Recombinant:Article Title: TBK1-Zyxin signaling controls tumor-associated macrophage recruitment to mitigate antitumor immunity.
Article Snippet: .. Reagents and tools table Reagent/Resource Reference or Source Identifier or Catalog Number Experimental models HEK293T ATCC CatCRL-3216 DLD1 ATCC Cat#CCL-221 NMuMG ATCC Cat#CRL-1636 B16-F10 ATCC Cat#CRL-6475 HeLa ATCC Cat#CCL-2 THP-1 ATCC Cat#TIB-202 HUVEC Wenzhou University N/A MC38 Prof. Xiaojian Wang N/A Peritoneal Macrophages This study N/A C57BL/6 mice wildtype (M. musculus) SLAC Laboratory Animal N/A C57BL/6 mice Zyxin−/− (M. musculus) Cyagen Biosciences N/A C57BL/6 mice Sting−/− (M. musculus) Cyagen Biosciences N/A C57BL/6 mice Irf3−/− (M. musculus) RIKEN BioResource Research Center N/A C57BL/6 mice Stingflox/flox (M. musculus) Shanghai Model Organisms N/A C57BL/6 mice Cx3cr1cre (M. musculus) The Jackson Laboratory N/A NSG mice (M. musculus) SLAC Laboratory Animal N/A Recombinant DNA This study Appendix Table S1 © The Author(s) The EMBO Journal 15 D ow nloaded from https://w w w .em bopress.org on Septem ber 24, 2024 from IP 2403:3800:3283:89ca:e8b2:b81b:46b4:9aa. .. Reagent/Resource Reference or Source Identifier or Catalog Number All-in-One cDNA Synthesis SuperMix Bimake Cat#24408 EvaGreen qPCR MasterMix Abm Cat#MasterMix-R Software GraphPad Prism 8.0 GraphPad https://www.graphpad.com/ scientific-software/prism/ Origin 9.0 OriginLab https://www.originlab.com/ index.aspx?go=PRODUCTS/ Origin ImageJ ImageJ https://imagej.nih.gov/ij/ Other Expression plasmids, reagents, and antibodies Expression plasmids encoding Flag-, Myc-, or HA-tagged wild-type or mutants of human TBK1, AKT1, IRF3 2SA, and STING have been described previously (Meng et al, 2021; Zhang et al, 2017).
Article Title: An epitranscriptomic program maintains skeletal stem cell quiescence via a METTL3-FEM1B-GLI1 axis.
Article Snippet: .. Reagents and tools table Reagent/resource Reference or source Identifier or catalog number Experimental models HEK293T ATCC CatCRL-3216 Mouse: Mettl3fl/fl Biocytogen N/A Mouse: Col2-cre Biocytogen N/A Mouse: C57BL/6J Jackson Laboratory Cat#000664 Recombinant DNA pLKO.1 Addgene Cat#8453 pLKO.1-shMettl3 This study N/A pMIR-REPORT ThermoFisher Cat#AM5795 pMIR-REPORT-FEM1B This study N/A MSCV-KD-G2P This study N/A MSCV-Mettl3-G2P This study N/A MSCVnm-Puro-IRES-GFP-Intron This study N/A MSCVnm-shFem1b-G2P This study N/A Antibodies m6A Abcam Cat#ab151230 Reagent/resource Reference or source Identifier or catalog number METTL3 Proteintech Cat#15073-1-AP FEM1B Proteintech Cat#11030-1-AP GLI1 Proteintech Cat#66905-1-Ig Ub Cell Signaling Technology Cat#3936s Tubulin Abcam Cat#ab6160 IgG Proteintech Cat#B900610 Biotin anti-mouse CD45 Biolegend Cat#103104 Biotin anti-mouse TER-119 Biolegend Cat#116204 Biotin anti-mouse CD202b (Tie-2) Biolegend