KEY RESOURCES TABLE
| REAGENT or RESOURCE | SOURCE | IDENTIFIER |
|---|---|---|
| Antibodies against human antigens | ||
| Rat monoclonal anti-AID, unconjugated (mAID-2) | eBioscience | Cat#14-5959 |
| Rabbit polyclonal anti-AID, unconjugated | Jayanta Chaudhuri | (Vuong et al., 2009) |
| Rat monoclonal anti-AID, AF647 (EK2-5G9) | BD | Cat#565785 |
| Mouse monoclonal anti-AIRE, unconjugated (C-2) | Santa Cruz | Cat#sc-373703 |
| Human recombinant anti-AIRE, APC (REA352) | Miltenyi Biotec | Cat#130-105-401 |
| Rat monoclonal anti-AIRE, eF570 (TM-724) | eBioscience | Cat#41-9534 |
| Human recombinant anti-AIRE, APC (REA352) | Miltenyi Biotec | Cat#130-105-359 |
| Mouse monoclonal anti-Bcl6, unconjugated (GI191E/A8) | Ventana | Cat# 227M |
| Mouse monoclonal anti-CD19, Biotin (HIB19) | Biolegend | Cat#302204 |
| Mouse monoclonal anti-CD19, eF450 (HIB19) | Tonbo | Cat#75-0199-T100 |
| Mouse monoclonal anti-CD19, PE (4G7) | BD | Cat#349209 |
| Mouse monoclonal anti-CD19, PE (HIB19) | BD | Cat#555413 |
| Mouse monoclonal anti-CD19, PE-CF594 (HIB19) | BD | Cat#562321 |
| Mouse monoclonal anti-CD23, FITC (9P25) | Beckman | Cat#IM0529 |
| Mouse monoclonal anti-CD19, PE-Cy7 (HIB19) | eBioscience | Cat#25-0199 |
| Mouse monoclonal anti-CD19, QDot655 (SJ25C1) | Thermo Fisher | Cat#Q10179 |
| Mouse monoclonal anti-CD19, BV786 (SJ25C1) | BD | Cat#563326 |
| Mouse monoclonal anti-CD24, APC-eF780 (eBioSN3) | eBioscience | Cat#47-0247 |
| Mouse monoclonal anti-CD27, AF647 (O323) | Biolegend | Cat#302812 |
| Mouse monoclonal anti-CD38, APC (HIT2) | Biolegend | Cat#303510 |
| Mouse monoclonal anti-CD38, PE-Cy7 (HIT2) | Biolegend | Cat#303516 |
| Mouse monoclonal anti-CD45, eF450 (2D1) | eBioscience | Cat#48-9459 |
| Mouse monoclonal anti-CD45, PE-Cy7 (HI30) | Tonbo | Cat#60-0459 |
| Mouse monoclonal anti-EpCAM, AF488 (9C4) | Biolegend | Cat#324210 |
| Rabbit monoclonal anti-Hsp90, unconjugated (C45G5) | Cell Signaling | Cat#4877 |
| Goat polyclonal F(ab)2 anti-IgD, Biotin | Southern Biotech | Cat#2032-08 |
| Goat polyclonal F(ab)2 anti-IgD, FITC | Southern Biotech | Cat#2032-02 |
| Mouse monoclonal anti-IgD, FITC (IA6-2) | BD | Cat#555778 |
| Rabbit polyclonal anti-Lamin B1 unconjugated (H-90) | Santa Cruz | Cat#sc-20682 |
| Rat monoclonal anti-Pax5, AF594 (1H9) | BioLegend | Cat#649711 |
| Mouse monoclonal anti-β-Actin, unconjugated (AC-15) | Sigma-Aldrich | Cat#A5441 |
| Antibodies against mouse antigens | ||
| Rat monoclonal anti-AID, unconjugated (mAID-2) | eBioscience | Cat#14-5959 |
| Rabbit polyclonal anti-AID, unconjugated | Jayanta Chaudhuri | (Vuong et al., 2009) |
