id1 expression plasmid  (Addgene inc)

Journal: bioRxiv

Article Title: Increases in BCL2L1 and ID1 dosage synergistically drive fate bias and competitive advantage in human pluripotent stem cells

doi: 10.64898/2026.03.26.714405

Figure Lengend Snippet: Effects of BCL2L1 and ID1 overexpression on hESC differentiation. (B) Quantitative RT-PCR analysis of BCL2L1 and ID1 expression levels in WT, 20q gain, and hESC lines overexpressing BCL2L1, ID1, or both (BCL2L1/ID1). (C) Immunofluorescence staining of the VUB03 cell line: WT, 20q gain, BCL2L1-OE, ID1-OE, and double (BCL2L1 + ID1) overexpression lines. The final condition represents wild-type cells treated with SB and BMP4. Stained markers include PAX6 (yellow; neuroectoderm, panel 1), TFAP2A (magenta; surface ectoderm, panel 2), KRT8 (turquoise; surface ectoderm, panel 2), EPCAM (turquoise; surface ectoderm, panel 3), and OCT4 (magenta; pluripotency, panel 3). (D) Quantification of neuroectoderm immunostaining of BCL2L1-OE, ID1-OE, and double (BCL2L1 + ID1) overexpression lines showing marker intensity similar to . (E) Immunofluorescence staining of VUB03 cells after 14 days of spontaneous differentiation. Stained markers include PAX6 (yellow; neuroectoderm, panel 1), TFAP2A (magenta; surface ectoderm, panel 2), OCT4 (turquoise; pluripotency, panel 2), EPCAM (turquoise; surface ectoderm, panel 3), and KRT8 (magenta; surface ectoderm, panel 3). (F) Quantification of immunostaining of BCL2L1-OE, ID1-OE, and double (BCL2L1 + ID1) overexpression lines after spontaneous differentiation showing marker intensity similar to . (G) Fraction of marker-positive cells after 8 days of neuroectoderm differentiation. (H) Fraction of marker-positive cells after 14 days of spontaneous differentiation.

Article Snippet: The ID1 expression plasmid (TFORF2860) was obtained from Addgene (plasmid #143643), with ID1 expression driven by the EF-1α promoter.

Techniques: Over Expression, Quantitative RT-PCR, Expressing, Immunofluorescence, Staining, Immunostaining, Marker

Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

Article Title: Inhibitor of Differentiation 1 (ID1) Facilitates the Efficacy of Sorafenib in Non-Small Cell Lung Cancer Cells through Suppressing Epithelial to Mesenchymal Transition

doi: 10.12659/MSM.922148

Figure Lengend Snippet: Sorafenib inhibits ID1 expression. ( A ) Response of cells to sorafenib detected by MTT assay. The average of 3 independent experiments and SD are shown. ( B ) H460 cells were treated with different concentrations of sorafenib. Real-time quantitative PCR was performed to determine the alterations in mRNA expression of ID1, PDGFR, and EGFR. ** P < 0.01; *** P <0.001. ( C ) H460 cells were treated with different concentrations of sorafenib. Western blot experiments were carried out to determine the alteration in ID1 protein expression. ( D ) Immunofluorescence staining for ID1 (red) was conducted in sorafenib-treated H460 cells, and merged images were obtained. DAPI fluorescence is shown in blue. ID1 – inhibitor of differentiation 1; MTT – 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium; SD – standard deviations; PCR – polymerase chain reaction; mRNA – messenger RNA; PDGFR – platelet-derived growth factor receptors; EGFR – epidermal growth factor receptor; DAPI – 4′,6-diamidino-2-phenylindole.

Article Snippet: ID1 over-expression plasmid pcDNA3-ID1 was purchased from Addgene.