Cat#124006 PerCP/Cyanin5.5 Streptavidin Biolegend Cat#405214 Anti-mouse CD51-PE Biolegend Cat#104106 Anti-mouse 6C3-PE-Cy7 Biolegend Cat#108314 Anti-mouse Thy1.1-APC-Cy7 Biolegend Cat#202520 Anti-mouse Thy1.2-APC-Cy7 Biolegend Cat#105328 Anti-mouse CD105-FITC Biolegend Cat#120406 Anti-mouse CD200-APC Biolegend Cat#123809 Anti-mouse Ki67-PE-CF594 Biolegend Cat#652427 Hoechst 33342 ThermoFisher Cat#H3570 7AAD Stem Cell Cat#75001 Oligonucleotides and other sequence-based reagents Primers for RT-PCR, see Appendix Table S1 This study N/A Primers for mouse genotyping, see Appendix Table S1 This study N/A Primers for shRNA, see Appendix Table S1 This study N/A Fem1b 3’UTR sequence for luciferase assay This study N/A Chemicals, enzymes and other reagents Polybrene Sigma-Aldrich Cat#H9268 Actinomycin D Sigma-Aldrich Cat#A9415 Cycloheximide(CHX) MCE Cat#HY-12320 Smoothened Agonist(SAG) Selleck Cat#S7779 Collagenase Sigma-Aldrich Cat#6885 Protein-A beads ThermoFisher Cat#10002D Library Preparation VATHS mRNA Capture Beads Vazyme Cat#N401-01 TruSeq Stranded mRNA Library Prep Kit Illumine Cat#RS-122-2101 Pure Beads Kit KAPA Cat#KK8002 KAPA Hifi HS RM KAPA Cat#KK2602 ReverTra Ace qPCR RT Kit TOYOBO Cat#FSQ-101 Dual-Luciferase Reporter Assay System Promega Cat#1980 Hematoxylin and Eosin Staining Kit Beyotime Cat#C0105S Safranin O- fast green cartilage staining kit Solarbio Cat#G1371 © The Author(s) The EMBO Journal Volume 44 | Issue 8 | April 2025 | 2263 – 2278 2273 D ow nloaded from https://w w w .em bopress.org on July 7, 2025 from IP 45.248.78.198. .. Reagent/resource Reference or source Identifier or catalog number Alcian Blue Cartilage Stain solution Servicebio Cat#G1027 Alizarin red S Sigma-Aldrich Cat#A5533 Software GraphPad Prism 8 GraphPad Software https:// www.graphpad.com/ FlowJo software (version 10.8.1) FlowJo https:// www.flowjo.com/ Other RNA-seq This study GEO: GSE284224 SLIM-seq This study GEO: GSE284224
other:Article Title: TBK1-Zyxin signaling controls tumor-associated macrophage recruitment to mitigate antitumor immunity.
Article Snippet: Reagent/Resource Reference or Source Identifier or Catalog Number All-in-One cDNA Synthesis SuperMix Bimake Cat#24408 EvaGreen qPCR MasterMix Abm Cat#MasterMix-R Software GraphPad Prism 8.0 GraphPad https://www.graphpad.com/ scientific-software/prism/ Origin 9.0 OriginLab https://www.originlab.com/ index.aspx?go=PRODUCTS/ Origin ImageJ ImageJ https://imagej.nih.gov/ij/ Other Expression plasmids, reagents, and antibodies Expression plasmids encoding Flag-, Myc-, or HA-tagged wild-type or mutants of human TBK1, AKT1, IRF3 2SA, and STING have been described previously (Meng et al, 2021; Zhang et al, 2017).
Sequencing:Article Title: An epitranscriptomic program maintains skeletal stem cell quiescence via a METTL3-FEM1B-GLI1 axis.
