Journal: bioRxiv

Article Title: Downregulation of a cell polarity protein potentiates Chikungunya Virus infection in host cells

doi: 10.1101/2023.07.24.550336

Figure Lengend Snippet: HEK293T cells were transfected with pEGFPC1 vector or pEGFPC1 plasmids carrying TF or 6K. Twenty-four hours post-transfection, cells were fixed, and immunostaining was performed for Scribble (red). Inset shows zoomed images of an EGFP-TF expressing cell. Arrowheads indicate Scribble puncta and co-localization with TF (A). Mutational analysis of TF and its effect on Scribble localization (B). Expression data of TF and TF mutants in HEK293T cells. Twenty-four hours post-transfection, cells were lysed, and 6K and TF expression were analyzed using western blot with anti-GFP antibody (C). L: protein ladder, C1: pEGFPC1 empty vector, TF Mut1: TF-HGAATV, TF Mut2: TF-AAAATV.

Article Snippet: The cDNA sequences of CHIKV 6K and TF (Supplementary Figure S1) were cloned in the pEGFPC1 vector (Clontech) to generate fusion proteins tagged with the Enhanced Green Fluorescent Protein (EGFP) at the N-terminus.

Techniques: Transfection, Plasmid Preparation, Immunostaining, Expressing, Western Blot

Journal: bioRxiv

Article Title: Downregulation of a cell polarity protein potentiates Chikungunya Virus infection in host cells

doi: 10.1101/2023.07.24.550336

Figure Lengend Snippet: HEK293T cells were infected with CHIKV at 0.1 MOI. Scribble levels were analyzed at 24 and 48 hpi using western blot and quantitated using densitometry (A). Twenty-four hours post-infection, the cells were fixed, and immunostaining was performed for Scribble and CHIKV capsid and E2 proteins. Inset shows zoomed images of an E2 expressing cell. Arrowheads indicate Scribble puncta (B). Effect of TF on Scribble expression. HEK293T cells were transfected with pEGFPC1-TF and its mutants. Only vector and pEGFPC1-6K were also transfected, and Scribble levels were analyzed 48 h post-transfection using western blot and quantitated using densitometry (C). Electron microscopy (EM) imaging of CHIKV-infected HEK293T cells. HEK293T cells were infected with CHIKV at 1.0 MOI and Scribble was visualized using a gold-labeled secondary antibody (D-i). EM images of an infected cell at higher magnification (D-ii). Immunostaining for ubiquitin and Scribble in CHIKV infected HEK293T cells (E). Data represents mean ± SEM of two biological replicates.

Article Snippet: The cDNA sequences of CHIKV 6K and TF (Supplementary Figure S1) were cloned in the pEGFPC1 vector (Clontech) to generate fusion proteins tagged with the Enhanced Green Fluorescent Protein (EGFP) at the N-terminus.

Techniques: Infection, Western Blot, Immunostaining, Expressing, Transfection, Plasmid Preparation, Electron Microscopy, Imaging, Labeling

Journal: eLife

Article Title: SAFB regulates hippocampal stem cell fate by targeting Drosha to destabilize Nfib mRNA

doi: 10.7554/eLife.74940

Figure Lengend Snippet:

Article Snippet: Recombinant DNA reagent , pEGFPC1-6XHis- FLKSRP (plasmid) , , RRID: Addgene_23001 , pEGFPC1 expression vector.

Techniques: Protease Inhibitor, Recombinant, Clone Assay, Bicinchoninic Acid Protein Assay, Mutagenesis, Transfection, Sequencing, esiRNA, Blocking Assay, Plasmid Preparation, Expressing, Software