| Rabbit polyclonal anti-AIRE, unconjugated (H-300) | Santa Cruz | Cat#sc-33188 |
| Goat polyclonal anti-AIRE, unconjugated (D-17) | Santa Cruz | Cat#sc-17986 |
| Rat monoclonal anti-AIRE, unconjugated (5H12) | eBioscience | Cat#14-5934 |
| Rat monoclonal anti-AIRE, eF660 (5H12) | eBioscience | Cat#50-5934 |
| Human recombinant anti-AIRE, APC (REA352) | Miltenyi Biotec | Cat#130-105-359 |
| Rat monoclonal anti-B220, APC-Cy7 (RA3-6B2) | Biolegend | Cat#103224 |
| Rat monoclonal anti-B220, Biotin (RA3-6B2) | Biolegend | Cat#103204 |
| Rat monoclonal anti-BAFF-R, APC (eBio7H22-E16) | eBioscience | Cat#17-5943 |
| Rat monoclonal anti-CD138, Biotin (281-2) | Biolegend | Cat#142514 |
| Rat monoclonal anti-CD138, PE-Cy7 (281-2) | Biolegend | Cat#142511 |
| Rat monoclonal anti-CD16/CD32, unconjugated (2.4G2) | Tonbo | Cat#70-0161 |
| Rat monoclonal anti-CD16/CD32, unconjugated (2.4G2) | BD | Cat#553141 |
| Rat monoclonal anti-CD19, Biotin (1D3) | BD | Cat#553784 |
| Rat monoclonal anti-CD19, BV650 (6D5) | Biolegend | Cat#115541 |
| Rat monoclonal anti-CD19, FITC (1D3) | Tonbo | Cat#35-0193 |
| Rat monoclonal anti-CD19, Pacific Blue (6D5) | Biolegend | Cat#115523 |
| Rat monoclonal anti-CD19, PE-CF594 (1D3) | BD | Cat#562291 |
| Rat monoclonal anti-CD21, APC (7G6) | BD | Cat#558658 |
| Rat monoclonal anti-CD23, PE (B3B4) | BD | Cat#553139 |
| Rat monoclonal anti-CD25, APC (PC61.5) | Tonbo | Cat#20-0251 |
| Rat monoclonal anti-CD3, APC-Cy7 (17A2) | Tonbo | Cat#25-0032 |
| Rat monoclonal anti-CD38, PE-Cy7 (90) | Biolegend | Cat#102717 |
| Rat monoclonal anti-CD4, unconjugated (GK1.5) | Biolegend | Cat#100401 |
| Rat monoclonal anti-CD4, FITC (GK1.5) | Biolegend | Cat#100406 |
| Rat monoclonal anti-CD4, FITC (GK1.5) | BD | Cat#553729 |
| Rat monoclonal anti-CD4, PE (GK1.5) | Biolegend | Cat#100408 |
| Rat monoclonal anti-CD4, PerCP-Cy5.5 (GK1.5) | Biolegend | Cat#100434 |
| Hamster monoclonal anti-CD40, unconjugated (HM40-3) | eBioscience | Cat#16-0402 |
| Hamster monoclonal anti-CD40, FITC (H40-3) | BD | Cat#553723 |
| Rat monoclonal anti-CD45, violetFluor450 (30-F11) | Tonbo | Cat#75-0451 |
| Rat monoclonal anti-CD45, biotin (30-F11) | eBioscience | Cat#13-0451-85 |
| Rat monoclonal anti-CD62L, PE-Cy7 (MEL-14) | BD | Cat#560516 |
| Hamster monoclonal anti-CD80, PerCP-Cy5.5 (16-10A1) | BD | Cat#560526 |
| Rat monoclonal anti-CD83, BV650 (Michel-19) | Biolegend | Cat#121515 |
| Rat monoclonal anti-CD86, APC (GL-1) | BD | Cat#558703 |
| Rat monoclonal anti-CD93, PE (AA4.1) | Biolegend | Cat#136503 |
| Rat monoclonal anti-CXCR4, Biotin (2B11) | eBioscience | Cat#13-9991 |
