Journal: Pathogens

Article Title: A Modular Hepatitis E Virus Replicon System for Studies on the Role of ORF1-Encoded Polyprotein Domains

doi: 10.3390/pathogens11030355

Figure Lengend Snippet: Luciferase activity of four parental HEV replicons in comparison to mock-transfected control (H 2 O) and inactivated RdRp GAA mutant replicon based on p6LucA26. HepG2 cells were transfected with replicon ivtRNA by electroporation and seeded across four wells of a 96-well plate as technical quadruplicate. Replication was estimated by measuring total luciferase activity (relative light units; RLU) of each replicon. Error bars indicate standard deviation and asterisks indicate significance level (*** p ≤ 0.001; ** 0.001 < p ≤ 0.01; ns 0.05 < p ≤ 1). The data depict a representative experiment of three separate transfections.

Article Snippet: HepG2 cells were purchased from CLS (Eppelheim, Germany) and grown in DMEM (Gibco 52100, ThermoFisher, Waltham, MA, USA), supplemented with NaHCO 3 and Na-pyruvate, and 10% fetal bovine serum, including 10 mg/L gentamicin (PAN-Biotech, Aidenbach, Germany), and 250 mg/L amphotericin B (PAN-Biotech, Aidenbach, Germany) in cell culture flasks with vented caps (Corning, Corning, NY, USA) in a humidified incubator with 5% CO 2 .

Techniques: Luciferase, Activity Assay, Comparison, Transfection, Control, Mutagenesis, Electroporation, Standard Deviation

Journal: Pathogens

Article Title: A Modular Hepatitis E Virus Replicon System for Studies on the Role of ORF1-Encoded Polyprotein Domains

doi: 10.3390/pathogens11030355

Figure Lengend Snippet: ( A ) Structure of nine chimeric replicons that were constructed using p6LucA26 as backbone. Subgenomic fragments containing partial sequences of ORF1 (MYP, nucleotide positions 1–2139 in p6; VXH, nucleotide positions 2144–4059 in p6; RJ, nucleotide positions 4064–5347 in p6) were replaced with the corresponding sequences from HEV strains 47832mc, rab52, and rab81. Percentages annotated in the highlighted fragments represent pairwise amino acid sequence identity of the encoded proteins with the corresponding ones encoded by parental p6 replicon. Multiple amino acid sequence alignments of each ORF1-encoded protein segment are included in the . ( B ) HepG2 cells were transfected with replicon ivtRNA by electroporation and seeded across four wells of a 96-well plate as technical quadruplicates. Replication was estimated by measuring total luciferase activity (RLU) of each replicon. Error bars indicate standard deviation; asterisks indicate significance level in reference to p6LucA26 (**** p ≤ 0.0001; *** 0.0001 < p ≤ 0.001; ** 0.001 < p ≤ 0.01; * 0.01 < p ≤ 0.05; ns 0.05 < p ≤ 1). The data depict a representative experiment of three separate transfections. Note that the controls (p6LucA26, p6GAALucA26, and mock) are shown in each plot for illustrative purposes. The data depicted are derived from a single experiment with all chimeric replicons and one group of controls.

Article Snippet: HepG2 cells were purchased from CLS (Eppelheim, Germany) and grown in DMEM (Gibco 52100, ThermoFisher, Waltham, MA, USA), supplemented with NaHCO 3 and Na-pyruvate, and 10% fetal bovine serum, including 10 mg/L gentamicin (PAN-Biotech, Aidenbach, Germany), and 250 mg/L amphotericin B (PAN-Biotech, Aidenbach, Germany) in cell culture flasks with vented caps (Corning, Corning, NY, USA) in a humidified incubator with 5% CO 2 .

Techniques: Construct, Sequencing, Transfection, Electroporation, Luciferase, Activity Assay, Standard Deviation, Derivative Assay