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Pgl3.0 vector

Manufactured by Promega
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Sourced in United States, China

The PGL3.0 vector is a plasmid vector used in molecular biology and genetic engineering applications. It serves as a backbone for cloning and expressing target genes or sequences. The core function of the PGL3.0 vector is to provide a stable and versatile platform for DNA manipulation and gene expression studies.

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Lab products found in correlation

Dual luciferase reporter assay kit, by Promega (2 mentions) Q5 site directed mutagenesis kit, by New England Biolabs (2 mentions) Modulus 2 microplate multimode reader, by Promega (1 mentions) Lipofectamine 2000, by Thermo Fisher Scientific (1 mentions) Fast mutagenesis system, by Transgene (1 mentions) Pmir report luciferase vector, by Thermo Fisher Scientific (1 mentions) Nf κb luciferase reporter plasmid, by Promega (1 mentions) Prl tk, by Promega (1 mentions) One step cloning kit, by Vazyme (1 mentions)

Close spelling variants of this product

PGL3 vector (739 citations) PGL3.0-basic vector (20 citations) PGL3.0 luciferase reporter vector (3 citations) PGL3.0-control vectors (2 citations)

6 protocols using pgl3.0 vector

1

Characterization of Mouse Wnt10b Promoter

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A plasmid containing 2000 to +216 bp DNA sequence of mouse
Wnt10b promoter was constructed by inserting a PCR
product of mouse genomic DNA into the pGL3.0 vector (Promega, Madison, WI).
mwnt10b-luc (a plasmid containing 705 to +216 bp DNA
sequence of mouse Wnt10b promoter constructed by inserting a PCR product of
mouse genomic DNA into the pGL3.0 vector) was a gift from Dr. D. J. Klemm,
University of Colorado, Denver. Other shortened constructs were subcloned
based on the construct of mwnt10b-luc (2000 bp to +216 bp).
The amplification template for all mutations is the
mwnt10b-luc (705 to +216 bp). The NFAT binding site
(5′-AGGAAAA-3′) at 282 to 276 bp was changed to
5′-AGcttAA-3′ using the Q5 Site-Directed Mutagenesis Kit (New
England BioLabs). Similarly, the SMAD binding site
(5′-GTCTAGA-3′) at 341 to 335 bp was mutated to
5′-catagcg-3′. The primers used are provided in Table S1.
Tyagi A.M., Yu M., Darby T.M., Vaccaro C., Li J.Y., Owens J.A., Hsu E., Adams J., Weitzmann M.N., Jones R.M, & Pacifici R. (2018). The Microbial Metabolite Butyrate Stimulates Bone Formation via T Regulatory Cell-Mediated Regulation of WNT10B Expression. Immunity, 49(6), 1116-1131.e7.
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2

Characterization of Mouse Wnt10b Promoter

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A plasmid containing 2000 to +216 bp DNA sequence of mouse
Wnt10b promoter was constructed by inserting a PCR
product of mouse genomic DNA into the pGL3.0 vector (Promega, Madison, WI).
mwnt10b-luc (a plasmid containing 705 to +216 bp DNA
sequence of mouse Wnt10b promoter constructed by inserting a PCR product of
mouse genomic DNA into the pGL3.0 vector) was a gift from Dr. D. J. Klemm,
University of Colorado, Denver. Other shortened constructs were subcloned
based on the construct of mwnt10b-luc (2000 bp to +216 bp).
The amplification template for all mutations is the
mwnt10b-luc (705 to +216 bp). The NFAT binding site
(5′-AGGAAAA-3′) at 282 to 276 bp was changed to
5′-AGcttAA-3′ using the Q5 Site-Directed Mutagenesis Kit (New
England BioLabs). Similarly, the SMAD binding site
(5′-GTCTAGA-3′) at 341 to 335 bp was mutated to
5′-catagcg-3′. The primers used are provided in Table S1.
Tyagi A.M., Yu M., Darby T.M., Vaccaro C., Li J.Y., Owens J.A., Hsu E., Adams J., Weitzmann M.N., Jones R.M, & Pacifici R. (2018). The Microbial Metabolite Butyrate Stimulates Bone Formation via T Regulatory Cell-Mediated Regulation of WNT10B Expression. Immunity, 49(6), 1116-1131.e7.
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3

