Prk5 ha ub k0

Manufactured by Addgene

Sourced in United States

PRK5-HA-Ub-K0 is a plasmid that contains a hemagglutinin (HA) tag fused to the ubiquitin (Ub) coding sequence. The plasmid does not have any lysine residues in the Ub sequence.

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Lab products found in correlation

Prk5 ha ub, by Addgene (1 mentions) Prk5 ha ub k48, by Addgene (1 mentions) Prk5 ha ub k63, by Addgene (1 mentions) Lipofectamine rnaimax reagent, by Thermo Fisher Scientific (1 mentions) Ltx reagent, by Thermo Fisher Scientific (1 mentions) Plus reagent, by Thermo Fisher Scientific (1 mentions) Prk5 ha ubiquitin wt, by Addgene (1 mentions) Gibson assembly master mix, by New England Biolabs (1 mentions) Ha ubiquitin, by Addgene (1 mentions) Pci ha nedd4, by Addgene (1 mentions) Plenti puro ha ubiquitin, by Addgene (1 mentions) Pegfp c1, by Takara Bio (1 mentions)

Spelling variants of this product

HA-Ub (16 citations) PRK5-HA-Ub (5 citations) HA-Ub-K0 (3 citations)

2 protocols using prk5 ha ub k0

Plasmid and siRNA transfection in HepG2 cells

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Flag-tagged PAQR3 plasmids and HA-tagged DDB2 plasmids were obtained from Vigene Bioscences (Jinan, China). pRK5-HA-Ub (RRID: Addgene_17608), pRK5-HA-Ub-K0 (RRID: Addgene_17603), pRK5-HA-Ub-K48 (RRID: Addgene_17605), and pRK5-HA-Ub-K63 (RRID: Addgene_17606) were obtained from Addgene (Watertown, MA, USA). Plasmids were handled according to the manufacturer's instructions with LTX reagent and PLUS reagent (ThermoFisher Scientific, Rockford, IL, USA). HepG2 cells were cultured 24 h prior to transfection, and then 2 μg of plasmid was transfected into cells with incubation for 48 h for further treatment.
The siRNA oligonucleotides of PAQR3 and DDB2 were purchased from GenePharma (Shuzhou, China), and the most effective sequences are listed in Supporting Information Table S3. Transfection of siRNA was performed according to the manufacturer's instructions with Lipofectamine RNAiMAX reagent (ThermoFisher Scientific). Transfected cells were incubated for 48 h for further treatment.

Xiao H., Sun X., Lin Z., Yang Y., Zhang M., Xu Z., Liu P., Liu Z, & Huang H. (2022). Gentiopicroside targets PAQR3 to activate the PI3K/AKT signaling pathway and ameliorate disordered glucose and lipid metabolism. Acta Pharmaceutica Sinica. B, 12(6), 2887-2904.

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Generation of TRPV4 and Ubiquitin Constructs

Cited 1 time

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TRPV4-FLAG in pcDNA3.1 was previously described (103 (link)). TRPV4-V5-FLAG was generated using Gibson Assembly Master Mix (New England Biolabs), by inserting the V5 tag sequence into the middle position of the first TRPV4 extracellular loop. TRPV4-V5-EGFP was generated by replacing the FLAG tag of the TRPV4-V5-FLAG construct with EGFP cloned from pEGFP-C1 (Clontech), as previously described (32 (link)). TRPV4 K>R mutants (All-K-IDR; 5K-IDR; 5K-CTD; K130, 136R), N-terminally Myc-tagged ITCH (NP_001311127.1), and ITCH C830A (ITCH-CI) were synthesized by GenScript in the pCI-Neo vector. Myc-NEDD4 and Myc-NEDD4-CI were generated by replacing the N-terminal HA tag in pCI-HA-NEDD4-CI (Addgene; plasmid nos.: 27002 and 26999) with a Myc tag using Gibson Assembly Master Mix (New England Biolabs). The expression plasmids for PACSIN1-Myc, PACSIN2-Myc, and PACSIN3-Myc were generous gifts from Dr Markus Plomann (University of Cologne).
The following plasmids were obtained from Addgene: pRK5-HA-Ub-K0 (17603), pRK5-HA-Ubiquitin-WT (catalog no.: 17608), pLenti-puro-HA-Ubiquitin (catalog no.: 74218-LV), pCI-HA-NEDD4 (catalog no.: 27002), pCI-HA-NEDD4-DD (catalog no.: 26999), HA-Ubiquitin (catalog no.: 18712), and RhoA-Myc (catalog no.: 12962).

Aisenberg W.H., McCray B.A., Sullivan J.M., Diehl E., DeVine L.R., Alevy J., Bagnell A.M., Carr P., Donohue J.K., Goretzki B., Cole R.N., Hellmich U.A, & Sumner C.J. (2022). Multiubiquitination of TRPV4 reduces channel activity independent of surface localization. The Journal of Biological Chemistry, 298(4), 101826.

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