| Rat monoclonal anti-CXCR5, PE (L138D7) | Biolegend | Cat#145504 |
| Hamster monoclonal anti-FAS, PE (Jo2) | BD | Cat#554258 |
| Rat monoclonal anti-Foxp3, V450 (MF23) | BD | Cat#561293 |
| Rabbit monoclonal anti-GAPDH, unconjugated (14C10) | Cell Signaling | Cat#2188 |
| Rat monoclonal anti-GL7, AF647 (GL7) | BD | Cat#561529 |
| Rabbit monoclonal anti-Hsp90, unconjugated (C45G5) | Cell Signaling | Cat#4877 |
| Rat monoclonal anti-I-Ab, PerCP-Cy5.5 (AF6-120.1) | Biolegend | Cat#116416 |
| Rat monoclonal anti-ICOSL, PE (HK5.3) | Biolegend | Cat#107405 |
| Rat monoclonal anti-IgA, FITC (C10-3) | BD | Cat#559354 |
| Goat polyclonal anti-IgA, HRP | Bethyl | Cat#A90-103P |
| Rat monoclonal anti-IgD, PE-Cy7 (11-26c.2a) | Biolegend | Cat#405727 |
| Rat monoclonal anti-IgD, PE-Cy7 (11-26c) | eBioscience | Cat#25-5993 |
| Horse polyclonal anti-IgG, ALP | Vector Laboratories | Cat#AP-2000 |
| Goat polyclonal anti-IgG1, HRP | Jackson Immuno | Cat#115-035-205 |
| Rat monoclonal anti-IgG1, PE-CF594 (A85-1) | BD | Cat#562559 |
| Goat polyclonal anti-IgG2b, HRP | Jackson Immuno | Cat#115-035-207 |
| Goat polyclonal anti-IgG3, HRP | Jackson Immuno | Cat#115-035-209 |
| Goat polyclonal F(ab)2 anti-IgM, unconjugated | Southern Biotech | Cat#1022-01 |
| Rat monoclonal anti-IgM, APC (RMM-1) | Biolegend | Cat#406509 |
| Goat polyclonal anti-IgM, HRP | Bethyl | Cat#A90-101P |
| Rat monoclonal anti-IL-17A, BV650 (TC11-18H10) | Biolegend | Cat#506929 |
| Goat polyclonal anti-IL-22, AF647 (Poly5164) | Biolegend | Cat#516406 |
| Rat monoclonal anti-Ly6G, AF647 (1A8) | Biolegend | Cat#127610 |
| Hamster monoclonal anti-PD-1, FITC (J43) | eBioscience | Cat#11-9985 |
| Hamster monoclonal anti-PD-1, PE-Cy7 (J43) | eBioscience | Cat#25-9985 |
| Rat monoclonal anti-PD-L1, PE-Cy7 (10F.9G2) | Biolegend | Cat#124314 |
| Rat monoclonal anti-PD-L2, PE (TY25) | BD | Cat#557796 |
| Goat polyclonal anti-Pol II, unconjugated | Bethyl | Cat#A303-835A |
| Rabbit polyclonal anti-Pol II Ser5, unconjugated | Bethyl | Cat#A304-408A |
| Rabbit polyclonal anti-Spt5, unconjugated | Santa Cruz | Cat#sc-28678 |
| Hamster monoclonal anti-TCRβ, PerCP-Cy5.5 (H57-597) | Biolegend | Cat#109227 |
| Hamster monoclonal anti-TCRγδ, BV421 (GL3) | Biolegend | Cat#118119 |
| Mouse monoclonal anti-β-Actin, unconjugated (AC-15) | Sigma-Aldrich | Cat#A5441 |
| Mouse monoclonal anti-IL17A, unconjugated (eBioMM17F3) | eBioscience | Cat#16-7173-81 |
| Mouse monoclonal anti-IL17F, unconjugated (RN17) | eBioscience | Cat#16-7473-82 |
| Mouse monoclonal anti-IL22, unconjugated (IL22JOP) | eBioscience | Cat#16-7222-82 |
| Antibodies against protein tags | ||
