lentivirus packaging mix  (Alstem Inc)

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Truncated tau interferes with the autophagy and endolysosomal pathway and results in lipid accumulation

doi: 10.1007/s00018-024-05337-6

Figure Lengend Snippet: Tau35 expression leads to a decrease in LC3-labelled autophagosomes and autolysosomes. a CHO cells transfected with EGFP-LC3 plasmid (green). Scale bar = 10 µm. (ai) Graph shows the number of EGFP-LC3 puncta per cell area. N = 3 independent experiments (70–100 cells). Kruskal–Wallis test with Dunn’s multiple comparisons test, ****P < 0.0001, **P < 0.005. b Immunofluorescence labelling of CHO cells with an LC3 antibody. Scale bar = 10 µm. (bi) Graph shows the number of LC3 puncta per cell area. N = 3 independent experiments. One-way ANOVA with Tukey’s multiple comparisons test, ****P < 0.0001, **P < 0.005. c CHO cells transduced with mCherry-GFP-LC3 lentivirus. Scale bar = 10 µm. Graphs show the number of (ci) mCherry + GFP + (autophagosomes) and (cii) mCherry + GFP− puncta (autolysosomes) per cell area. N = 3 independent experiments. Kruskal–Wallis test with Dunn’s multiple comparisons test, ****P < 0.0001. d Primary cortical neurons from WT and Tau35 mice, transduced with mCherry-GFP-LC3 lentivirus. Blue arrowheads point to autolysosomes. White arrowheads point to autophagosomes. Scale bar = 10 µm. Graphs show the number of (di) mCherry + GFP + (autophagosomes) and (dii) mCherry + GFP− puncta (autolysosomes) per soma area. N = 3 independent experiments. Unpaired t- test, *P < 0.05

Article Snippet: FUW-mCherry-GFP-LC3 lentivirus was produced by transfection of the plasmid (21 μg DNA) in HEK293T cells with 30 μl TransIT ® lentivirus packaging mix (Mirius, Cambridge Bioscience) in a T-175 flask.

Techniques: Expressing, Transfection, Plasmid Preparation, Immunofluorescence, Transduction

Journal: Heliyon

Article Title: The role and mechanism of IFITM1 in developing acquired cisplatin resistance in small cell lung cancer

doi: 10.1016/j.heliyon.2024.e30806

Figure Lengend Snippet: Silencing of IFITM1 gene in cisplatin resistant SCLC cells and overexpression of IFITM1 in their parent cells by using lentivirus A) IC 50 value analysis of cisplatin-resistant SCLC cells (NCI–H466 and HOP-92) and their parent cells. The concentration of cisplatin was calculated for each cell line at 48h after treatments. B) Lentivirus infection efficiency analysis. At 72 h after infection with Lv-NC, we roughly estimated the infection efficiency of lentivirus to these cells by counting the proportion of cells with GFP expression in all. C) At 72 h after infection, the mRNA levels of IFITM1 were detected by RT-qPCR. β-actin were used as the references. D) After 72 h of infection, the protein expression of IFITM1 were detected by western blotting. β-actin were used as the references respectively. Each experiment was repeated at least for three times(n = 3). Data are presented as means ± SD. * p < 0.05, ** p < 0.01.

Article Snippet: Two micrograms of each vector and 10 μg of lentivirus Packaging Plasmid Mix (System Biosciences, CA, USA) were co-transfected using Lipofectamine 2000 (Invitrogen) in accordance with the manufacturer's protocol.

Techniques: Over Expression, Concentration Assay, Infection, Expressing, Quantitative RT-PCR, Western Blot