Article Snippet: .. Reagents and tools table Reagent/resource Reference or source Identifier or catalog number Experimental models HEK293T ATCC CatCRL-3216 Mouse: Mettl3fl/fl Biocytogen N/A Mouse: Col2-cre Biocytogen N/A Mouse: C57BL/6J Jackson Laboratory Cat#000664 Recombinant DNA pLKO.1 Addgene Cat#8453 pLKO.1-shMettl3 This study N/A pMIR-REPORT ThermoFisher Cat#AM5795 pMIR-REPORT-FEM1B This study N/A MSCV-KD-G2P This study N/A MSCV-Mettl3-G2P This study N/A MSCVnm-Puro-IRES-GFP-Intron This study N/A MSCVnm-shFem1b-G2P This study N/A Antibodies m6A Abcam Cat#ab151230 Reagent/resource Reference or source Identifier or catalog number METTL3 Proteintech Cat#15073-1-AP FEM1B Proteintech Cat#11030-1-AP GLI1 Proteintech Cat#66905-1-Ig Ub Cell Signaling Technology Cat#3936s Tubulin Abcam Cat#ab6160 IgG Proteintech Cat#B900610 Biotin anti-mouse CD45 Biolegend Cat#103104 Biotin anti-mouse TER-119 Biolegend Cat#116204 Biotin anti-mouse CD202b (Tie-2) Biolegend Cat#124006 PerCP/Cyanin5.5 Streptavidin Biolegend Cat#405214 Anti-mouse CD51-PE Biolegend Cat#104106 Anti-mouse 6C3-PE-Cy7 Biolegend Cat#108314 Anti-mouse Thy1.1-APC-Cy7 Biolegend Cat#202520 Anti-mouse Thy1.2-APC-Cy7 Biolegend Cat#105328 Anti-mouse CD105-FITC Biolegend Cat#120406 Anti-mouse CD200-APC Biolegend Cat#123809 Anti-mouse Ki67-PE-CF594 Biolegend Cat#652427 Hoechst 33342 ThermoFisher Cat#H3570 7AAD Stem Cell Cat#75001 Oligonucleotides and other sequence-based reagents Primers for RT-PCR, see Appendix Table S1 This study N/A Primers for mouse genotyping, see Appendix Table S1 This study N/A Primers for shRNA, see Appendix Table S1 This study N/A Fem1b 3’UTR sequence for luciferase assay This study N/A Chemicals, enzymes and other reagents Polybrene Sigma-Aldrich Cat#H9268 Actinomycin D Sigma-Aldrich Cat#A9415 Cycloheximide(CHX) MCE Cat#HY-12320 Smoothened Agonist(SAG) Selleck Cat#S7779 Collagenase Sigma-Aldrich Cat#6885 Protein-A beads ThermoFisher Cat#10002D Library Preparation VATHS mRNA Capture Beads Vazyme Cat#N401-01 TruSeq Stranded mRNA Library Prep Kit Illumine Cat#RS-122-2101 Pure Beads Kit KAPA Cat#KK8002 KAPA Hifi HS RM KAPA Cat#KK2602 ReverTra Ace qPCR RT Kit TOYOBO Cat#FSQ-101 Dual-Luciferase Reporter Assay System Promega Cat#1980 Hematoxylin and Eosin Staining Kit Beyotime Cat#C0105S Safranin O- fast green cartilage staining kit Solarbio Cat#G1371 © The Author(s) The EMBO Journal Volume 44 | Issue 8 | April 2025 | 2263 – 2278 2273 D ow nloaded from https://w w w .em bopress.org on July 7, 2025 from IP 45.248.78.198. .. Reagent/resource Reference or source Identifier or catalog number Alcian Blue Cartilage Stain solution Servicebio Cat#G1027 Alizarin red S Sigma-Aldrich Cat#A5533 Software GraphPad Prism 8 GraphPad Software https:// www.graphpad.com/ FlowJo software (version 10.8.1) FlowJo https:// www.flowjo.com/ Other RNA-seq This study GEO: GSE284224 SLIM-seq This study GEO: GSE284224
shRNA:Article Title: An epitranscriptomic program maintains skeletal stem cell quiescence via a METTL3-FEM1B-GLI1 axis.
