Journal: iScience

Article Title: Widespread ribosome stalling in a genome-reduced bacterium and the need for translational quality control

doi: 10.1016/j.isci.2021.102985

Figure Lengend Snippet:

Article Snippet: Briefly, approximately 10 8 cells and 2 μg of DNA were incubated for 15 min in ice-cold electroporation buffer (8 mM HEPES·Na pH 7.2, 272 mM sucrose) in a 1-mm gapped electroporation cuvette (Bio-Rad).

Techniques: Virus, Recombinant, Protease Inhibitor, Mutagenesis, Bicinchoninic Acid Protein Assay, Sample Prep, RNA Sequencing, Software, Electroporation, Sequencing, Imaging, Sonication, Mass Spectrometry

Journal: Journal of Virology

Article Title: Stable Cell Clones Harboring Self-Replicating SARS-CoV-2 RNAs for Drug Screen

doi: 10.1128/jvi.02216-21

Figure Lengend Snippet: Initial design of SARS-CoV-2 replicons. (A) Top, genome organization of SARS-CoV-2. Leader sequence (red), transcriptional regulatory sequence within the leader sequence (TRS-L) and within the body (TRS-B) are highlighted in green. Bottom, the design of SARS-CoV-2-Rep-NanoLuc-Neo. (B, C, D, E) Replication kinetics of SARS-CoV-2-Rep-NanoLuc-Neo in Vero E6 (B), A549 (C), Huh7.5.1 (D), and BHK-21 cells (E). Nano luciferase was measured at indicated time points post-electroporation. Notably, nanoluciferase continued to decrease even under G418 selection. (F) Generation of BHK21 stable cells that express NP in a doxycycline-inducible manner. Cells were induced with 0.5 μg/mL doxycycline and lysed at 48 h postinduction for Western blotting with anti-NP and anti-actin antibodies. Numbers on the left refer to the positions of marker proteins that are given in kilodalton (kDa).

Article Snippet: Cells were pelleted by centrifugation (1,000 × g for 5 min at 4°C), washed twice with ice-cold DMEM, and resuspended in ice-cold Gene Pulser Electroporation Buffer (Bio-Rad) at 1 × 10 7 cells/mL.

Techniques: Sequencing, Luciferase, Electroporation, Selection, Western Blot, Marker