cDNA was generated by PCR and inserted into the firefly luciferase plasmid. The luciferase activity was measured by Dual-luciferase reporter assay system (Promega). Firefly luciferase activity was normalized to that of Renilla luciferase.
In addition, RP1-59D14.5 or LATS1/2 fragment covering miR-147a wild-type (Wt) or mutant (Mut)-type binding sites was inserted into pmirGLO luciferase reporter vectors (Promega), and then co-transfected with miR-147a mimics or NC mimics into cells. After 48 h transfection, all luciferase activities were examined by use of luciferase reporter assay system (Promega). The experiment was conducted at least three times.
Zhong B., Zhao Z, & Jiang X. (2022). RP1-59D14.5 triggers autophagy and represses tumorigenesis and progression of prostate cancer via activation of the Hippo signaling pathway. Cell Death & Disease, 13(5), 458.
+ Open protocol
+ Expand
10

Identifying miRNA-144-5p Binding to ITGA3

Check if the same lab product or an alternative is used in the 5 most similar protocols
The pmirGLO luciferase reporter vectors (Promega, USA) inserted with the ITGA3 wild-type (ITGA3-WT) and mutant (ITGA3-MUT) 3′-UTR were constructed to identify the binding relationship of miRNA-144-5p to the 3′-UTR of ITGA3. TC cell line TPC-1 was seeded in triplicate in 96-well plates (3 × 105 cells/well), and 100 nM miR-mimics/miR-NC and ITGA3-WT/ITGA3-MUT plasmids were cotransfected into the cells. Cultured for 48 h, the dual-luciferase reporter system (Promega, USA) was then used for measuring the luciferase activity.
Zhang J., Zhong Y., Sang Y, & Ren G. (2021). miRNA-144-5p/ITGA3 Suppressed the Tumor-Promoting Behaviors of Thyroid Cancer Cells by Downregulating ITGA3. Computational and Mathematical Methods in Medicine, 2021, 9181941.
+ Open protocol
+ Expand

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

Sign up now

Revolutionizing how scientists
search and build protocols!