other:Article Title: A TRIM21-based bioPROTAC highlights the therapeutic benefit of HuR degradation.
Article Snippet: Prior to in vitro mRNA transcription, pcGEMHE or pcDNA 3.1 constructs were linearised via PCR 5’-capped modified RNA was synthesised according to the manufacturer’s protocol using HiScribe T7 ARCA mRNA Kit (New England Biolabs) or HiScribe T7 High Yield RNA Synthesis Kit (New England Biolabs), CleanCap (Trilink) and 5- Methoxyuridine-5′-Triphosphate (5-moUTP) (40% final concentration) (Trilink) respectively.
Cell Culture:Article Title: Identification of RNF114 as ADPr-Ub reader through non-hydrolysable ubiquitinated ADP-ribose
Article Snippet: .. Briefly, human U2OS osteosarcoma (American Type Culture Collection (ATCC), HTB-96) cells were cultured in Dulbecco’s modified Eagle’s medium (Sigma-Aldrich) supplemented with 10% fetal bovine serum (FBS; Gibco) and penicillin-streptomycin (100 U/ml; Gibco) in a humidified atmosphere at 37 °C with 5% CO 2 . .. Cells were plated on an eight-well μ-slide glass bottom chamber slide (ibidi) and transfected with an expression plasmid for YFP-RNF114 WT, YFP-RNF114 UIM mutant (V220G,L221G,S224G), YFP-RNF114 ΔUIM (1-208), or ADPr binding mutants YFP-RNF114, Y186A, and C176A together with PaTagRFP-H2B using TransIT-LT1 Transfection Reagent (Mirus Bio), according to the manufacturer’s instructions.
Article Title: Specificity and recognition of the ADP-ribosyl-ubiquitin modification in the DNA damage response.
Article Snippet: .. Briefly, human U2OS osteosarcoma (American Type Culture Collection (ATCC HTB-96) WT, DTX2KO, DTX3LKO, [69] or HPF1KO [70] cells were cultured in Dulbecco’s modified Eagle’s medium (Sigma-Aldrich) supplemented with 10% fetal bovine serum (FBS; Gibco) and penicillin-streptomycin (100 U/ml; Gibco) in a humidified atmosphere at 37°C with 5% CO 2 . .. Cells were plated on an eight- well μ-slide glass bottom chamber slide (ibidi) and transfected with an expression plasmid for YFP-RNF114 WT, YFPRNF114 Znf2-3 + UIM (amino acid 140−228 of RNF114), RNF138, RNF166 or YFP-RNF166 ZnF2-3 + UIM (amino acids 149−237 of RNF166), mCherry-RNF114 or YFP-DTX2 using TransIT-LT1 Transfection Reagent (Mirus Bio), according to the manufacturer’s instructions.
Article Title: Specificity and recognition of the ADP-ribosyl-ubiquitin modification in the DNA damage response
Article Snippet: .. Briefly, human U2OS osteosarcoma (American Type Culture Collection (ATCC HTB-96) WT, DTX2KO, DTX3LKO, [ ] or HPF1KO [ ] cells were cultured in Dulbecco’s modified Eagle’s medium (Sigma-Aldrich) supplemented with 10% fetal bovine serum (FBS; Gibco) and penicillin-streptomycin (100 U/ml; Gibco) in a humidified atmosphere at 37°C with 5% CO 2 . .. Cells were plated on an eight-well μ-slide glass bottom chamber slide (ibidi) and transfected with an expression plasmid for YFP-RNF114 WT, YFP-RNF114 Znf2-3 + UIM (amino acid 140−228 of RNF114), RNF138, RNF166 or YFP-RNF166 ZnF2-3 + UIM (amino acids 149−237 of RNF166), mCherry-RNF114 or YFP-DTX2 using TransIT-LT1 Transfection Reagent (Mirus Bio), according to the manufacturer’s instructions.
Article Title: Identification of RNF114 as ADPr-Ub reader through non-hydrolysable ubiquitinated ADP-ribose
Article Snippet: .. Briefly, human U2OS osteosarcoma (American Type Culture Collection (ATCC), HTB-96) cells were cultured in Dulbecco’s modified Eagle’s medium (Sigma-Aldrich) supplemented with 10% fetal bovine serum (FBS; Gibco) and penicillin-streptomycin (100 U/ml; Gibco) in a humidified atmosphere at 37°C with 5% CO2. .. Cells were plated on an eight-well μ-slide glass bottom chamber slide (ibidi) and transfected with an expression plasmid for YFP-RNF114 WT, YFP-RNF114 UIM mutant (V220G,L221G,S224G), YFP-RNF114 ΔUIM (1-208), or YFP-RNF114 ZnF2+ZnF3 mutant C176A together with PaTagRFP-H2B using TransIT-LT1 Transfection Reagent (Mirus Bio), according to the manufacturer’s instructions.
