Journal: Scientific Reports

Article Title: Ubiquitin-mediated DNA damage response is synthetic lethal with G-quadruplex stabilizer CX-5461

doi: 10.1038/s41598-021-88988-w

Figure Lengend Snippet: Pooled CRISPR-Cas9 screens reveal genetic determinants of sensitivity to G4 drugs. ( a ) Schematic of pooled screen. ( b ) Experimental design of drug screens. HCT116 cells were transfected with the LCV2-puro library at an MOI of 0.3. After 24 h, 0.5 µg/ml puromycin was added and the cells kept in puromycin for 3 days. On day 5 drug/vehicle treatments were started in triplicate and continued until day 19. Cells were passaged with freshly added vehicle or drugs, and harvested on days 1, 4, 5, 12, 15 and 19 to make DNA for the downstream sequencing library preparation. ( c ) Top 81 depleted genes from MAGeCK output. Each row represents the corresponding drug treatment groups. Heatmap shows the MAGeCK depletion scores of the genes. The heatmap is ordered according to the depletion scores of CX-5461 (IC 50 ) group, ranking the most depleted genes on the left. To emphasize genes with the highest depletion score, the median of the color gradient (white) was shifted to 0.2. ( d ) UpSet plot showing the intersection of candidate genes undergoing negative selection with G4 stabilizers (MAGeCK P -values < 0.045) within each drug condition. In the bottom matrix, each row represents the set of genes identified for the indicated drug condition, and each column represents the intersection of the drug condition gene sets indicated by black dots. The histogram to the right of the matrix indicates the number of genes identified for each input drug condition. The histogram on the top shows the number of genes in each intersection set. Gene hits common in all CX-5461 and PDS conditions are shown in yellow.

Article Snippet: Individual sgRNAs were designed using the Deskgen Cloud CRISPR Design Software (Desktop Genetics). sgRNA cloning into LCV2-DRX2 vector was performed as previously described , .

Techniques: CRISPR, Transfection, Sequencing, Selection

Journal: 3 Biotech

Article Title: An overview of designing and selection of sgRNAs for precise genome editing by the CRISPR-Cas9 system in plants

doi: 10.1007/s13205-019-1760-2

Figure Lengend Snippet: Summary of the CRISPR-Cas9-mediated effects on target organisms (human, animals and plants)

Article Snippet: Cas CRISPR-associated protein/gene CRISPR Clustered regularly interspaced short palindromic repeats sgRNA Single-guide RNA crRNA CRISPR RNA PAM Protospacer adjacent motif CFD Cutting frequency determination ZFN Zinc finger nuclease CLD CRISPR library designer TALEN Transcription activator-like effector nuclease tracRNA Trans -activating CRISPR RNA DESKGEN Desktop genetics NGS Next-generation sequencing

Techniques: CRISPR, Knock-Out, Disruption, Mutagenesis, Virus, Clinical Proteomics

Journal: 3 Biotech

Article Title: An overview of designing and selection of sgRNAs for precise genome editing by the CRISPR-Cas9 system in plants

doi: 10.1007/s13205-019-1760-2

Figure Lengend Snippet: A representation of sgRNA structure. a Sequence composition of sgRNA (crRNA and tracrRNA components) in CRISPR-Cas9 type II system. The crRNA consists of spacer sequence (blue; 18–20 nt in length) and crRNA repeat sequence (brown 22 nt). Complementary spacer sequence pairs with the target sequence and complementary repeat sequence pairs with tracrRNA (purple). b Chimeric sgRNA sequence composition. The sgRNA is formed by the fusion of crRNA and tracrRNA with engineered tetra loop structure (green). The sgRNA sequence consists of four different sequences, spacer (blue), crRNA repeat (brown), tetra loop (green) and tracrRNA (purple). It also has six structural secondary structure, spacer, lower stem, bulge, upper stem, nexus and hairpins. c The repeat sequence of crRNA dictates recognition lobe of spCas9 localization when there is complimentary base pairing between target DNA and guide RNA. Nexus region lies in center of sgRNA: Cas9 interaction and helps in DNA cleavage. The hairpin secondary structures support in formation of this stable complex

Article Snippet: Cas CRISPR-associated protein/gene CRISPR Clustered regularly interspaced short palindromic repeats sgRNA Single-guide RNA crRNA CRISPR RNA PAM Protospacer adjacent motif CFD Cutting frequency determination ZFN Zinc finger nuclease CLD CRISPR library designer TALEN Transcription activator-like effector nuclease tracRNA Trans -activating CRISPR RNA DESKGEN Desktop genetics NGS Next-generation sequencing

Techniques: Sequencing, CRISPR

Journal: 3 Biotech

Article Title: An overview of designing and selection of sgRNAs for precise genome editing by the CRISPR-Cas9 system in plants

doi: 10.1007/s13205-019-1760-2

Figure Lengend Snippet: Potential sgRNAs (only crRNA component) selected from Benchling using IDM2 and IDM3 genes as input queries

Article Snippet: Cas CRISPR-associated protein/gene CRISPR Clustered regularly interspaced short palindromic repeats sgRNA Single-guide RNA crRNA CRISPR RNA PAM Protospacer adjacent motif CFD Cutting frequency determination ZFN Zinc finger nuclease CLD CRISPR library designer TALEN Transcription activator-like effector nuclease tracRNA Trans -activating CRISPR RNA DESKGEN Desktop genetics NGS Next-generation sequencing

Techniques:

Journal: 3 Biotech

Article Title: An overview of designing and selection of sgRNAs for precise genome editing by the CRISPR-Cas9 system in plants

doi: 10.1007/s13205-019-1760-2

Figure Lengend Snippet: Potential sgRNAs (only crRNA component) selected from Deskgen using IDM2 and IDM3 genes as input queries

Article Snippet: Cas CRISPR-associated protein/gene CRISPR Clustered regularly interspaced short palindromic repeats sgRNA Single-guide RNA crRNA CRISPR RNA PAM Protospacer adjacent motif CFD Cutting frequency determination ZFN Zinc finger nuclease CLD CRISPR library designer TALEN Transcription activator-like effector nuclease tracRNA Trans -activating CRISPR RNA DESKGEN Desktop genetics NGS Next-generation sequencing

Techniques: Sequencing, Activity Assay

Journal: 3 Biotech

Article Title: An overview of designing and selection of sgRNAs for precise genome editing by the CRISPR-Cas9 system in plants

doi: 10.1007/s13205-019-1760-2

Figure Lengend Snippet: Potential sgRNAs (only crRNA component) selected from CRISPR-P using IDM2 and IDM3 genes as input queries

Article Snippet: Cas CRISPR-associated protein/gene CRISPR Clustered regularly interspaced short palindromic repeats sgRNA Single-guide RNA crRNA CRISPR RNA PAM Protospacer adjacent motif CFD Cutting frequency determination ZFN Zinc finger nuclease CLD CRISPR library designer TALEN Transcription activator-like effector nuclease tracRNA Trans -activating CRISPR RNA DESKGEN Desktop genetics NGS Next-generation sequencing

Techniques: CRISPR