| Mouse monoclonal anti-6xHis, unconjugated (J099B12) | Biolegend | Cat#652502 |
| Mouse monoclonal anti-c-Myc, unconjugated (9E10) | Biolegend | Cat#626802 |
| Rabbit polyclonal anti-FLAG, unconjugated | Cell Signaling | Cat#2368 |
| Mouse monoclonal anti-FLAG, magnetic beads (M2) | Sigma-Aldrich | Cat#F3165 |
| Isotype control antibodies | ||
| Goat polyclonal IgG, unconjugated | Santa Cruz | Cat#sc-2028 |
| Goat polyclonal IgG, AF647 (Poly24030) | Biolegend | Cat#403006 |
| Goat polyclonal F(ab)2 IgG, Biotin | Southern Biotech | Cat#0110-08 |
| Goat polyclonal F(ab)2 IgG, FITC | Southern Biotech | Cat#0110-02 |
| Hamster monoclonal IgG2, PE (B81-3) | BD | Cat#550085 |
| Hamster monoclonal IgG2, PerCP-Cy5.5 (B81-3) | BD | Cat#560562 |
| Hamster monoclonal IgM, FITC (G235-1) | BD | Cat#553960 |
| Mouse monoclonal IgG1, AF647 (MOPC-21) | Biolegend | Cat#400155 |
| Mouse monoclonal IgG1, APC (MOPC-21) | BD | Cat#555751 |
| Mouse monoclonal IgG1, APC (MOPC-21) | Biolegend | Cat#400120 |
| Mouse monoclonal IgG1, Biotin (MOPC-21) | Biolegend | Cat#400103 |
| Mouse monoclonal IgG1, PE (MOPC-21) | BD | Cat#555749 |
| Mouse monoclonal IgG1, PE-Cy7 (MOPC-21) | BD | Cat#555872 |
| Mouse monoclonal IgG1, PE-Cy7 (MOPC-21) | Biolegend | Cat#400126 |
| Mouse monoclonal IgG2a, FITC (X39) | BD | Cat#349051 |
| Rabbit polyclonal IgG, unconjugated | Santa Cruz | Cat#sc-2027 |
| Rabbit polyclonal IgG, unconjugated | R&D | Cat#AF-008 |
| Rat monoclonal IgG1, BV650 (RTK2071) | Biolegend | Cat#400437 |
| Rat monoclonal IgG2a, unconjugated (RTK2758) | Biolegend | Cat#400501 |
| Rat monoclonal IgG2a, APC (RTK2758) | Biolegend | Cat#400511 |
| Rat monoclonal IgG2a, Biotin (eBR2a) | eBioscience | Cat#13-4321 |
| Rat monoclonal IgG2a, eF570 (eBR2a) | eBioscience | Cat#41-4321 |
| Rat monoclonal IgG2a, PE (eBR2a) | eBioscience | Cat#12-4321 |
| Rat monoclonal IgG2a, PerCP-Cy5.5 (RTK2758) | Biolegend | Cat#400531 |
| Rat monoclonal IgG2a, PE-Cy7 (RTK2758) | Biolegend | Cat#400521 |
| Rat monoclonal IgG2b, unconjugated (eB149/10H5) | eBioscience | Cat#14-4031 |
| Rat monoclonal IgG2b, AF647 (A95-1) | BD | Cat#557691 |
| Rat monoclonal IgG2b, Biotin (RTK4530) | Biolegend | Cat#400603 |
| Rat monoclonal IgG2b, eF660 (eB149/10H5) | eBioscience | Cat#50-4031 |
| Rat monoclonal IgG2b, PE (A95-1) | BD | Cat#553989 |
| Rat monoclonal IgG2b, PE-Cy7 (RTK4530) | Biolegend | Cat#400617 |
| Human recombinant IgG, APC (REA293) | Miltenyi Biotec | Cat#130-104-615 |
| Human recombinant IgG, PE (REA293) | Miltenyi Biotec | Cat#130-104-613 |
| Secondary antibodies | ||
