qpcr data analysis  (Roche)

Journal: iScience

Article Title: Gene delivery in mosquitos with a vesicular stomatitis virus vector

doi: 10.1016/j.isci.2025.113510

Figure Lengend Snippet: Susceptibility of adult Ae. aegypti mosquitoes to rVSV (A–D) Twelve mosquitoes were microinjected (A and B) or blood-fed (C and D) with increasing doses of rVSV (A and C) Infection rate was determined as the number of infected mosquitoes relative to the total number of mosquitoes ( n = 3). (B and D) The viral titers of individual mosquito were determined by TCID 50 assay at 7 dpi. Data are presented as mean ± SD. (E–H) Tissue preference of rVSV infection in mosquitoes. Six-day-old female adults were blood-fed with 1 × 10 8 FFU/mL rVSV. Total RNAs from eight tissues of fifteen mosquitoes were extracted at 7 dpi, and the viral RNAs were detected by real-time qPCR (E). Total RNAs from midguts (F), ovaries (G), and fat bodies (H) of eight mosquitoes were extracted at different days after infection, and the viral RNA was detected by real-time qPCR. Control (Con.) represents uninfected mosquito groups.

Article Snippet: • All original western blot images, confocal microscopy images, along with processed real-time qPCR analysis data used for this study, have been deposited at Mendeley Data: https://data.mendeley.com/datasets/3pb4vh7hjz/1 .

Techniques: Infection, Control

Journal: iScience

Article Title: Gene delivery in mosquitos with a vesicular stomatitis virus vector

doi: 10.1016/j.isci.2025.113510

Figure Lengend Snippet: The insertion positions of GFP gene affected its expression level (A) Schematic diagram of GFP gene insertion positions. (B) Vero cells were infected with rVSVs at an MOI of 0.1. Western blotting of the cell lysates was performed at 24 h after infection using anti-GFP or anti-GAPDH mouse antibodies. Band quantification was carried out using Image Studio software (Licor), and the numbers indicated the ratio of GFP to GAPDH. (C) Real-time qPCR of GFP expression levels relative to β-actin at 24 h after rVSV infection. Data are presented as mean ± SD of triplicate measurements. Con. represents mock-infected Vero cells. (D) GFP fluorescence in rVSV-infected Vero cells was detected at 24 h after infection under a fluorescence microscope. (F) Mosquitoes were blood-fed with indicated rVSVs, and their midguts were visualized under fluorescence microscopy at 5 dpi. GFP was shown in green, and the nucleus was stained with Hoechst 33342 (blue). (E and G) Quantitative results of the GFP fluorescence intensity in (D) or (F). Kruskal-Wallis test followed by Dunn’s post hoc tests. ∗ p < 0.05, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. ns, not significant. These results are representative of three independent experiments.

Article Snippet: • All original western blot images, confocal microscopy images, along with processed real-time qPCR analysis data used for this study, have been deposited at Mendeley Data: https://data.mendeley.com/datasets/3pb4vh7hjz/1 .

Techniques: Expressing, Infection, Western Blot, Software, Fluorescence, Microscopy, Staining

Journal: iScience

Article Title: Gene delivery in mosquitos with a vesicular stomatitis virus vector

doi: 10.1016/j.isci.2025.113510

Figure Lengend Snippet: Overexpression of FoxO gene in adult mosquitoes did not affect lifespan (A) Schematic diagram of FoxO gene insertion position in the rVSV. (B) The growth curves of rVSV and rVSV-FoxO in Vero cells at an MOI of 0.01 ( n = 3, unpaired t test, two-sided). (C–F) Six-day-old females were blood-fed with rVSV-FoxO or rVSV diluted to 1 × 10 8 FFU/mL. Con. was fed with medium. Total RNAs were extracted from 8 mosquitoes (C), 8 midguts (D), 8 ovaries (E), or 8 fat bodies (F) at indicated dpi, and the FoxO expression was measured by real-time qPCR. Data are presented as mean ± SD of triplicate measurements ( n = 3). (G and H) Ten mosquitoes per group were treated with iFoxO and iGFP via microinjection or infected with rVSV-FoxO (rFoxO) and rVSV via blood feeding. Western blotting of these mosquito homogenates was performed using anti-FoxO or anti-GAPDH antibodies. (I) Band intensities of FoxO in (H) were quantified using LiCor Image Studio and normalized to GAPDH. Results are shown as mean ± SD from three independent experiments ( n = 3 biologically independent mixtures; two-way ANOVA test followed by Tukey’s post hoc tests). (J and K) Relative FoxO mRNA levels of ten mosquitoes at 7 days post treatment (unpaired t test, two-sided). (L and M) Survival curves of mosquitoes per group upon different treatments (Kaplan-Meier curves and log rank test). Error bars in graphs represent the mean ± SD. ∗ p < 0.05, ∗∗∗∗ p < 0.0001. ns, not significant; These results are representative of three independent experiments.

