Key Resources Table
| REAGENT or RESOURCE | SOURCE | IDENTIFIER |
|---|---|---|
| Antibodies | ||
| Anti-Ku80 mouse monoclonal 111 | Invitrogen | MA5-12933 |
| Anti-XLF Rabbit polyclonal antibody | Invitrogen | PA5-96310 |
| Anti-DNA-PKcs monoclonal antibody 42-27 | Gift from Tim Carter | N/A |
| Anti-laminB Goat polyclonal antibody | Santa Cruz | 6217 |
| Bacterial and virus strains | ||
| Biological samples | ||
| Chemicals, peptides, and recombinant proteins | ||
| Calicheamicin | Thermo-Fisher | 50-202-9283 |
| Etoposide | Sigma-Aldrich | E1383 |
| Zeocin | Invitrogen | R25001 |
| Teniposide | Millipore Sigma | 28767-20-2 |
| Camptothecin | Sigma-Aldrich | C9911 |
| KU55933 | Tocris | 3544 |
| Okadaic Acid | Thermo-Fisher | J60155 |
| Polyethylenimine | Polysciences | 23966 |
| Critical commercial assays | ||
| TOPO TA cloning kit | Invitrogen | K459540 |
| Deposited data | ||
| Experimental models: Cell lines | ||
| V3 DNA-PKcs deficient CHO cell strain | gift from Martin Gellert | CVCL_K039 |
| xrs6 Ku80 deficient CHO cell strain | gift from David Roth | CVCL_4340 |
| 293T | ATCC | CRL-3216 |
| XLF-deficient 293T cells | Roy et al. 2015 | N/A |
| XLF-deficient V3 cells | This paper | N/A |
| TDP2-deficient V3 cells | This paper | N/A |
| Experimental models: Organisms/strains | ||
| Oligonucleotides | ||
| Upstream coding joint primer: CGGTGGGAGGTCTATATAAGCA | IDT | N/A |
| Downstream coding joint primer: CTACACCGTGGTGGAGCAGTA | IDT | N/A |
| TDP2 upstream gRNA top: CACCGCTGTTGTAGGCGAGTGTGGG | IDT | N/A |
| TDP2 upstream gRNA bottom: AAACCCCACACTCGCCTACAACAGC | IDT | N/A |
| TDP2 downstream gRNA top: CACCGCATGAAGTAAGTAACAGAGT | IDT | N/A |
| TDP2 downstream gRNA bottom: AAACACTCTGTTACTTACTTCATGC | IDT | N/A |
| TDP2 5’ PCR primer: GTGCTAACTGTATGGTTTCAGTTG | IDT | N/A |
| TDP2 3’ PCR primer: CCATACCTTTGATAAAAGGAACCC | IDT | N/A |
| XLF 5’ PCR primer: TCTCTCCAGCCTCAGATTCTAA | IDT | N/A |
| XLF 3’ PCR primer: CCAATTCAACCTTCCTTGCCAAC | IDT | N/A |
| Recombinant DNA | ||
| pcDNA6A | Invitrogen | |
| WT Ku80 in pcDNA6 | Douglas et al. 2005 | N/A |
| Ku80 insert 1 mutant in pcDNA6 | This paper | N/A |
| Ku80 Insert 2 mutant in pcDNA6 | This paper | N/A |
| Ku80 Δ3 Ku80 mutant in pcDNA6 | This paper | N/A |
| Ku80 Δ6 Ku80 mutant in pcDNA6 | This paper | N/A |
| Ku80 Δ10 Ku80 mutant in pcDNA6 | This paper | N/A |
| Ku80 Δ13 Ku80 mutant in pcDNA6 | This paper | N/A |
| Ku80 Δ15 Ku80 mutant in pcDNA6 | This paper | N/A |
| Ku80 ΔCTD Ku80 mutant in pcDNA6 | This paper | N/A |
| murine RAG1 Expression Plasmid in pMAL-1 | Landree et al. 1999 | N/A |
| murine RAG2 Expression Plasmid in pMAL-1 | Landree et al. 1999 | N/A |
| murine RAG2 frameshift mutant in pMAL-1 | Corneo et al. 2007 | N/A |
| 290-Crimson/ZS VDJ coding joint substrate plasmid | Meek 2020 | N/A |
| Alt-VDJ substrate | Deriano et al., 2009 | N/A |
| Fill-in substrate | This paper | N/A |
| A/T Coding joint substrate to minimize microhomology | This paper | N/A |
| pDSRED express | Takara | 632539 |
| pEF6V5HisA | Invitrogen | V96120 |
| WT XLF in pEF6 | Roy et al. 2015 | N/A |
| L115A XLF in pEF6 | Roy et al. 2015 | N/A |
| Ku Mut XLF in pEF6 | This paper | N/A |
| L297W XLF in pEF6 | This paper | N/A |
| Ku Mut + L297W XLF in pEF6 | This paper | N/A |
| L115A + Ku Mut XLF in pEF6 | This paper | N/A |
| L115A + L297W XLF in pEF6 | This paper | N/A |
| L115A + Ku Mut + L297W XLF in pEF6 | This paper | N/A |
| pCMV6 expression plasmid | Kienker et al. 2000 | N/A |
| human DNA-PKcs in pCMV6 | Kienker et al. 2000 | N/A |
| human DNA-PKcs in RMCE | Neal et al. 2014 | N/A |
| 898 DNA-PKcs in RMCE | Chaplin et al. 2021 | N/A |
| 2569 DNA-PKcs in RMCE | Chaplin et al. 2021 | N/A |
| 4xala DNA-PKcs in RMCE | Chaplin et al. 2021 | N/A |
| 898 + 4xala DNA-PKcs in RMCE | Chaplin et al. 2021 | N/A |
| 2569 + 4xala DNA-PKcs in RMCE | This paper | N/A |
| 898+2569 DNA-PKcs in RMCE | This paper | N/A |
| 4xasp DNA-PKcs in RMCE | This paper | N/A |
| 898+902asp DNA-PKcs in RMCE | This paper | N/A |
| 898+902asp + 4xasp DNA-PKcs in RMCE | This paper | N/A |
| pSpCas9(BB)-2A-Puro | Addgene | PX459 |
| Software and algorithms | ||
| Junction Analysis | Luedean et al. 2022 | https://github.com/aluthman/Ramsden-Lab/tree/main/Luedeman_et_al_2022 |
| PRISM 9 for MacOS | GraphPad | version 9.4.1 |
| Other | ||