| Mouse monoclonal anti-Biotin, magnetic microbeads | Miltenyi Biotec | Cat#130-090-485 |
| Donkey polyclonal anti-goat IgG, HRP | Santa Cruz | Cat#sc-2020 |
| Donkey polyclonal anti-mouse IgG, AF546 | Thermo Fisher | Cat#A10036 |
| Donkey polyclonal anti-mouse IgG, CF647 | Sigma-Aldrich | Cat#SAB4600176 |
| Donkey polyclonal anti-mouse IgG, HRP | Santa Cruz | Cat#sc-2318 |
| Donkey polyclonal F(ab’)2 anti-rat IgG, HRP | Jackson Immuno | Cat#712-036-153 |
| Goat polyclonal F(ab’)2 anti-mouse IgG, Cy3 | Jackson Immuno | Cat#115-166-006 |
| Goat polyclonal F(ab’)2 anti-mouse IgG, FITC | Southern Biotech | Cat#1032-02 |
| Goat polyclonal F(ab’)2 anti-rabbit IgG, Cy3 | Jackson Immuno | Cat#111-166-047 |
| Goat polyclonal anti-rabbit IgG, HRP | Santa Cruz | Cat#sc-2004 |
| Bacterial, Fungal and Viral Strains | ||
| Candida albicans | ATCC | Cat#MYA-2876 |
| Heat Inactivated C. albicans pseudohyphae | Made in house | This paper |
| Live C. albicans pseudohyphae | Made in house | This paper |
| Biological Samples | ||
| Human blood | American Red Cross | This paper |
| Human spleen | Children’s Hospital of Michigan | This paper |
| Human thymus | Children’s Hospital of Michigan | This paper |
| Human tonsil, healthy | Children’s Hospital of Michigan | This paper |
| Human tonsil, HIGM3 | Mount Sinai Medical Center | This paper |
| Sheep red blood cell | Innovative | Cat#IC100-0210 |
| Chemicals, Peptides, and Recombinant Proteins | ||
| NP8, FITC | BioSearch | Cat#N-5050F |
| NP36, PE | BioSearch | Cat#N-5070-1 |
| NP32, KLH | BioSearch | Cat#N-5060 |
| NP4, BSA | BioSearch | Cat#N-5050L-10 |
| NP29, BSA | BioSearch | Cat#N-5050H-10 |
| UEA-1, Biotin | Vector Laboratories | Cat#B-1065 |
| Streptavidin, AF488 | Thermo Fisher | Cat#S11223 |
| Streptavidin, AF546 | Thermo Fisher | Cat#S11225 |
| Streptavidin, PerCP-Cy5.5 | BD | Cat#551419 |
| Streptavidin, BV605 | Biolegend | Cat#405229 |
| Streptavidin, BV785 | Biolegend | Cat#405249 |
| Streptavidin, Qdot605 | Thermo Fisher | Cat#Q10101MP |
| Aldehyde Reactive Probe | Cayman Chemical | Cat#10009350 |
| RNase A | Sigma | Cat# R6513 |
| Proteinase K | Qiagen | Cat# 9133 |
| HaeIII | New England Biolabs | Cat# R0108S |
| O-Allylhydroxylamine hydrochloride (AA7) | Sigma | Cat# 05983 |
| Methoxyamine hydrochloride | Sigma | Cat# 226904 |
| Alkoxyamine-6 (AA6) | Ashok Bhagwat | (Wei et al., 2017) |
| Alkoxyamine-3 (AA3) | Ashok Bhagwat | (Wei et al., 2015) |
| DBCO -Cy5 | Click Chemistry Tools | Cat# A130-1 |
| Cy5 Azide | Lumiprobe | Cat# A3330 |
| Copper bromide | Sigma | Cat# 254185 |
| TBTA Tris[(1-benzyl-1H-1,2,3-triazol-4-yl)methyl]amine | Sigma | Cat# 678937 |