Techniques: Expressing, MTT Assay, Real-time Polymerase Chain Reaction, Western Blot, Immunofluorescence, Staining, Fluorescence, Polymerase Chain Reaction, Derivative Assay

Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

Article Title: Inhibitor of Differentiation 1 (ID1) Facilitates the Efficacy of Sorafenib in Non-Small Cell Lung Cancer Cells through Suppressing Epithelial to Mesenchymal Transition

doi: 10.12659/MSM.922148

Figure Lengend Snippet: ID1 downregulation enhances sorafenib efficacy. ( A, B ) H460 cells were transfected with siRNAs targeting ID1. Following treatment with various concentrations of sorafenib for 24 hours ( A ) and 48 hours ( B ), the cell survival rate was determined by MTT assay. ( C, D ) H358 cells were transfected with ID1 overexpression plasmids. Following treatment with various concentrations of sorafenib for 24 hours ( C ) and 48 hours ( D ), the cell survival rate was determined by MTT assay. ID1 – inhibitor of differentiation 1; siRNAs – small interfering RNAs; MTT – 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium.

Article Snippet: ID1 over-expression plasmid pcDNA3-ID1 was purchased from Addgene.

Techniques: Transfection, MTT Assay, Over Expression

Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

Article Title: Inhibitor of Differentiation 1 (ID1) Facilitates the Efficacy of Sorafenib in Non-Small Cell Lung Cancer Cells through Suppressing Epithelial to Mesenchymal Transition

doi: 10.12659/MSM.922148

Figure Lengend Snippet: MG132 inhibits the effect of sorafenib on ID1. ( A ) H460 cells were incubated with various concentrations of sorafenib for 24 hours, with or without pretreatment with MG132. Western blotting was performed to determine the alterations in ID1 expression. ( B, C ) H460 cells transfected with siRNAs targeting ID1 were incubated with various concentrations of sorafenib, with or without pretreatment with MG132. The MTT assay was used to determine the cell survival rate. ID1 – inhibitor of differentiation; siRNAs – small interfering RNAs; MTT – 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium.

Article Snippet: ID1 over-expression plasmid pcDNA3-ID1 was purchased from Addgene.

Techniques: Incubation, Western Blot, Expressing, Transfection, MTT Assay

Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

Article Title: Inhibitor of Differentiation 1 (ID1) Facilitates the Efficacy of Sorafenib in Non-Small Cell Lung Cancer Cells through Suppressing Epithelial to Mesenchymal Transition

doi: 10.12659/MSM.922148

Figure Lengend Snippet: ID1 expression was negatively correlated with EMT biomarkers. ( A ) The pictographs show the morphology of H460, A549, and H358 cells. ( B ) Western blot analysis of EMT markers (E-cadherin and vimentin) and ID1 in H460, A549, and H358 cells. The intensity of the bands was quantified using ImageJ software and normalized to GAPDH ( right panel ). * P <0.05; ** P <0.01; *** P <0.001. ( C ) Immunofluorescence was used to detect the fluorescence intensity of E-cadherin, vimentin and ID1 in H460 and H358 cells. ID1 – inhibitor of differentiation 1; EMT – epithelial to mesenchymal transition.

Article Snippet: ID1 over-expression plasmid pcDNA3-ID1 was purchased from Addgene.

Techniques: Expressing, Western Blot, Software, Immunofluorescence, Fluorescence

Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

Article Title: Inhibitor of Differentiation 1 (ID1) Facilitates the Efficacy of Sorafenib in Non-Small Cell Lung Cancer Cells through Suppressing Epithelial to Mesenchymal Transition

doi: 10.12659/MSM.922148

Figure Lengend Snippet: ID1 suppresses EMT in NSCLC. ( A ) Western blot analysis for the expression of EMT-related markers in H460 cells with a negative control and siRNAs targeting ID1 ( left panel ). Western blot analysis for the expression of EMT-related markers in H358 cells with the vector alone as the control and ID1 overexpression plasmids ( right panel ). ( B ) Representative images of wound healing in H460 cells with negative control and siRNAs targeting ID1 are shown in the left panel . Representative images of wound healing in H358 cells with the vector alone as the control and ID1 overexpression plasmids are shown in the right panel . ID1 – inhibitor of differentiation 1; EMT – epithelial to mesenchymal transition; NSCLC – non-small cell lung cancer; siRNAs – small interfering RNAs.