Article Snippet: .. Reagents and tools table Reagent/resource Reference or source Identifier or catalog number Experimental models HEK293T ATCC CatCRL-3216 Mouse: Mettl3fl/fl Biocytogen N/A Mouse: Col2-cre Biocytogen N/A Mouse: C57BL/6J Jackson Laboratory Cat#000664 Recombinant DNA pLKO.1 Addgene Cat#8453 pLKO.1-shMettl3 This study N/A pMIR-REPORT ThermoFisher Cat#AM5795 pMIR-REPORT-FEM1B This study N/A MSCV-KD-G2P This study N/A MSCV-Mettl3-G2P This study N/A MSCVnm-Puro-IRES-GFP-Intron This study N/A MSCVnm-shFem1b-G2P This study N/A Antibodies m6A Abcam Cat#ab151230 Reagent/resource Reference or source Identifier or catalog number METTL3 Proteintech Cat#15073-1-AP FEM1B Proteintech Cat#11030-1-AP GLI1 Proteintech Cat#66905-1-Ig Ub Cell Signaling Technology Cat#3936s Tubulin Abcam Cat#ab6160 IgG Proteintech Cat#B900610 Biotin anti-mouse CD45 Biolegend Cat#103104 Biotin anti-mouse TER-119 Biolegend Cat#116204 Biotin anti-mouse CD202b (Tie-2) Biolegend Cat#124006 PerCP/Cyanin5.5 Streptavidin Biolegend Cat#405214 Anti-mouse CD51-PE Biolegend Cat#104106 Anti-mouse 6C3-PE-Cy7 Biolegend Cat#108314 Anti-mouse Thy1.1-APC-Cy7 Biolegend Cat#202520 Anti-mouse Thy1.2-APC-Cy7 Biolegend Cat#105328 Anti-mouse CD105-FITC Biolegend Cat#120406 Anti-mouse CD200-APC Biolegend Cat#123809 Anti-mouse Ki67-PE-CF594 Biolegend Cat#652427 Hoechst 33342 ThermoFisher Cat#H3570 7AAD Stem Cell Cat#75001 Oligonucleotides and other sequence-based reagents Primers for RT-PCR, see Appendix Table S1 This study N/A Primers for mouse genotyping, see Appendix Table S1 This study N/A Primers for shRNA, see Appendix Table S1 This study N/A Fem1b 3’UTR sequence for luciferase assay This study N/A Chemicals, enzymes and other reagents Polybrene Sigma-Aldrich Cat#H9268 Actinomycin D Sigma-Aldrich Cat#A9415 Cycloheximide(CHX) MCE Cat#HY-12320 Smoothened Agonist(SAG) Selleck Cat#S7779 Collagenase Sigma-Aldrich Cat#6885 Protein-A beads ThermoFisher Cat#10002D Library Preparation VATHS mRNA Capture Beads Vazyme Cat#N401-01 TruSeq Stranded mRNA Library Prep Kit Illumine Cat#RS-122-2101 Pure Beads Kit KAPA Cat#KK8002 KAPA Hifi HS RM KAPA Cat#KK2602 ReverTra Ace qPCR RT Kit TOYOBO Cat#FSQ-101 Dual-Luciferase Reporter Assay System Promega Cat#1980 Hematoxylin and Eosin Staining Kit Beyotime Cat#C0105S Safranin O- fast green cartilage staining kit Solarbio Cat#G1371 © The Author(s) The EMBO Journal Volume 44 | Issue 8 | April 2025 | 2263 – 2278 2273 D ow nloaded from https://w w w .em bopress.org on July 7, 2025 from IP 45.248.78.198. .. Reagent/resource Reference or source Identifier or catalog number Alcian Blue Cartilage Stain solution Servicebio Cat#G1027 Alizarin red S Sigma-Aldrich Cat#A5533 Software GraphPad Prism 8 GraphPad Software https:// www.graphpad.com/ FlowJo software (version 10.8.1) FlowJo https:// www.flowjo.com/ Other RNA-seq This study GEO: GSE284224 SLIM-seq This study GEO: GSE284224
Luciferase:Article Title: An epitranscriptomic program maintains skeletal stem cell quiescence via a METTL3-FEM1B-GLI1 axis.
