gecko v2 mouse crispr knockout pooled library sanjana  (Addgene inc)

Journal: Cell reports

Article Title: CRISPR-Cas9 screening identifies an IRF1-SOCS1-mediated negative feedback loop that limits CXCL9 expression and antitumor immunity.

doi: 10.1016/j.celrep.2023.113014

Figure Lengend Snippet: Figure 5. Targeting SOCS1 in macrophages enhances CXCL9 expression CRISPR-Cas9 editing was used to generate control or Irf1- or Socs1-deficient bone marrow-derived macrophages as per Figure 3E. 1 3 105 macrophages were stimulated for 17 h with 1 ng/mL IFNg. (A and B) Expression of Cxcl9 determined (A) by flow cytometry and (B) by cytometric bead array. *p < 0.05, ****p < 0.0001, two-way ANOVA. (C) MA plot of RNA-seq analysis performed on control and Socs1 KO bone marrow-derived macrophages stimulated with 1 ng/mL IFNg for 17 h.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Deposited data Raw data This paper GSE237974 Murine scRNA sequencing data Qu et al.6 GSE150970 Human scRNA sequencing data Bassez et al.29 EGAS00001004809 STAT1 CHIPseq Hogg et al.28 GSE94134 Experimental models: Cell lines Mouse: AT-3 Trina Stewart N/A Mouse: E0771 Robin Anderson N/A Mouse: B16F10 ATCC CRL-6475 Human: OVCAR-3 ATCC HTB-161 Human: MCF7 ATCC HTB-22 Human: HEK293T ATCC CRL-3216 Experimental models: Organisms/strains Mouse: Human-Her2 (hHer2) Bred in house N/A Mouse: OTI Bred in house N/A Mouse: C57BL/6 WEHI N/A Mouse: C57BL/6 Rag1 / WEHI, Australian Bioresources N/A Mouse: Ly5.1 WEHI, Australian Bioresources N/A Oligonucleotides See Table S3 for sgRNA Sequences Synthego N/A See Table S4 for Primer Sequences Integrated DNA Technologies N/A Recombinant DNA Plasmid: FUCas9Cherry Aubrey et al.45 Addgene Plasmid:70182 Pooled library: Mouse CRISPR Knockout Pooled Library (Brie) Doench et al.46 Addgene Cat:73633 Pooled library: Mouse CRISPR Knockout Pooled Library (Gecko) Sanjana et al.47 Plasmid: pMDLg/pRRE Dull et al.48 Addgene Plasmid:12251 Plasmid: pRSV-Rev Dull et al.48 Addgene Plasmid:12253 Plasmid: pMD2.G Dider Trono: Trono Laboratory Packaging and Envelope Plasmids (unpublished) Addgene Plasmid:12259 Homology repair template: Cxcl9-GFP Knock In (See Table S5 for sequence) This Paper N/A Homology repair template: BFP into Irf1 Knock In (See Table S5 for sequence) This Paper N/A Homology repair template: Irf1 + BFP into Irf1 Knock In (See Table S5 for sequence) This Paper N/A Software and algorithms CutAdapt Martin.49 https://cutadapt.readthedocs.io/en/stable/ MAGeCK v0.5.7 Li et al.50 https://sourceforge.net/projects/mageck/ RNASeQC DeLuca et al.51 www.broadinstitute.org/rna-seqc/ HISAT2 Kim et al.52 https://github.com/DaehwanKimLab/ hisat2 EdgeR v3.8.5 Robinson, McCarthy, and Smyth53 McCarthy, Chen, and Smyth54 https://bioconductor.org/packages/ release/bioc/html/edgeR.html Seurat v4.0.4 Stuart et al.55 https://github.com/satijalab/seurat/ releases/tag/v4.0.4 GENIE3 v1.16.0 Huynh-Thu et al.56 https://github.com/vahuynh/GENIE3 Bowtie2 v2.3.3 Langmead et al.57 https://bowtie-bio.sourceforge.net/index. shtml (Continued on next page) Cell Reports 42, 113014, August 29, 2023 19

Techniques: Expressing, CRISPR, Control, Derivative Assay, Cytometry, RNA Sequencing