Journal: iScience

Article Title: PERIOD 2 regulates low-dose radioprotection via PER2/pGSK3β/β-catenin/Per2 loop

doi: 10.1016/j.isci.2022.105546

Figure Lengend Snippet:

Article Snippet: Eagle’s MEM , Corning CellGro , Cat# 10-010-CV.

Techniques: Western Blot, Immunoprecipitation, Recombinant, Luciferase, Transfection, Library Quantification, ATP Assay, Bicinchoninic Acid Protein Assay, Software, Control, Sequencing, Plasmid Preparation, Gene Expression, Microscopy

Journal:

Article Title: Identification of a Novel Virulence Factor in Recombinant Pneumonia Virus of Mice ▿

doi: 10.1128/JVI.00364-07

Figure Lengend Snippet: rPVM-GFP7 viruses with mutations in the G gene. (A) Schematic diagrams (not drawn to scale) of the G genes of the biologically derived virulent and nonpathogenic isolates of PVM strain 15 (23, 34). Open rectangles, ORFs. The amino acid length of the encoded protein is given in each rectangle. Translational start (filled triangles pointing right) or stop (filled vertical bars) codons are identified by their nucleotide positions in the respective G gene. The GS and GE signals are indicated, and nontranslated gene regions are shown as thin horizontal lines. The nucleotide sequence (negative sense) of positions 166 to 190 of the G gene is given, and the inserted U residue that results in a frameshift is shown in parentheses. The complement of the translational stop codon (nucleotides 188 to 190) that is accessed by the frameshift is italicized, and the complement to the alternative start codon at nucleotides 183 to 185 (Met-34) is boldfaced. (B) Schematic representations (not drawn to scale) of rPVM-GFP-ΔG and rPVM-GFP-Gt. The diagram in the middle represents the rPVM-GFP7 genome (filled box, GFP gene). The box above the diagram illustrates the deletion of the entire G gene to create rPVM-GFP-ΔG. The nucleotide sequence (negative sense) shows the GFP/F gene junction and the AgeI site (italicized) left following the 1,350-nt deletion. The sequence is numbered according to the sequence of rPVM-GFP7, and the GFP GE and F GS signals are underlined. The box below the diagram illustrates the deletion of the cytoplasmic tail of G to create rPVM-GFP-Gt. The unmodified G gene is depicted as in panel A, with the nucleotide positions of its ends numbered according to the complete rPVM-GFP7 sequence. The two potential translational start codons at positions 83 (Met-1) and 182 (Met-34) relative to the G gene sequence are indicated. The nucleotide sequence (negative sense) shows the G GS signal (underlined), the 167-nt deletion that deletes the cytoplasmic tail, and the UAC triplet at positions 182 to 184 that is the complement of the new translational initiation site (Met-34 in the original G ORF). (C) Characterization of rPVM-GFP7, rPVM-GFP-Gt, and rPVM-GFP-ΔG with respect to the expression of G protein. Vero cell monolayers in six-well plates were infected with serial dilutions of the indicated viruses and incubated at 32°C under a methylcellulose overlay for 5 days. (Upper panels) GFP-expressing foci were photographed without further treatment by using an inverted fluorescence microscope. (Lower panels) This was followed by fixation and staining using a G-specific antiserum as described for the plaque assay procedure. Each pair of upper and lower panels depicts approximately the same field of view. (D) Multistep growth kinetics of rPVM, rPVM-GFP7, rPVM-GFP-ΔG, and rPVM-GFP-Gt in BHK-21 cells. Duplicate monolayers were infected at an input MOI of 0.01 PFU per cell. At the indicated days postinfection, the medium supernatants were harvested and flash frozen, and fresh medium was added. Viral titers were determined by a plaque assay. Means from two independent experiments were used to generate the diagram. Error bars, standard errors of the means.

Article Snippet: BHK-21 cells and the BHK-derived BSR-T7/5 cells ( 9 ) were maintained in Glasgow minimal essential medium (MEM) (Invitrogen GIBCO), and Vero cells (ATCC CCL-81) were maintained in Eagle's MEM (Biochrom), all supplemented with 10% fetal calf serum.

Techniques: Derivative Assay, Sequencing, Residue, Expressing, Infection, Incubation, Fluorescence, Microscopy, Staining, Plaque Assay