Article Snippet: .. Reagents and tools table Reagent/resource Reference or source Identifier or catalog number Experimental models HEK293T ATCC CatCRL-3216 Mouse: Mettl3fl/fl Biocytogen N/A Mouse: Col2-cre Biocytogen N/A Mouse: C57BL/6J Jackson Laboratory Cat#000664 Recombinant DNA pLKO.1 Addgene Cat#8453 pLKO.1-shMettl3 This study N/A pMIR-REPORT ThermoFisher Cat#AM5795 pMIR-REPORT-FEM1B This study N/A MSCV-KD-G2P This study N/A MSCV-Mettl3-G2P This study N/A MSCVnm-Puro-IRES-GFP-Intron This study N/A MSCVnm-shFem1b-G2P This study N/A Antibodies m6A Abcam Cat#ab151230 Reagent/resource Reference or source Identifier or catalog number METTL3 Proteintech Cat#15073-1-AP FEM1B Proteintech Cat#11030-1-AP GLI1 Proteintech Cat#66905-1-Ig Ub Cell Signaling Technology Cat#3936s Tubulin Abcam Cat#ab6160 IgG Proteintech Cat#B900610 Biotin anti-mouse CD45 Biolegend Cat#103104 Biotin anti-mouse TER-119 Biolegend Cat#116204 Biotin anti-mouse CD202b (Tie-2) Biolegend Cat#124006 PerCP/Cyanin5.5 Streptavidin Biolegend Cat#405214 Anti-mouse CD51-PE Biolegend Cat#104106 Anti-mouse 6C3-PE-Cy7 Biolegend Cat#108314 Anti-mouse Thy1.1-APC-Cy7 Biolegend Cat#202520 Anti-mouse Thy1.2-APC-Cy7 Biolegend Cat#105328 Anti-mouse CD105-FITC Biolegend Cat#120406 Anti-mouse CD200-APC Biolegend Cat#123809 Anti-mouse Ki67-PE-CF594 Biolegend Cat#652427 Hoechst 33342 ThermoFisher Cat#H3570 7AAD Stem Cell Cat#75001 Oligonucleotides and other sequence-based reagents Primers for RT-PCR, see Appendix Table S1 This study N/A Primers for mouse genotyping, see Appendix Table S1 This study N/A Primers for shRNA, see Appendix Table S1 This study N/A Fem1b 3’UTR sequence for luciferase assay This study N/A Chemicals, enzymes and other reagents Polybrene Sigma-Aldrich Cat#H9268 Actinomycin D Sigma-Aldrich Cat#A9415 Cycloheximide(CHX) MCE Cat#HY-12320 Smoothened Agonist(SAG) Selleck Cat#S7779 Collagenase Sigma-Aldrich Cat#6885 Protein-A beads ThermoFisher Cat#10002D Library Preparation VATHS mRNA Capture Beads Vazyme Cat#N401-01 TruSeq Stranded mRNA Library Prep Kit Illumine Cat#RS-122-2101 Pure Beads Kit KAPA Cat#KK8002 KAPA Hifi HS RM KAPA Cat#KK2602 ReverTra Ace qPCR RT Kit TOYOBO Cat#FSQ-101 Dual-Luciferase Reporter Assay System Promega Cat#1980 Hematoxylin and Eosin Staining Kit Beyotime Cat#C0105S Safranin O- fast green cartilage staining kit Solarbio Cat#G1371 © The Author(s) The EMBO Journal Volume 44 | Issue 8 | April 2025 | 2263 – 2278 2273 D ow nloaded from https://w w w .em bopress.org on July 7, 2025 from IP 45.248.78.198. .. Reagent/resource Reference or source Identifier or catalog number Alcian Blue Cartilage Stain solution Servicebio Cat#G1027 Alizarin red S Sigma-Aldrich Cat#A5533 Software GraphPad Prism 8 GraphPad Software https:// www.graphpad.com/ FlowJo software (version 10.8.1) FlowJo https:// www.flowjo.com/ Other RNA-seq This study GEO: GSE284224 SLIM-seq This study GEO: GSE284224
Real-time Polymerase Chain Reaction:Article Title: An epitranscriptomic program maintains skeletal stem cell quiescence via a METTL3-FEM1B-GLI1 axis.
