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Flag usp28

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Flag-USP28 is a recombinant protein that contains the Flag epitope tag fused to the USP28 (ubiquitin specific peptidase 28) enzyme. USP28 is a deubiquitinating enzyme that is involved in DNA damage response pathways.

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Lab products found in correlation

Pgex 4t 1, by GE Healthcare (2 mentions) Myc usp11, by Addgene (2 mentions) Flag usp15, by Addgene (2 mentions) Quikchange site directed mutagenesis kit, by Agilent Technologies (2 mentions) Lipofectamine 3000, by Thermo Fisher Scientific (2 mentions) Hbx 41108, by Cayman Chemical (1 mentions) Pr 619, by Selleck Chemicals (1 mentions) Cycloheximide (chx), by Avantor (1 mentions) Mg132, by Selleck Chemicals (1 mentions)

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PDZ-FLAG-USP28 (2 citations)

3 protocols using flag usp28

1

Plasmid-Based Deubiquitinase Assay

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Flag-USP13, Flag-USP28, HA-CYLD lysine 63 deubiquitinase (CYLD), HA-USP46, pRetroSuper, pRetro-shUSP28-1, and pRetro-shUSP28-3 plasmids were purchased from Addgene (Watertown, MA, USA). HA-PIM1 and HA-PIM2 were gifts from Dr. Andrew Kraft. USP28-Myc, GFP-PIM3, and Flag-F-box and WD repeat domain-containing 7 (FBW7) were created using Gateway cloning. Cells were cultured to approximately 70% confluency, at which point plasmids were transfected using Lipofectamine 3000 (Thermo Fisher, Waltham, MA, USA).
MG-132, PR-619, and AZD5383 were purchased from Selleck Chemical (Houston, TX, USA). HBX 41108 was purchased from Cayman Chemical (Ann Arbor, MI, USA). Cycloheximide (CHX) was purchased from VWR (Randor, PA, USA).
Toth R.K., Solomon R, & Warfel N.A. (2022). Stabilization of PIM Kinases in Hypoxia Is Mediated by the Deubiquitinase USP28. Cells, 11(6), 1006.
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2

Detailed Plasmid Expression Constructs

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USP2, USP4, USP5, USP8, USP13, USP14, USP16, USP18, USP25, USP26, USP29, USP30, USP36, USP39, USP46, and USP48 expression plasmids were kindly provided by Dr. Jianhua Yang (Texas Children’s Cancer Center). Flag-HA-USP1, USP3, USP21, USP22, and CYLD were kindly provided by Dr. Hui-Kuan Lin (MD Anderson Cancer Center). Myc-USP11, Flag-USP15, and Flag-USP28 were from Addgene. Myc-GSK3β-WT, Myc-GSK3β-CA, and Myc-GSK3β-KD were kindly provided by Dr. Mien-Chie Hung (MD Anderson Cancer Center). Flag-KDM1A was kindly provided by Dr. Yang Shi (Harvard University). USP22 cDNA from Flag-HA-USP22 plasmid was further cloned into pcDNA3-HA vector and pGEX-4T-1 vector.
Deletion mutants of KDM1A were constructed into 3×Flag vector. Site-directed mutagenesis in KDM1A was introduced using the QuikChange site-directed mutagenesis kit (Agilent Technologies). GST-tagged KDM1A or KDM1A S683A/S687A was constructed by cloning the wild-type or mutant cDNA into pGEX-4T-1 (GE Healthcare). Lentiviral expression of KDM1A-shR, KDM1A S683D, or KDM1A S683A was achieved by cloning the cDNA into a pLVX-IRES-Hyg vector. The cDNAs of three CK1 isoforms were amplified and cloned into pcDNA3-HA vector. All plasmids were confirmed by DNA sequencing. Supplementary Table 2 contains the detail information about the sequence of the used primers.
Zhou A., Lin K., Zhang S., Chen Y., Zhang N., Xue J., Wang Z., Aldape K.D., Xie K., Woodgett J.R, & Huang S. (2016). Nuclear GSK3β promotes tumorigenesis by phosphorylating KDM1A and inducing its deubiquitination by USP22. Nature cell biology, 18(9), 954-966.
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3

Detailed Plasmid Expression Constructs

Check if the same lab product or an alternative is used in the 5 most similar protocols
USP2, USP4, USP5, USP8, USP13, USP14, USP16, USP18, USP25, USP26, USP29, USP30, USP36, USP39, USP46, and USP48 expression plasmids were kindly provided by Dr. Jianhua Yang (Texas Children’s Cancer Center). Flag-HA-USP1, USP3, USP21, USP22, and CYLD were kindly provided by Dr. Hui-Kuan Lin (MD Anderson Cancer Center). Myc-USP11, Flag-USP15, and Flag-USP28 were from Addgene. Myc-GSK3β-WT, Myc-GSK3β-CA, and Myc-GSK3β-KD were kindly provided by Dr. Mien-Chie Hung (MD Anderson Cancer Center). Flag-KDM1A was kindly provided by Dr. Yang Shi (Harvard University). USP22 cDNA from Flag-HA-USP22 plasmid was further cloned into pcDNA3-HA vector and pGEX-4T-1 vector.
Deletion mutants of KDM1A were constructed into 3×Flag vector. Site-directed mutagenesis in KDM1A was introduced using the QuikChange site-directed mutagenesis kit (Agilent Technologies). GST-tagged KDM1A or KDM1A S683A/S687A was constructed by cloning the wild-type or mutant cDNA into pGEX-4T-1 (GE Healthcare). Lentiviral expression of KDM1A-shR, KDM1A S683D, or KDM1A S683A was achieved by cloning the cDNA into a pLVX-IRES-Hyg vector. The cDNAs of three CK1 isoforms were amplified and cloned into pcDNA3-HA vector. All plasmids were confirmed by DNA sequencing. Supplementary Table 2 contains the detail information about the sequence of the used primers.
Zhou A., Lin K., Zhang S., Chen Y., Zhang N., Xue J., Wang Z., Aldape K.D., Xie K., Woodgett J.R, & Huang S. (2016). Nuclear GSK3β promotes tumorigenesis by phosphorylating KDM1A and inducing its deubiquitination by USP22. Nature cell biology, 18(9), 954-966.
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