Journal: PLoS ONE

Article Title: Prickle isoform participation in distinct polarization events in the Drosophila eye

doi: 10.1371/journal.pone.0262328

Figure Lengend Snippet:

Article Snippet: Recombinant DNA reagent , pDsRedattp , Addgene , RRID: Addgene_51019 , Donor recombinant DNA backbone.

Techniques: Western Blot, Recombinant, CRISPR, Sequencing

Journal: Nature Communications

Article Title: A high-content RNAi screen reveals multiple roles for long noncoding RNAs in cell division

doi: 10.1038/s41467-020-14978-7

Figure Lengend Snippet: a Schematic diagram of ectopic overexpression of linc00899. Linc00899 and TPPP expression were analysed by qPCR after lentiviral overexpression using lincXpress vector encoding linc00899 cDNA in HeLa (left) and RPE1 cells (right). The expression was normalised to the scrambled linc00899 vector (negative control). n = 3–4 biological replicates, * P < 0.05 by two-tailed Student’s t test. b Rescue of linc00899 function after RNAi- or LNA-mediated depletion of linc00899 in HeLa cells through ectopic overexpression of linc00899 . n = 3 biological replicates, * P < 0.05, ** P < 0.01 and **** P < 0.0001 by two-tailed Student’s t test. c Schematic diagram of endogenous overexpression of linc00899 using CRISPRa . Linc00899 and TPPP expression were analysed by qPCR after transduction of dCas9-VP64 and gRNAs targeting different regions of the linc00899 promoter in HeLa (left) and RPE1 cells (right). The expression was normalised to the negative guide (negative control). n = 4 biological replicates, * P < 0.05, *** P < 0.001 and **** P < 0.0001 by two-tailed Student’s t test. Data are shown as mean ± S.E.M for a – c . Source data are provided as a Source Data file.

Article Snippet: For CRISPR activation (CRISPRa), pHAGE EF1alpha dCAS-VP64-HA (Addgene, #50918), pU6-sgRNA EF1Alpha-puro-T2A-BFP (Addgene, #60955), second-generation packaging plasmid psPAX2 (Addgene, #12260) and the envelope plasmid pMD2.G (Addgene, #12259) were used.

Techniques: Over Expression, Expressing, Plasmid Preparation, Negative Control, Two Tailed Test, Transduction

Journal: Nature genetics

Article Title: The impact of nonsense-mediated mRNA decay on genetic disease, gene editing and cancer immunotherapy

doi: 10.1038/s41588-019-0517-5

Figure Lengend Snippet: a , a decrease in protein expression due to tiling sgRNAs placed along the length of human and mouse genes (y axis quantifies the sgRNA fold difference between a low-expressing versus high-expressing set of cells) reveals, overall, similar associations with the non-canonical start-proximal NMD rule to the canonical NMD last-exon rule. The CD13 gene demonstrates the effect of the non-canonical long-exon rule. Shaded regions are 95% confidence interval of the loess fit to protein expression. Pearson correlation coefficients and two-sided tests for association were computed by comparing the loess fit to the NMDetective -A NMD efficacy scores. b , evading NMD attenuates the loss of fitness (y axis) due to knockout of essential genes. Data for non-essential genes are in . P values are by Mann-Whitney U test. The knock-out efficiency compares the reduction of sgRNAs in NMD evading regions to the reduction in regions that trigger NMD. c , a ‘saturation genome editing’ CRISPR experiment shows strongly reduced mRNA levels for nonsense mutations in BRCA1, except for those introduced into regions covered by the start-proximal (top) and last-exon NMD evasion rules (bottom panel).

Article Snippet: We therefore used the CRISPR design tools E-CRISP and ‘CRISPRko’ offered by the Genetic Perturbation Platform of the Broad Institute, to design sgRNAs for knock-out experiments of the top 100 most cited genes (from http://doi.org/10.5281/zenodo.1066066 ).

Techniques: Expressing, Knock-Out, MANN-WHITNEY, CRISPR

Journal: Nature genetics

Article Title: The impact of nonsense-mediated mRNA decay on genetic disease, gene editing and cancer immunotherapy

doi: 10.1038/s41588-019-0517-5

Figure Lengend Snippet: a , fitness loss upon targeting a non-essential gene (left) versus an essential gene (right) using a sgRNA directed at gene sections which are covered by various NMD-evasion rules. b - e , distribution of loci targeted by sgRNAs that are NMD-detected or NMD-evading (according to the individual NMD rules) for genome-wide CRISPR libraries ( b , c ) or by sgRNA design tools ( d , e ).

Article Snippet: We therefore used the CRISPR design tools E-CRISP and ‘CRISPRko’ offered by the Genetic Perturbation Platform of the Broad Institute, to design sgRNAs for knock-out experiments of the top 100 most cited genes (from http://doi.org/10.5281/zenodo.1066066 ).

Techniques: Genome Wide, CRISPR