Article Snippet: .. Reagents and tools table Reagent/resource Reference or source Identifier or catalog number Experimental models HEK293T ATCC CatCRL-3216 Mouse: Mettl3fl/fl Biocytogen N/A Mouse: Col2-cre Biocytogen N/A Mouse: C57BL/6J Jackson Laboratory Cat#000664 Recombinant DNA pLKO.1 Addgene Cat#8453 pLKO.1-shMettl3 This study N/A pMIR-REPORT ThermoFisher Cat#AM5795 pMIR-REPORT-FEM1B This study N/A MSCV-KD-G2P This study N/A MSCV-Mettl3-G2P This study N/A MSCVnm-Puro-IRES-GFP-Intron This study N/A MSCVnm-shFem1b-G2P This study N/A Antibodies m6A Abcam Cat#ab151230 Reagent/resource Reference or source Identifier or catalog number METTL3 Proteintech Cat#15073-1-AP FEM1B Proteintech Cat#11030-1-AP GLI1 Proteintech Cat#66905-1-Ig Ub Cell Signaling Technology Cat#3936s Tubulin Abcam Cat#ab6160 IgG Proteintech Cat#B900610 Biotin anti-mouse CD45 Biolegend Cat#103104 Biotin anti-mouse TER-119 Biolegend Cat#116204 Biotin anti-mouse CD202b (Tie-2) Biolegend Cat#124006 PerCP/Cyanin5.5 Streptavidin Biolegend Cat#405214 Anti-mouse CD51-PE Biolegend Cat#104106 Anti-mouse 6C3-PE-Cy7 Biolegend Cat#108314 Anti-mouse Thy1.1-APC-Cy7 Biolegend Cat#202520 Anti-mouse Thy1.2-APC-Cy7 Biolegend Cat#105328 Anti-mouse CD105-FITC Biolegend Cat#120406 Anti-mouse CD200-APC Biolegend Cat#123809 Anti-mouse Ki67-PE-CF594 Biolegend Cat#652427 Hoechst 33342 ThermoFisher Cat#H3570 7AAD Stem Cell Cat#75001 Oligonucleotides and other sequence-based reagents Primers for RT-PCR, see Appendix Table S1 This study N/A Primers for mouse genotyping, see Appendix Table S1 This study N/A Primers for shRNA, see Appendix Table S1 This study N/A Fem1b 3’UTR sequence for luciferase assay This study N/A Chemicals, enzymes and other reagents Polybrene Sigma-Aldrich Cat#H9268 Actinomycin D Sigma-Aldrich Cat#A9415 Cycloheximide(CHX) MCE Cat#HY-12320 Smoothened Agonist(SAG) Selleck Cat#S7779 Collagenase Sigma-Aldrich Cat#6885 Protein-A beads ThermoFisher Cat#10002D Library Preparation VATHS mRNA Capture Beads Vazyme Cat#N401-01 TruSeq Stranded mRNA Library Prep Kit Illumine Cat#RS-122-2101 Pure Beads Kit KAPA Cat#KK8002 KAPA Hifi HS RM KAPA Cat#KK2602 ReverTra Ace qPCR RT Kit TOYOBO Cat#FSQ-101 Dual-Luciferase Reporter Assay System Promega Cat#1980 Hematoxylin and Eosin Staining Kit Beyotime Cat#C0105S Safranin O- fast green cartilage staining kit Solarbio Cat#G1371 © The Author(s) The EMBO Journal Volume 44 | Issue 8 | April 2025 | 2263 – 2278 2273 D ow nloaded from https://w w w .em bopress.org on July 7, 2025 from IP 45.248.78.198. .. Reagent/resource Reference or source Identifier or catalog number Alcian Blue Cartilage Stain solution Servicebio Cat#G1027 Alizarin red S Sigma-Aldrich Cat#A5533 Software GraphPad Prism 8 GraphPad Software https:// www.graphpad.com/ FlowJo software (version 10.8.1) FlowJo https:// www.flowjo.com/ Other RNA-seq This study GEO: GSE284224 SLIM-seq This study GEO: GSE284224
Reporter Assay:Article Title: An epitranscriptomic program maintains skeletal stem cell quiescence via a METTL3-FEM1B-GLI1 axis.
