Fuw teto hoct4

Manufactured by Addgene

The FUW-tetO-hOct4 is a plasmid vector used for the inducible expression of the human Oct4 gene. It contains a tetracycline-responsive promoter element (tetO) that can drive the expression of the Oct4 coding sequence upon the addition of a tetracycline-like inducer.

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Lab products found in correlation

Fudeltagw rtta, by Addgene (2 mentions) Fibronectin, by Santa Cruz Biotechnology (1 mentions) Latrunculin a, by Merck Group (1 mentions) D luciferin, by Merck Group (1 mentions) Doxycycline hyclate, by Merck Group (1 mentions) 8xgtiic lux, by Addgene (1 mentions) Pap1 luc, by Takara Bio (1 mentions) Prl tk, by Promega (1 mentions)

2 protocols using fuw teto hoct4

Inducible Expression of YAP Constructs

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Doxycycline hyclate, latrunculin A, D-luciferin, DAPT and DBZ were from Sigma; fibronectin was from Santa Cruz Biotechnology; cytochalasin D was from Calbiochem; CPRG was from Roche. For the inducible expression of YAP constructs, cDNA for human YAP1 5SA (LATS-mutant sites)21 (link) and human YAP1 5SA/S94A (TEAD-binding mutant)34 (link) were made insensitive to YAP siRNA #1 and subcloned, together with an HA-tag, in FUW-tetO-MCS, obtained by substituting the Oct4 sequence in FUW-tetO-hOct4 (Addgene #20726)65 (link) with a new multiple cloning site (MCS). This generated FUW-tetO-HA-YAP5SA and FUW-tetO-HA-YAP5SA/S94A were used throughout this study. FUW-tetO-MCS (empty vector) or FUW-tetO-RFP plasmids, obtained by subcloning the RFP coding sequence from the pTomo vector (Addgene #26291), were used as controls. FUdeltaGW-rtTA was from Addgene (#19780)66 (link). FUW-tetO-N1ICD-Myc and FUW-tetO-GFP-DNMAML1 were obtained by subloning in the FUW-tetO-MCS the corresponding coding sequences, respectively, from pcDNA3 N1ICD-Myc67 (link) and MigRI-DNMAML1-GFP38 (link). All constructs were confirmed by sequencing. The 8xGTIIC-lux (Addgene #34615) was previously described22 (link). The 3D.A-Lux was gently provided by Guido Posern68 (link). Primers for RT–PCR are listed in Supplementary Data 3.

Totaro A., Castellan M., Battilana G., Zanconato F., Azzolin L., Giulitti S., Cordenonsi M, & Piccolo S. (2017). YAP/TAZ link cell mechanics to Notch signalling to control epidermal stem cell fate. Nature Communications, 8, 15206.

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Regulatory Transcription Factor Plasmid Library

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pCS2-FLAG-mTAZ, pBABE-hygro-FLAG-mTAZ wt and S89A, pCDNA-FLAG-YAP, FU-tet-o-EGFP-ires-PURO, 8xGTIIC-luc were previously described7 (link),16 (link),33 (link),38 (link). TAZS51A and YAPS94A were generated by PCR-mediated mutagenesis and cloned into pBABE retroviral vectors. pCMV6-FLAG-MYC-TEAD1 was from Origene. FU-tet-o-hc-myc (#19775, Ref 34 (link)) and FUdeltaGW-rtTA (#19780, Ref 34 (link)) were purchased from Addgene. pBABE-puro-JUN~FOSL1-FLAG was a kind gift of L. Bakiri26 (link). Doxycycline-inducible JUN-DN lentiviral construct was obtained by substituting the Oct4 sequence in FUW-tetO-hOCT4 (Addgene #20726) with JUN-DN cDNA from pMIEG3-JunDN (Addgene #40350). pAP1-luc was from Clontech and pRL-TK from Promega. CTGF and ANKRD1 luciferase reporters were generated by amplifying CTGF (hg19, chr6:132272417-132273191)/ANKRD1(chr10:92680870-92681128) promoters by PCR from genomic DNA and cloning into pGL3 basic; for AP-1 mutants, point mutations were introduced by PCR in AP-1 binding sites; for TEAD mutants, a single deletion comprising the three TEAD binding sites was introduced in CTGF reporter, whereas point mutations were generated in ANKRD1 promoter. All constructs were confirmed by sequencing.

Zanconato F., Forcato M., Battilana G., Azzolin L., Quaranta E., Bodega B., Rosato A., Bicciato S., Cordenonsi M, & Piccolo S. (2015). Genome-wide association between YAP/TAZ/TEAD and AP-1 at enhancers drives oncogenic growth. Nature cell biology, 17(9), 1218-1227.

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