Journal: Current biology : CB

Article Title: Centriole-less pericentriolar material serves as a microtubule organizing center at the base of C. elegans sensory cilia

doi: 10.1016/j.cub.2021.03.022

Figure Lengend Snippet: Key Resources Table

Article Snippet: Plasmid: L4440: RNAi empty vector , Addgene , Cat#1654.

Techniques: Recombinant, Mutagenesis, CRISPR, Sequencing, Plasmid Preparation, Software

Journal: The Journal of Biological Chemistry

Article Title: Ingestion of single guide RNAs induces gene overexpression and extends lifespan in Caenorhabditis elegans via CRISPR activation

doi: 10.1016/j.jbc.2022.102085

Figure Lengend Snippet: Components for CRISPR activation in C. elegans by bacterial delivery of sgRNAs. A , domain organization of the dCas9::VP64 fusion protein, including two nuclear localization signals (NLSs), and a FLAG- and HA-tag. B , schematic representation of the region upstream of a given transcription start site (TSS) from which promoter-specific sgRNAs are selected. C , full sequence of the two sgRNA expression cassettes in vector L4440_BioBrick-sgRNA, including BioBrick cloning sites and with individual features as indicated by color. D , Western blot analysis of total protein extracts from WT N2 nematodes versus dCas9::VP64 nematodes outcrossed x0, x2, or x4 (MIR249). The 171 kDa dCas9::VP64 protein (dC9V) is detected with a FLAG antibody in all dCas9::VP64 nematodes and an additional unspecific (unsp.) band is detected in all samples at 25 kDa. E – G , lifespan assay of WT versus dCas9::VP64 nematodes on OP50 bacteria ( E ), HT115 bacteria carrying the L4440 vector ( F ), or HT115 sgRNA scramble control (SCR) bacteria ( G ). p -values of C. elegans lifespan assays were determined by log-rank test. See Table S1 for detailed lifespan assay statistics. HA, hemagglutinin; sgRNA, single-guide RNA.

Article Snippet: For the expression of C. elegans promoter-specific sgRNAs in E . coli HT115 bacteria, the L4440 RNAi empty vector (Addgene; plasmid #1654) was modified to contain two consecutive sgRNA expression cassettes ( , ) under control of individual T7 promoters and together flanked by BioBrick cloning sites ( ) ( C ), resulting in the vector L4440_BioBrick-sgRNA.

Techniques: CRISPR, Activation Assay, Sequencing, Expressing, Plasmid Preparation, Clone Assay, Western Blot