Article Snippet: • All original western blot images, confocal microscopy images, along with processed real-time qPCR analysis data used for this study, have been deposited at Mendeley Data: https://data.mendeley.com/datasets/3pb4vh7hjz/1 .

Techniques: Over Expression, Expressing, Microinjection, Infection, Western Blot

Journal: iScience

Article Title: Gene delivery in mosquitos with a vesicular stomatitis virus vector

doi: 10.1016/j.isci.2025.113510

Figure Lengend Snippet: E93 overexpression shortens pupation time of larvae (A) Schematic diagram of the rVSV carrying E93 gene. (B) The growth curves of rVSV-E93-RB, rVSV-E93-RC, and rVSV after infection of Vero cells at an MOI of 0.01 ( n = 3, one-way ANOVA test). (C) Florescence images of larvae and pupa infected with rVSV and rVSV-E93. (D) Real-time qPCR analysis of E93 transcripts in larvae infected with 10 9 FFU/mL rVSV-E93 or rVSV ( n = 15, unpaired t test, two-sided). Error bars in graphs represent the mean ± SD. (E) Pupation time of the larvae infected with rVSV-E93 or rVSV (unpaired t test, two-sided). Data are presented as mean ± SD of triplicate measurements. ns, not significant; ∗∗ p < 0.01; ∗∗∗∗ p < 0.0001. These results are representative of three independent experiments.

Article Snippet: • All original western blot images, confocal microscopy images, along with processed real-time qPCR analysis data used for this study, have been deposited at Mendeley Data: https://data.mendeley.com/datasets/3pb4vh7hjz/1 .

Techniques: Over Expression, Infection

Journal: iScience

Article Title: Gene delivery in mosquitos with a vesicular stomatitis virus vector

doi: 10.1016/j.isci.2025.113510

Figure Lengend Snippet: Delivery of miR-10 by rVSV reduced the susceptibility of adult mosquitoes to ZIKV (A) The growth curves of rVSV-miR-10 and rVSV after infection of Vero cells at an MOI of 0.01 ( n = 3). (B and C) Mosquitoes were microinjected with anta-miR-10, antagomir-negative control (anta-NC), or rVSV-miR-10, rVSV within 24 h post eclosion. The RNA levels of miR-10 were examined by real-time qPCR. (D and E) Mosquitoes were blood-fed with ZIKV after microinjection with antagomirs or rVSVs. ZIKV RNA levels were measured using real-time qPCR at 7 dpi ( n ≥ 12). Percentage represents the mosquito infection rate. Data are shown as mean ± SD. The figure illustrated one of three representative independent experiments. The experiments were repeated three times with similar results. ns, not significant; ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001 (unpaired t test, two-sided). These results are representative of three independent experiments.

Article Snippet: • All original western blot images, confocal microscopy images, along with processed real-time qPCR analysis data used for this study, have been deposited at Mendeley Data: https://data.mendeley.com/datasets/3pb4vh7hjz/1 .

Techniques: Infection, Negative Control, Microinjection

Journal: International journal of food microbiology

Article Title: Presence and characterization of the human pathogenic Vibrio species in the microbiota of Manila clams using cultural and molecular methods.

doi: 10.1016/j.ijfoodmicro.2025.111113

Figure Lengend Snippet: Fig. 4. qPCR data visualization. (A) Absolute amount of Vibrio spp. in homogenized samples (HO), marine agar (MA), and on thiosulfate-citrate-bile-salts-sucrose agar (TCBS) incubated at 22 ◦C and 37 ◦C with the Y-axis representing the log copies number of cells. (B) Absolute amount of V. parahaemolyticus in Chioggia (two sites, 1CH and 2CH), Porto Marghera (PM), Scardovari (SC), Goro (GO) Marano (MA), and Colmata (CO) with the Y-axis representing the log copies number of cells. (C) Bubble chart showing the samples that resulted quantifiable, positive but not quantifiable, and negative using V. parahaemolyticus, V. vulnificus, and Vibrio spp. primers in qPCR.

Article Snippet: The cycling conditions were as follows: initial incubation at 50 ◦C for 2 min, followed by 2 min at 95 ◦C, and 45 cycles at 95 ◦C for 10 s and 60 ◦C for 40 s. All reactions were performed in duplicate. qPCR data analysis was performed using LightCycler 480 software version 1.5 (Roche).

Techniques: Incubation