Article Snippet: .. Reagents and tools table Reagent/resource Reference or source Identifier or catalog number Experimental models HEK293T ATCC CatCRL-3216 Mouse: Mettl3fl/fl Biocytogen N/A Mouse: Col2-cre Biocytogen N/A Mouse: C57BL/6J Jackson Laboratory Cat#000664 Recombinant DNA pLKO.1 Addgene Cat#8453 pLKO.1-shMettl3 This study N/A pMIR-REPORT ThermoFisher Cat#AM5795 pMIR-REPORT-FEM1B This study N/A MSCV-KD-G2P This study N/A MSCV-Mettl3-G2P This study N/A MSCVnm-Puro-IRES-GFP-Intron This study N/A MSCVnm-shFem1b-G2P This study N/A Antibodies m6A Abcam Cat#ab151230 Reagent/resource Reference or source Identifier or catalog number METTL3 Proteintech Cat#15073-1-AP FEM1B Proteintech Cat#11030-1-AP GLI1 Proteintech Cat#66905-1-Ig Ub Cell Signaling Technology Cat#3936s Tubulin Abcam Cat#ab6160 IgG Proteintech Cat#B900610 Biotin anti-mouse CD45 Biolegend Cat#103104 Biotin anti-mouse TER-119 Biolegend Cat#116204 Biotin anti-mouse CD202b (Tie-2) Biolegend Cat#124006 PerCP/Cyanin5.5 Streptavidin Biolegend Cat#405214 Anti-mouse CD51-PE Biolegend Cat#104106 Anti-mouse 6C3-PE-Cy7 Biolegend Cat#108314 Anti-mouse Thy1.1-APC-Cy7 Biolegend Cat#202520 Anti-mouse Thy1.2-APC-Cy7 Biolegend Cat#105328 Anti-mouse CD105-FITC Biolegend Cat#120406 Anti-mouse CD200-APC Biolegend Cat#123809 Anti-mouse Ki67-PE-CF594 Biolegend Cat#652427 Hoechst 33342 ThermoFisher Cat#H3570 7AAD Stem Cell Cat#75001 Oligonucleotides and other sequence-based reagents Primers for RT-PCR, see Appendix Table S1 This study N/A Primers for mouse genotyping, see Appendix Table S1 This study N/A Primers for shRNA, see Appendix Table S1 This study N/A Fem1b 3’UTR sequence for luciferase assay This study N/A Chemicals, enzymes and other reagents Polybrene Sigma-Aldrich Cat#H9268 Actinomycin D Sigma-Aldrich Cat#A9415 Cycloheximide(CHX) MCE Cat#HY-12320 Smoothened Agonist(SAG) Selleck Cat#S7779 Collagenase Sigma-Aldrich Cat#6885 Protein-A beads ThermoFisher Cat#10002D Library Preparation VATHS mRNA Capture Beads Vazyme Cat#N401-01 TruSeq Stranded mRNA Library Prep Kit Illumine Cat#RS-122-2101 Pure Beads Kit KAPA Cat#KK8002 KAPA Hifi HS RM KAPA Cat#KK2602 ReverTra Ace qPCR RT Kit TOYOBO Cat#FSQ-101 Dual-Luciferase Reporter Assay System Promega Cat#1980 Hematoxylin and Eosin Staining Kit Beyotime Cat#C0105S Safranin O- fast green cartilage staining kit Solarbio Cat#G1371 © The Author(s) The EMBO Journal Volume 44 | Issue 8 | April 2025 | 2263 – 2278 2273 D ow nloaded from https://w w w .em bopress.org on July 7, 2025 from IP 45.248.78.198. .. Reagent/resource Reference or source Identifier or catalog number Alcian Blue Cartilage Stain solution Servicebio Cat#G1027 Alizarin red S Sigma-Aldrich Cat#A5533 Software GraphPad Prism 8 GraphPad Software https:// www.graphpad.com/ FlowJo software (version 10.8.1) FlowJo https:// www.flowjo.com/ Other RNA-seq This study GEO: GSE284224 SLIM-seq This study GEO: GSE284224
Staining:Article Title: An epitranscriptomic program maintains skeletal stem cell quiescence via a METTL3-FEM1B-GLI1 axis.
