The genes HPV6 E6, HPV6 E7, HPV11 E6, HPV11 E7, HPV16 E6 and HPV16 E7 were amplified by PCR containing restriction enzyme sites. These PCR products were digested with restriction enzymes, Bgl II and Xho I (New England Biolab), and ligated to pMSCV-puro vector (Clontech). Plasmids expressing EBV LMP-1, (i.e. pMSCV-neo-LMP-1) were prepared as previously described [15 (link)]. pNF-κB-TA-luc, pp53-TA-luc, pβ-gal-basic (Clontech) and pcDNA3.1/Zeo (+) (Invitrogen) were also used for assays.
Pnf κb ta luc
Manufactured by Takara Bio
The PNF-κB-TA-luc is a reporter construct designed to measure the activity of the NF-κB transcription factor. It consists of a luciferase gene under the control of tandem NF-κB response elements and a TATA-like promoter sequence.
Automatically generated - may contain errors
Lab products found in correlation
Pp53 ta luc, by Takara Bio (2 mentions)
Pcdna3.1 zeo, by Thermo Fisher Scientific (1 mentions)
Pmscv puro vector, by Takara Bio (1 mentions)
Bglii, by New England Biolabs (1 mentions)
Xhoi, by New England Biolabs (1 mentions)
Pmscv puro retroviral vector, by Takara Bio (1 mentions)
Pcre luc, by Takara Bio (1 mentions)
Phse luc, by Takara Bio (1 mentions)
Pe2f ta luc, by Takara Bio (1 mentions)
Pap1 luc, by Takara Bio (1 mentions)
Psre luc, by Takara Bio (1 mentions)
Pstat3 ta luc, by Takara Bio (1 mentions)
Pisre ta luc, by Takara Bio (1 mentions)
Fopflash, by Addgene (1 mentions)
Topflash, by Addgene (1 mentions)
Pmirglo vector, by Promega (1 mentions)
Pta luc, by Takara Bio (1 mentions)
Pgl3 promoter, by Promega (1 mentions)
Luciferase assay system, by Promega (1 mentions)
3 protocols using pnf κb ta luc
1
Cloning of HPV Oncogenes into Retroviral Vector
2
Cloning and Profiling of miR-424-5p
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The human miR-424-5p gene was PCR amplified from genomic DNA and cloned into a pMSCV-puro retroviral vector (Clontech, Japan). Pathway Profiling System, including pAP1-luc, pCRE-luc, pE2F-TA-luc, pERE-luc, pGRE-luc, pHSE-luc, pISRE-TA-luc, pMYC-luc, pNF-κB-TA-luc, pp53-TA-luc, pSRE-luc, and pSTAT3-TA-luc, was purchased from Clontech (PT3286-1). HOPFlash (Catalog [Cat.] 83467), HIPFlash (Cat. 83466), TOPFlash (Cat. 12456), and FOPFlash (Cat. 12457) were purchased from Addgene. The 3′ UTRs of WWC1, SAV1, and LATS2 were PCR amplified from genomic DNA and cloned into pmirGLO vectors (Promega, USA), and the list of primers used in cloning reactions was provided in Table S3 . Anti-miR-424-5p was synthesized and purified by GENECHEM (China). The primers of the mutant plasmid for WWC1, SAV1, and LAST2 were synthesized and purified by HDbio (Guangzhou, China). Transfection of plasmids was performed as previously described.46 (link)
3
Evaluating DSF's Impact on RANKL-Induced Signaling
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To examine the effects of DSF on RANKL-induced NF-κB and NFATc1 activation, RAW264.7 cells were transiently transfected with luciferase reporter genes (p-NF-κB-TA-Luc, p-NFAT-TA-Luc, pTA-Luc (Clontech, Mercury Pathway Profiling System) or pGL3-promoter (Promega)) as previously described [26 (link), 27 (link)]. Cells were seeded (1.5×105/well) into 48-well plates. After attachment, cells were pretreated with DSF for 1h and then stimulated with RANKL (100 ng/ml) for an indicated period of time. Luciferase activities were measured in cell lysates using the Promega Luciferase Assay System according the manufacturer’s instructions.
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