| DNA Clean and Concentrator kit | Zymo research | Cat# D4014 |
| SYBR Gold nucleic acid gel stain | Invitrogen | Cat# S11494 |
| Zeta probe membrane | Bio-Rad | Cat# 1620165 |
| Complete Freund’s Adjuvant | Thermo Fisher | Cat#77140 |
| Incomplete Freund’s Adjuvant | Thermo Fisher | Cat#77145 |
| Histopaque-1077 | Sigma-Aldrich | Cat#10771 |
| Ammonium-Chloride-Potassium (ACK) Lysis Buffer | Thermo Fisher | Cat#A1049201 |
| Collagenase II | Worthington | Cat#LS004177 |
| Collagenase V | Sigma-Aldrich | Cat#C9263-1G |
| DNase I | Roche | Cat#11284932001 |
| CD40L (Human) | Peprotech | Cat#310-02 |
| IL-4 (Human) | Peprotech | Cat#200-21 |
| IFN-β (Human) | Peprotech | Cat#300-02 |
| CD40L (Mouse) | Peprotech | Cat#315-15 |
| IL-4 (Mouse) | Peprotech | Cat#214-14 |
| IL-4 (Mouse) | R&D | Cat#404-ML |
| IL-17A (Mouse) | Peprotech | Cat#210-17 |
| IL-17F (Mouse) | Peprotech | Cat#210-17F |
| IL-17RA homodimer (Mouse) | R&D | Cat# 4481-MR-100 |
| IL-21 (Mouse) | Peprotech | Cat#210-21 |
| IL-22 (Mouse) | Peprotech | Cat#210-22 |
| IL-22Rα1 (Mouse) | R&D | Cat# 4294-MR-050 |
| TGF-β1 (Mouse) | R&D | Cat#7666-MB/CF |
| AIRE and AID WT and mutant proteins, Table S2 | Kang Chen | This paper |
| Caffeic Acid Phenethyl Ester (CAPE) | Cayman | Cat#70750 |
| Carboxyfluorescein Succinimidyl Ester (CFSE) | Biolegend | Cat#422701 |
| Lipofectamine 3000 | Thermo Fisher | Cat#L3000015 |
| Protease Inhibitor Cocktail | Sigma-Aldrich | Cat#P8340 |
| TRIzol | Thermo Fisher | Cat#15596026 |
| RNALater | Thermo Fisher | Cat#AM7020 |
| RNAProtect | QIAGEN | Cat#76526 |
| NP-40 | Sigma-Aldrich | Cat#I8896 |
| 4-20% Bis-Tris Gel | GeneScript | Cat#M42012 |
| 10% Tris-Glycine Gel | Bio-Rad | Cat#4561034 |
| Polyvinylidene fluoride (PVDF) Membrane | Bio-Rad | Cat#1620177 |
| Nitrocellulose Membrane | Bio-Rad | Cat#1620115 |
| ECL Substrate | Bio-Rad | Cat#170-5061 |
| Western Blotting Chemiluminescence Luminol Reagent | Santa Cruz | Cat#sc-2048 |
| Fc Blocking Reagent | Miltenyi Biotec | Cat#130-059-901 |
| Fc Blocking Reagent | Tonbo Biosciences | Cat#70-0161 |
| CytoFix/CytoPerm | BD | Cat#554722 |
| Transcription Factor Buffer | BD | Cat#562725 |
| 7-aminoactinomycin D (7AAD) | Tonbo Biosciences | Cat#13-6993-T500 |
| 7-aminoactinomycin D (7AAD) | BD | Cat#559925 |
| Ghost Dye Violet 510 (GV510) | Tonbo Biosciences | Cat#13-0870-T500 |
| Anti-Biotin Magnetic Microbeads | Miltenyi Biotec | Cat#130-090-485 |
| 4’,6-diamidine-2’-phenylindole dihydrochloride (DAPI) | Sigma-Aldrich | Cat#D9542 |
| BluePhos Phosphate Substrate | KPL | Cat#50-88-02 |
| BbsI restriction enzyme | Thermo Fisher | Cat#ER1011 |
