Journal: eLife

Article Title: Yap1 safeguards mouse embryonic stem cells from excessive apoptosis during differentiation

doi: 10.7554/eLife.40167

Figure Lengend Snippet:

Article Snippet: For inducible overexpression (OE), a pLVX-IRES-ZsGreen1 vector (Clontech) containing the gene of interest and pLVX-TRE3G vector (Clontech) were transfected separately with packaging plasmids.

Techniques: Transfection, Recombinant, Knock-Out, LDH Cytotoxicity Assay, SYBR Green Assay, Membrane, Luciferase, Binding Assay, Software, Microscopy

Journal: Cancers

Article Title: LdrB Toxin with In Vitro and In Vivo Antitumor Activity as a Potential Tool for Cancer Gene Therapy

doi: 10.3390/cancers11071016

Figure Lengend Snippet: HCT-116 and MCF-7 cells stably express the ldrB gene after transfection. ( A ) The plasmid pLVX-TRE3G-mCherry-IdrB was identified by XhoI and ApaI digestion. M: DNA Ladder 10,000; 1, undigested circular plasmid; 2, plasmid digested with XhoI and ApaI (expected Size: 1187+8K). ( B ) HCT-116 and MCF-7 cells were transfected with the ldrB gene using Tet-On 3G Inducible Expression System (with mCherry). Stable, transfected cells colored red were selected. In the absence of Dox, protein fluorescence is not observed. After 24 h of Dox administration, red fluorescence emitted by m-cherry through LdrB toxin expression emerged (a, b, and c 20× and e, f, and g 10×). ( C ) Total RNA was extracted from HCT-116, MCF-7, HCT-ldrB, and MCF-ldrB cells. After 24 h of dox induction, ldrB expression levels were analyzed with qRT-PCR. ( D ) Effects of the ldrB gene on HCT-116 and MCF-7 cell lines (a) HCT-116 cells (b) HCT-116-Dox, and (c) HCT-ldrB-Dox after 15 d of culture; (d) MCF-7 cells, (e) MCF-7-Dox, and (f) MCF-ldrB-Dox after 15 d of culture (a, b, c, d, e, f, and g 10×). ( E ) Determination of the effects of ldrB gene expression on HCT-116 and ( F ) MCF-7 cell proliferation. Cells were cultured for 15 d to determine the growth rate. Data were obtained by an EnSight system with well imaging technology. Values represent means ± SD of quadruplicate cultures (*** p < 0.001).

Article Snippet: The ldrB gene flanked by the NdeI site ant 5′ and EcoRI site at 3′ was synthesized and subcloned into pLVX-TRE3G-mCherry (Clontech, Palo Alto, CA, USA) previously digested with the same restriction enzymes.

Techniques: Stable Transfection, Transfection, Plasmid Preparation, Expressing, Fluorescence, Quantitative RT-PCR, Cell Culture, Imaging