Journal: Vaccines

Article Title: Development of a Novel Live Attenuated Influenza A Virus Vaccine Encoding the IgA-Inducing Protein

doi: 10.3390/vaccines9070703

Figure Lengend Snippet: In vitro growth kinetics. ( A ) Alignment of the predicted IGIP in different mammalian species. The mature swine IGIP sequence used in this study is shown. ( B ) Schematic representation of the IGIP-H1 gene. The IGIP sequence and the components of the intergenic region are indicated. ( C ) Growth kinetics profiles of IGIP-H1att and H1att in MDCK and MDCK STAT1 KO cells. Experiments were performed two times independently, each time in triplicate. Titers were determined by RT-qPCR and expressed as log 10 TCID50 equivalents. Gray area represents the area below the level of detection of the assay. ( D ) IGIP-H1att virus was serially passaged five times in MDCK (E1C5) cells and SPF eggs (E5), and the HA was amplified by RT-PCR, showing that the IGIP-H1 rearrangement is stable. Abbreviations: FP, fusion peptide; TM, transmembrane domain; CT, c-terminal region; G4S, poly-glycine protein linker; Furin CS, furin cleavage site; Tav2A, Thosea assigna virus 2A protein sequence; GlucS, Signal peptide of Gaussia luciferase.

Article Snippet: MDCK STAT1 KO cells (CCL-34-VHG) were purchased from ATCC.

Techniques: In Vitro, Sequencing, Quantitative RT-PCR, Virus, Amplification, Reverse Transcription Polymerase Chain Reaction, Luciferase

Journal: Cell Proliferation

Article Title: RNAi‐mediated inhibition of the desmosomal cadherin (desmoglein 3) impairs epithelial cell proliferation

doi: 10.1111/j.1365-2184.2011.00765.x

Figure Lengend Snippet: Modulation of Dsg3 expression affected cell proliferation in MDCK cells. (a) Western blotting of cell lysates extracted from cells with either overexpression (knock‐in: KI) or knockdown (KD) of Dsg3. (b) Confocal microscopy of MDCK cells with ectopic Dsg3 expression [myc‐tag in (i)] or with Dsg3 knockdown (iii). Images in (ii) were vector control cells. (c) Dispase fragmentation assay of MDCK cells with either vector or Dsg3 shRNAi transduction. Cells were grown to confluence before being treated with 2.4 units/ml dispase for about 30 min to detach epithelial sheets. These sheets were subjected to mechanical stress by pipetting five times with 1 ml tips. Epithelial fragments were quantified by ImageJ and increased fragments (up to 2‐fold) were seen in cells with Dsg3 silencing by shRNAi‐1 or shRNAi‐2, respectively. Data are averages of duplicates in each group. (d) Growth curve of matched MDCK cells with up‐ or down‐regulation of Dsg3. Cells with overexpression had higher proliferation, but those with Dsg3 knockdown exhibited lower growth rate compared to matched control cells.

Article Snippet: HuSH TM shRNA plasmids (29‐mer) used for transduction in MDCK cells were purchased from OriGene Technologies Inc (Rockville, MD, USA).

Techniques: Expressing, Western Blot, Over Expression, Knock-In, Confocal Microscopy, Plasmid Preparation, Transduction