| Ficoll-Paque | GE Healthcare | Cat#17-1440-03 |
| Golgi Plug | BD | Cat#55029 |
| PMA | Sigma-Aldrich | Cat#P1585-1MG |
| Ionomycin | Thermo Fisher | Cat#I24222 |
| Quick T4 Ligase | New Engl. Biolabs | Cat#M2200L |
| Opti-MEM | Thermo Fisher | Cat#31985070 |
| Protein G Beads | Cell Signaling | Cat#8740 |
| Protein G Beads | Thermo Fisher | Cat#88847 |
| CelLytic M Buffer | Sigma-Aldrich | Cat#C2978 |
| Halt Phosphatase Inhibitor | Thermo Fisher | Cat#78426 |
| FluorSave mounting medium | EMD Millipore | Cat#345789 |
| HRP conjugation kit | Abcam | Cat#Ab102890 |
| Image-iT FX Signal Enhancer | Thermo Fisher | Cat#I36933 |
| Critical Commercial Assays | ||
| Mouse B Cell Isolation kit | Miltenyi Biotec | Cat#130-090-862 |
| Click-iT EdU Flow Cytometry Assay kit | Thermo Fisher | Cat#C10418 |
| Phusion Site-Directed Mutagenesis kit | Thermo Fisher | Cat#F541 |
| Amaxa Cell Line Nucleofector Kit V | Lonza | Cat#VCA-1003 |
| BCA Protein Assay kit | Thermo Fisher | Cat#23225 |
| iTaq Universal SYBR Green One-Step kit | Bio-Rad | Cat#172-5150 |
| Cells-to-CT 1-step SYBR Green kit | Thermo Fisher | Cat#A25601 |
| ChIP Assay kit | EMD Millipore | Cat#17-295 |
| Mouse IgA ELISA | Bethyl | Cat#E90-103 |
| Superscript III First Strand Synthesis | Thermo Fisher | Cat#18808051 |
| PowerSYBR Green Master Mix | Thermo Fisher | Cat#4367660 |
| Experimental Models: Cell Lines | ||
| CH12F3, WT | Tasuku Honjo | (Nakamura et al., 1996) |
| CH12F3, Aire−/− (clones 43, 53 and 69) | Kang Chen | This Paper |
| CH12F3, Acida−/− | Kefei Yu | This Paper |
| CH12F3, Ung−/− | Ashok Bhagwat | This Paper |
| HKB-11 | ATCC | Cat#12568 |
| Experimental Models: Organisms/Strains | ||
| C57BL/6J | Jackson | Cat#000664 |
| Aire+/− (B6.129S2-Airetm1.1Doi/J) | Jackson | Cat#004743 |
| μMT (B6.129S2-Ighmtm1Cgn/J) | Jackson | Cat#002288 |
| B6.AireAdig reporter | Mark Anderson | (Gardner et al., 2008) |
| Aicda −/− | Tasuku Honjo | (Muramatsu et al., 2000) |
| CD45.1 (B6.SJL-Ptprca Pepcb/BoyJ) | Jackson | Cat#002014 |
| Oligonucleotides | ||
| A) qRT-PCR Primers | ||
| Human Primers: | ||
|
ACTB: _F: AGAGCTACGAGCTGCCTGAC _R: AGCACTGTGTTGGCGTACAG |
Sigma-Aldrich | (Wang et al., 2013) |
| Iμ_F: GTGATTAAGGAGAAACACTTTGAT | Sigma-Aldrich | (Chen et al., 2009) |
| Cγ1_R: CCAGGGCTGCTGTGCCCCCA | Sigma-Aldrich | This Paper |
| Cγ3_R: CCAGGGCCGCTGTGCCCCCA | Sigma-Aldrich | This Paper |
|
AIRE: _F: CCAGGCTCTCAACTGAAGGC _R: GAATCCCGTTCCCGAGTGG |
Sigma-Aldrich | (Dudakovic et al., 2015) |
| Mouse Primers: | ||
|
Actb _F: TGCGTGACATCAAAGAGAAG _R: CGGATGTCAACGTCACACTT |
Sigma-Aldrich | (Steuerwald et al., 2000) |
