empty vector control (lightswitch 3 ́utr reporter vector  (SwitchGear Genomics)

Journal: bioRxiv

Article Title: Role of a childhood cancer-linked BRIP1/FANCJ germline variant in genomic instability and cancer cell vulnerability

doi: 10.64898/2026.03.24.714005

Figure Lengend Snippet: a, HCT116 cells stably expressing BRIP1 R162Q were transiently transfected using PEI with empty vector (EV-EGFP) or RNaseH1-WT-eGFP. Transfection efficiency was assessed via EGFP fluorescence. b, immunoblot analysis confirming expression of GFP-tagged constructs in transfected cells. c, Immunofluorescence detection of R-loops using the S9.6 antibody in cells transfected with EGFP or EGFP-RNaseH1. d, Quantification of S9.6 mean fluorescence intensity (MFI) per nucleus. Data represent mean ± SEM of three independent experiments. e, Immunofluorescence detection of G-quadruplex (G4) structures using the BG4 antibody in cells transfected with EGFP or EGFP-RNaseH1. f, Quantification of BG4 fluorescence intensity per nucleus. Data represent mean ± SEM of three independent experiments. g, Representative DNA fiber assay images from HCT116 cells harboring BRIP1 R162Q transfected with empty vector (EV-eGFP) or RNaseH1-WT. DNA replication tracts were sequentially labeled with CldU (red) and IdU (green). h, Quantification of DNA fiber types. RNaseH1-WT expression reduced the proportion of stalled replication forks in cells harboring BRIP1 R162Q. i, While the overall fork speed was not significantly increased upon RNaseH1-WT expression, tracts displayed a more balanced distribution of CldU and IdU incorporation. j, Fork asymmetry analysis. RNaseH1-WT expression reduced fork asymmetry, indicating that left- and right-moving sister forks progressed at more similar speeds. Labeled tracks were visualized by confocal microscopy, and tract lengths were measured on raw LSM images. Lengths (µm) were converted to replication speed (kb/min) using a factor of 2.59 kb/µm. All data represent mean ± SEM from three independent experiments. At least 150 fibers per condition were analyzed in each replicate. Statistical significance was determined using paired two-tailed Student’s t-test (*p < 0.05, **p < 0.01, ***p < 0.001, ns = not significant). c, e, Representative confocal images are shown.

Article Snippet: For R-loop removal, cells were transiently transfected with an EGFP empty vector control (pEGFP-C1, Takara Bio, #632470), EGFP-tagged RNaseH1 wild-type (WT), or the catalytically inactive RNaseH1 D145N mutant as indicated.

Techniques: Stable Transfection, Expressing, Transfection, Plasmid Preparation, Fluorescence, Western Blot, Construct, Immunofluorescence, Labeling, Confocal Microscopy, Two Tailed Test

Journal: Cell Biology and Toxicology

Article Title: Endothelial progenitor cell derived extracellular vesicles promotes wound healing in diabetic mice via activating mobilization and neovascularization

doi: 10.1007/s10565-025-10134-3

Figure Lengend Snippet: Overexpression of SNHG1 or HDAC6 partially reverses the pro-angiogenic effect of EPC-EVs on MAECs. MAECs were transfected with SNHG1 overexpression vectors (oe-SNHG1), HDAC6 overexpression vectors (oe-HDAC6), or empty vectors (oe-NC, control) in HG (30 mM) medium. A : Transfection efficiency of oe-SNHG1 and oe-HDAC6 was verified by qRT-PCR (n = 3). Cells were then co-treated with EPC-EVs-miR in HG medium. B : SNHG1 and HDAC6 expression was detected by qRT-PCR (n = 3). C : Expression of HDAC6, CD31, and VEGFA was measured by Western blot (n = 3). D : Cell viability was assessed by CCK-8 assay (n = 3). E : Angiogenic capacity was evaluated by tube formation assay (n = 3). F : Cell migration was measured by scratch wound healing assay (n = 3). Data are presented as mean ± standard deviation. Data in panel A were analyze using student’s t-test. Data in panels B , D , E , and F were analyzed using one-way ANOVA. Data in panel C were analyzed using two-way ANOVA, followed by Tukey’s multiple comparisons test. * p < 0.05 and ** p < 0.01

Article Snippet: For lentiviral infection, GFP-labeled miR-204-5p lentiviral overexpression vectors, SNHG1 lentiviral overexpression vectors, or control empty vectors (NC) (Genechem, Shanghai, China) were applied to transfection with 293 T cells (CRL-1573; ATCC, Manassas, VA, USA).

Techniques: Over Expression, Transfection, Control, Quantitative RT-PCR, Expressing, Western Blot, CCK-8 Assay, Tube Formation Assay, Migration, Wound Healing Assay, Standard Deviation