Article Snippet: .. Reagents and tools table Reagent/resource Reference or source Identifier or catalog number Experimental models HEK293T ATCC CatCRL-3216 Mouse: Mettl3fl/fl Biocytogen N/A Mouse: Col2-cre Biocytogen N/A Mouse: C57BL/6J Jackson Laboratory Cat#000664 Recombinant DNA pLKO.1 Addgene Cat#8453 pLKO.1-shMettl3 This study N/A pMIR-REPORT ThermoFisher Cat#AM5795 pMIR-REPORT-FEM1B This study N/A MSCV-KD-G2P This study N/A MSCV-Mettl3-G2P This study N/A MSCVnm-Puro-IRES-GFP-Intron This study N/A MSCVnm-shFem1b-G2P This study N/A Antibodies m6A Abcam Cat#ab151230 Reagent/resource Reference or source Identifier or catalog number METTL3 Proteintech Cat#15073-1-AP FEM1B Proteintech Cat#11030-1-AP GLI1 Proteintech Cat#66905-1-Ig Ub Cell Signaling Technology Cat#3936s Tubulin Abcam Cat#ab6160 IgG Proteintech Cat#B900610 Biotin anti-mouse CD45 Biolegend Cat#103104 Biotin anti-mouse TER-119 Biolegend Cat#116204 Biotin anti-mouse CD202b (Tie-2) Biolegend Cat#124006 PerCP/Cyanin5.5 Streptavidin Biolegend Cat#405214 Anti-mouse CD51-PE Biolegend Cat#104106 Anti-mouse 6C3-PE-Cy7 Biolegend Cat#108314 Anti-mouse Thy1.1-APC-Cy7 Biolegend Cat#202520 Anti-mouse Thy1.2-APC-Cy7 Biolegend Cat#105328 Anti-mouse CD105-FITC Biolegend Cat#120406 Anti-mouse CD200-APC Biolegend Cat#123809 Anti-mouse Ki67-PE-CF594 Biolegend Cat#652427 Hoechst 33342 ThermoFisher Cat#H3570 7AAD Stem Cell Cat#75001 Oligonucleotides and other sequence-based reagents Primers for RT-PCR, see Appendix Table S1 This study N/A Primers for mouse genotyping, see Appendix Table S1 This study N/A Primers for shRNA, see Appendix Table S1 This study N/A Fem1b 3’UTR sequence for luciferase assay This study N/A Chemicals, enzymes and other reagents Polybrene Sigma-Aldrich Cat#H9268 Actinomycin D Sigma-Aldrich Cat#A9415 Cycloheximide(CHX) MCE Cat#HY-12320 Smoothened Agonist(SAG) Selleck Cat#S7779 Collagenase Sigma-Aldrich Cat#6885 Protein-A beads ThermoFisher Cat#10002D Library Preparation VATHS mRNA Capture Beads Vazyme Cat#N401-01 TruSeq Stranded mRNA Library Prep Kit Illumine Cat#RS-122-2101 Pure Beads Kit KAPA Cat#KK8002 KAPA Hifi HS RM KAPA Cat#KK2602 ReverTra Ace qPCR RT Kit TOYOBO Cat#FSQ-101 Dual-Luciferase Reporter Assay System Promega Cat#1980 Hematoxylin and Eosin Staining Kit Beyotime Cat#C0105S Safranin O- fast green cartilage staining kit Solarbio Cat#G1371 © The Author(s) The EMBO Journal Volume 44 | Issue 8 | April 2025 | 2263 – 2278 2273 D ow nloaded from https://w w w .em bopress.org on July 7, 2025 from IP 45.248.78.198. .. Reagent/resource Reference or source Identifier or catalog number Alcian Blue Cartilage Stain solution Servicebio Cat#G1027 Alizarin red S Sigma-Aldrich Cat#A5533 Software GraphPad Prism 8 GraphPad Software https:// www.graphpad.com/ FlowJo software (version 10.8.1) FlowJo https:// www.flowjo.com/ Other RNA-seq This study GEO: GSE284224 SLIM-seq This study GEO: GSE284224
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