Key resources table.
| Reagent type (species) or resource |
Designation | Source or reference | RRID identifiers |
|---|---|---|---|
| Cell line (human) |
HEK293T | ATCC | RRID:CVCL_0063 |
| Cell line (human) |
hTERT-RPE1 | ATCC | RRID:CVCL_4388 |
| Cell line (human) |
RPE1 | Ian Cheeseman | |
| Antibody | Anti-GFP (B-2) | Santa Cruz Biotechnology |
RRID:AB_627695 |
| Antibody | Anti-alpha Tubulin | DSHB | RRID:AB_2315509 |
| Antibody | Goat anti-Mouse IgG (H + L) Secondary Antibody |
Thermo Fisher Scientific |
RRID:AB_228307 |
| Antibody | Goat anti-Rabbit IgG (H + L) Secondary Antibody |
Thermo Fisher Scientific |
RRID:AB_228341 |
| Bacteria (E. coli) | K12 CJ236 | NEB (E4141) | |
| Bacteria (E. coli) | 10 beta | NEB (C3020K) | |
| Recombinant DNA reagent |
pLentiGuide | Addgene (52963) | |
| Recombinant DNA reagent |
pLentiCRISPRv2 | Addgene (52961) | |
| Recombinant DNA reagent |
PLKO.1 | Addgene (8453) | |
| Recombinant DNA reagent |
pPax2 | Addgene (12260) | |
| Recombinant DNA reagent |
pMD2.G | Addgene (12259) | |
| Commercial kit | E.Z.N.A. M13 DNA Mini Kit |
Omega Bio-Tek (D69001-01) |
|
| Commercial kit | GeneJET Gel extraction kit |
Thermo Fisher (K0692) |
|
| Commercial kit | Plasmid Maxi Kit | Qiagen (12163) | |
| Commercial kit | PureLink Genomic DNA Mini Kit |
Invitrogen (K1820-01) |
|
| Chemical compound, drug |
Ampicillin | Roth (K029.2) | |
| Chemical compound, drug |
Chloramphenicol | Roth (3886.1) | |
| Chemical compound, drug |
Kanamycin | Roth (T832.3) | |
| Chemical compound, drug |
NaCl | Roth (31434) | |
| Chemical compound, drug |
ATP | NEB (756) | |
| Chemical compound, drug |
DTT | Cell Signaling Technology Europe (7016) |
|
| Chemical compound, drug |
dNTP mix | Roth (0178.1/2) | |
| Chemical compound, drug |
Penicillin-streptomycin | Sigma-Aldrich (P4333) |
|
| Chemical compound, drug |
Hygromycin | Capricorn Scientific (HYG-H) |
|
| Chemical compound, drug |
Lipofectamin 2000 | Thermo Fisher (11668019) |
|
| Chemical compound, drug |
Polybrene | Sigma Aldrich (H9268) |
|
| Chemical compound, drug |
Doxorubicin | Selleckchem (S1208) |
|
| Chemical compound, drug |
Bay-CysLT2 | Cayman Chemical (10532) |
|
| Chemical compound, drug |
Bay-u9773 | Tocris Bioscience (3138) |
|
| Other | T4 DNA ligase | NEB (M0202) | |
| Other | T7 DNA polymerase (unmodified) | NEB (M0274) | |
| Other | 2 mm electroporation cuvette | BTX (45–0125) | |
| Other | Gene Pulser electroporation system | BioRad (164–2076) | |
| Other | I-SceI | NEB (R0694) | |
| Other | DMEM | Thermo Fisher (41965–039) |
|
| Other | DMEM/F12 | Thermo Fisher (11320–074) |
|
| Other | FBS | Thermo Fisher (10270) |
|
| Other | M13KO7 helper phage | NEB (N0315) | |
| Other | Polyethylene glycol | Roth (263.2) | |
| Other | SOC outgrowth medium | Thermo Fisher (15544034) |
|
| Other | 2YT medium | Roth (6676.2) | |
| Other | T4 polynucleotide kinase | NEB (M0201) | |
| Other | T7 endonuclease | NEB (M0302) | |
| Other | OneTaq DNA polymerase | NEB (M0480) | |
| Other | Next High-Fidelity 2x PCR Master Mix | NEB (M0541) | |
| Other | NextSeq 500 | Illumina | |
| Software, algorithm |
bcl2fastq | Illumina | RRID:SCR_015058 |
| Software, algorithm |
cutadapt v.1.15 | Martin, 2011 | RRID:SCR_011841 |
| Software, algorithm |
CasOFFinder v2.4 | Bae et al., 2014 | |
| Software, algorithm |
SnpEff 4.3T | Cingolani et al., 2012 | RRID:SCR_005191 |
| Software, algorithm |
MAGeCK | Li et al., 2014 |