Modification:Article Title: Identification of RNF114 as ADPr-Ub reader through non-hydrolysable ubiquitinated ADP-ribose
Article Snippet: .. Briefly, human U2OS osteosarcoma (American Type Culture Collection (ATCC), HTB-96) cells were cultured in Dulbecco’s modified Eagle’s medium (Sigma-Aldrich) supplemented with 10% fetal bovine serum (FBS; Gibco) and penicillin-streptomycin (100 U/ml; Gibco) in a humidified atmosphere at 37 °C with 5% CO 2 . .. Cells were plated on an eight-well μ-slide glass bottom chamber slide (ibidi) and transfected with an expression plasmid for YFP-RNF114 WT, YFP-RNF114 UIM mutant (V220G,L221G,S224G), YFP-RNF114 ΔUIM (1-208), or ADPr binding mutants YFP-RNF114, Y186A, and C176A together with PaTagRFP-H2B using TransIT-LT1 Transfection Reagent (Mirus Bio), according to the manufacturer’s instructions.
Article Title: Specificity and recognition of the ADP-ribosyl-ubiquitin modification in the DNA damage response.
Article Snippet: .. Briefly, human U2OS osteosarcoma (American Type Culture Collection (ATCC HTB-96) WT, DTX2KO, DTX3LKO, [69] or HPF1KO [70] cells were cultured in Dulbecco’s modified Eagle’s medium (Sigma-Aldrich) supplemented with 10% fetal bovine serum (FBS; Gibco) and penicillin-streptomycin (100 U/ml; Gibco) in a humidified atmosphere at 37°C with 5% CO 2 . .. Cells were plated on an eight- well μ-slide glass bottom chamber slide (ibidi) and transfected with an expression plasmid for YFP-RNF114 WT, YFPRNF114 Znf2-3 + UIM (amino acid 140−228 of RNF114), RNF138, RNF166 or YFP-RNF166 ZnF2-3 + UIM (amino acids 149−237 of RNF166), mCherry-RNF114 or YFP-DTX2 using TransIT-LT1 Transfection Reagent (Mirus Bio), according to the manufacturer’s instructions.
Article Title: Specificity and recognition of the ADP-ribosyl-ubiquitin modification in the DNA damage response
Article Snippet: .. Briefly, human U2OS osteosarcoma (American Type Culture Collection (ATCC HTB-96) WT, DTX2KO, DTX3LKO, [ ] or HPF1KO [ ] cells were cultured in Dulbecco’s modified Eagle’s medium (Sigma-Aldrich) supplemented with 10% fetal bovine serum (FBS; Gibco) and penicillin-streptomycin (100 U/ml; Gibco) in a humidified atmosphere at 37°C with 5% CO 2 . .. Cells were plated on an eight-well μ-slide glass bottom chamber slide (ibidi) and transfected with an expression plasmid for YFP-RNF114 WT, YFP-RNF114 Znf2-3 + UIM (amino acid 140−228 of RNF114), RNF138, RNF166 or YFP-RNF166 ZnF2-3 + UIM (amino acids 149−237 of RNF166), mCherry-RNF114 or YFP-DTX2 using TransIT-LT1 Transfection Reagent (Mirus Bio), according to the manufacturer’s instructions.
Article Title: Identification of RNF114 as ADPr-Ub reader through non-hydrolysable ubiquitinated ADP-ribose
Article Snippet: .. Briefly, human U2OS osteosarcoma (American Type Culture Collection (ATCC), HTB-96) cells were cultured in Dulbecco’s modified Eagle’s medium (Sigma-Aldrich) supplemented with 10% fetal bovine serum (FBS; Gibco) and penicillin-streptomycin (100 U/ml; Gibco) in a humidified atmosphere at 37°C with 5% CO2. .. Cells were plated on an eight-well μ-slide glass bottom chamber slide (ibidi) and transfected with an expression plasmid for YFP-RNF114 WT, YFP-RNF114 UIM mutant (V220G,L221G,S224G), YFP-RNF114 ΔUIM (1-208), or YFP-RNF114 ZnF2+ZnF3 mutant C176A together with PaTagRFP-H2B using TransIT-LT1 Transfection Reagent (Mirus Bio), according to the manufacturer’s instructions.
Multiple Displacement Amplification:
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