|
Aicda _F: GAAAGTCACGCTGGAG _R: TCTCATGCCGTCCCTT |
Sigma-Aldrich | (Xu et al., 2015) |
|
Aire (B6.129S2-Airetm1.1Doi/J): _F: GACAAGTTCCAGGAGACGCT _R: AAGCCGTCCAGGATGCTATG |
Sigma-Aldrich | This Paper |
| Iα-Cβ circle transcript: Iα_F: CCAGGCATGGTTGAGATAGAGATAG Cβ_R: AATGGTGCTGGGCAGGAAGT |
Sigma-Aldrich | (Cao et al., 2015) |
| Iγ1-Cμ circle transcript: Iγ1_F: GGCCCTTCCAGATCTTTGAG Cμ_R: AATGGTGCTGGGCAGGAAGT |
Sigma-Aldrich | (Doi et al., 2008) |
| Iμ-Cμ germline transcript: Iμ_F: CTCTGGCCCTGCTTATTGTTG Cμ’_R: GAAGACATTTGGGAAGGACTG |
Sigma-Aldrich | (Muramatsu et al., 2000) |
| Iα-Cα germline transcript: Iα_F: CCTGGCTGTTCCCCTATGAA Cα_R: GAGCTCGTGGGAGTGTCAGTG |
Sigma-Aldrich | (Boersma et al., 2015) |
| Sμ (after ChIP): Sμ_F: TAGTAAGCGAGGCTCTAAAAAGCAT Sμ_R: AGAACAGTCCAGTGTAGGCAGTAGA |
Sigma-Aldrich | (Nowak et al., 2011) |
| Iμ (after ChIP): Iμ_F: GCTCAGCCTGGACTTTCGGTTTGGT Iμ_R: GGAGTCAAGATGGCCGATCAGAACC |
Sigma-Aldrich | (Nowak et al., 2011) |
| Sγ1 (after ChIP): Sγ1_F: TATGATGGAAAGAGGGTAGCATTCACC Sγ1_R: CTCCTTCCCAATCTCCCGTG |
Sigma-Aldrich | (Nowak et al., 2011) |
| sgRNA (CH12 mutant 69): GCACCGCACCGAGATCGCGG (PAM:TGG) |
Feng Zhang | http://crispr.mit.edu |
| sgRNA (CH12 mutants 43 and 53): ACCTAAACCAGTCCCGGAAA (PAM:GGG) |
Feng Zhang | http://crispr.mit.edu |
| B) Primers for screening CRISPR mutants | ||
| CH12 Mutant 69: _F: CTTTCCCGCTTCCTCTATCC _R: ACTGTCTATGGCCACCGC |
Sigma-Aldrich | This paper |
| CH12 Mutant 43: _F1: ACCTAAACCAGTCCCGGAAA _F2 (Second Mutation): CCATTGTTCCTGCCCCTG _R: ACCGTTTCCAAGAGGAAGGT |
Sigma-Aldrich | This paper |
| CH12 Mutant 53: _F: ACCTAAACCAGTCCCGGAAA _R: ACCGTTTCCAAGAGGAAGGT |
Sigma-Aldrich | This paper |
| C) Primers used to generate AIRE and AICDA mutants, Table S3 | Sigma-Aldrich | This paper |
| D) Oligos used to clone sgRNA for CH12 CRISPR mutants, Table S3 | Sigma-Aldrich | This paper |
| E) Primers to amplify β-Actin gDNA after DNAse I treatment | ||
|
ACTB_gDNA:_F: CAAGGCCAACCGCGAGAAGA _R: TGTGCTGGGGTCTTGGGATG |
Sigma-Aldrich | This paper |
| Recombinant DNA | ||
| pSpCas9(BB)-2A-Puro | Addgene | Cat#48139 |
| pFLAG-CMV2 | Sigma-Aldrich | Cat#E7033 |
| pcDNA3.1(-) | Thermo Fisher | Cat#V38520 |
| pcDNA3-eGFP | Thilo Hagen | N/A |
| pCMV-Tag1 | Agilent Technologies | Cat#211170 |
| Software and Algorithms | ||
| FlowJo 7 and 10 | Tree Star | N/A |
| Adobe Photoshop CS6 | Adobe | N/A |
| Prism 6 | GraphPad | N/A |
| StepOne RT-PCR software | Applied Biosystems | N/A |
| IDEAS 6.1 | Amnis | N/A |
| VectorNTI 10 | Thermo Fisher | N/A |
| ImageQuant TL 8.1 | GE Healthcare | N/A |
| IMonitor 1.1.0 | Wei Zhang | (Zhang et al., 2015) |