Investigating Transcriptional Regulation in Embryonic Development

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The Shh, Fgfr2, and Fgf8 promoter fragment (0 to −2000 bp) was cloned from mouse genomic DNA and inserted into the pGL3.0 vector (E1751, Promega). Isl1 was amplified using the primers containing the NheI–XhoI restriction sites and inserted into the pcDNA3.1 vector. An empty luciferase reporter vector was used as a control. The primer sequence is in Supplementary Table S3.
The human embryonic kidney 293FT cells were maintained in Dulbecco’s modified Eagle’s medium supplemented with 10% fetal bovine serum and 1% penicillin–streptomycin at 37 °C with 5% CO2. Cells were transfected with Isl1 expression vector, Fgf8, Fgfr2, Shh luciferase reporter vector, and pTK-Ranilla vector using Lipofectamine 2000 (Invitrogen) according to the manufacturer’s instruction. Twenty four hours post transfection, luciferase activity was measured by using Dual-Luciferase Reporter Assay Kit (E1910; Promega) on a Modulus II Microplate Multimode Reader (Turner Biosystems, Sunnyvale, CA, USA). The values were normalized against Renilla luciferase activity. At least four independent experiments were performed.
Su T., Liu H., Zhang D., Xu G., Liu J., Evans S.M., Pan J, & Cui S. (2019). LIM homeodomain transcription factor Isl1 affects urethral epithelium differentiation and apoptosis via Shh. Cell Death & Disease, 10(10), 713.
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4

Cloning and Characterization of acsl6 Promoter

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Genomic DNA of muscle tissue was extracted from common carp using the standard phenol-chloroform extraction method and was used as a template for cloning the candidate promoter. The 1790-bp upstream sequence of the acsl6 gene was obtained from the genomic sequencing data of common carp. For identifying the core promoter region of acsl6 gene, four promoter fragments (SD1–SD4) were amplified using Transfer-PCR (TPCR) [48 (link)] with one of the specific forward primers (SD1-F, SD2-F, SD3-F, and SD4-F; Table 1) and a common reverse primer (SD-R; Table 1) and inserted into pGL3.0 vector (Promega, USA). The upstream sequence in the four insert fragments including SD4, SD3, SD2, and SD1 was of −1789 bp, −1231 bp, −758 bp, and −198 bp lengths to the putative transcription start site (TSS +1), respectively (Figure 4). These sequences were checked by sequencing in Shanghai Sangon Biotech Co., Ltd. (Shanghai, China).
Xie D., He Z., Dong Y., Gong Z., Nie G, & Li Y. (2020). Molecular Cloning, Characterization, and Expression Regulation of Acyl-CoA Synthetase 6 Gene and Promoter in Common Carp Cyprinus carpio. International Journal of Molecular Sciences, 21(13), 4736.
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5

Cloning and Mutagenesis of CD34 3'UTR

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The 3′ UTR of human CD34 gene was amplified by PCR using the CD34 3′ UTR primers, and then was mutated by Fast Mutagenesis System (Transgen Biotech Corporation, Beijing, China) according to the instructions. To construct luciferase reporter plasmids, the 3′ UTR and the mutant 3′ UTR of CD34 was respectively inserted into the pMIR-REPORTTM Luciferase Vector (Ambion, Carlsbad, CA). To construct pGL3-luciferase reporter plasmids, the 5′ fragments of hsa-mir-665 were inserted into pGL3.0 Vector (Promega Beijing, China) according to the manufacturer’s protocol.
Fan J., Li H., Nie X., Yin Z., Zhao Y., Zhang X., Yuan S., Li Y., Chen C, & Wang D.W. (2018). MiR-665 aggravates heart failure via suppressing CD34-mediated coronary microvessel angiogenesis. Aging (Albany NY), 10(9), 2459-2479.
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6

Transcriptional Regulation of Adhesion Molecules

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By using the One Step Cloning Kit (C112-02; Vazyme Biotech, Nanjing, China), NF-κB luciferase reporter plasmid, purchased from Tsingke Biotechnology (Tsingke, China), promoters of intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), E-selectin, and P-selectin, were assembled into pGL3.0 vector (Promega, Madison, Wis). The aforementioned primers are listed in Table E3. HEK293 cells were transfected with an internal control plasmid (pRL-TK; Promega) and plasmids containing other promoter regions simultaneously. Luciferase activity were detected with the Dual Luciferase Reporter Assay Kit (Promega).
Rao C., Liu B., Huang D., Chen R., Huang K., Li F, & Dong N. (2021). Nucleophosmin contributes to vascular inflammation and endothelial dysfunction in atherosclerosis progression. The Journal of thoracic and cardiovascular surgery, 161(5).
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