Article Snippet: ID1 over-expression plasmid pcDNA3-ID1 was purchased from Addgene.

Techniques: Western Blot, Expressing, Negative Control, Plasmid Preparation, Control, Over Expression

Journal: PLoS Biology

Article Title: Olig2 and Hes regulatory dynamics during motor neuron differentiation revealed by single cell transcriptomics

doi: 10.1371/journal.pbio.2003127

Figure Lengend Snippet: (A) Identification of an evolutionarily conserved element containing an E-box in the vicinity of the Hes5 genomic locus in chick, mouse, and human (Hes5(e1)). (B) Analysis of Olig2 Chip-Seq data from reveals Olig2 binding sites in the vicinity of the Hes1 and Hes5 genes. The peak corresponding to the Hes5(e1) element is highlighted in red. (C) Electrophoretic mobility shift assays show that both Olig2 and E12 homodimers can individually bind to the Hes5(e1) E-box and do not form any heterodimeric complexes (lanes 1–4). Positions of the different protein complexes are indicated by colored arrows. Binding depends on the E-box, as both proteins fail to bind probes containing an E-box mutation (Hes5(e1ΔE)) (lanes 5–7). Olig2 binding to Hes5(e1) can be abolished by the addition of unlabelled Hes5(e1) probes, but not those containing the E-box mutation (lanes 8–14). (D) Id1 inhibits binding of E12, but not of Olig2 or Ngn2, to the Hes5(e1) element. Olig2, E12, and Ngn2 alone or Ngn2/E12 heterodimers can bind the Hes5(e1) element. Mixing Olig2 or Ngn2 with Id1 does not inhibit their homodimeric binding activities (lanes 2, 5, 8, and 10). In contrast, Id1 strongly inhibits binding of both E12/E12 and Ngn2/E12 complexes (lanes 6 and 10). The addition of E12 without and with Id1 does not affect Olig2 binding efficiency (lanes 2, 4, and 7). ATG, translational initation codon; Chip-Seq, chromatin immunoprecipitation-sequence; E-box, bHLH transcription factor binding site; N2, Ngn2 protein; O2, Olig2 protein.

Article Snippet: pCS2+ plasmid expression vectors for Olig2 , E12 , Ngn2 , and Id1 [ ] were transcribed and translated in vitro using the Promega TNT Coupled Wheat Germ Extract System.

Techniques: ChIP-sequencing, Binding Assay, Electrophoretic Mobility Shift Assay, Mutagenesis, Chromatin Immunoprecipitation, Sequencing

Journal: bioRxiv

Article Title: Olig2 and Hes regulatory dynamics during motor neuron differentiation revealed by single cell transcriptomics

doi: 10.1101/104307

Figure Lengend Snippet: (A) Identification of an evolutionary conserved element containing an E-box in the vicinity of the Hes5 genomic locus in chick, mouse and human (Hes5(e1)). (B) Analysis of Olig2 Chip-Seq data from reveals Olig2 binding sites in the vicinity of the Hes1 and Hes5 genes. The peak corresponding to the Hes5(e1) element is highlighted in red. (C) Electrophoretic mobility shift assays show that both Olig2 and E12 homodimers can individually bind to the Hes5(e1) E-box, and do not form any heterodimeric complexes (lanes 1-4). Positions of the different protein complexes are indicated by colored arrows. Binding depends on the E-box as both proteins fail to bind probes containing an E-box mutation (Hes5(e1ΔE) (lanes 5-7). Olig2 binding to Hes5(e1) can be abolished by the addition of unlabelled Hes5(e1) probes, but not those containing the E-box mutation (lanes 8-14). (D) Id1 inhibits binding of E12, but not of Olig2 or Ngn2, to the Hes5(e1) element. Olig2, E12 and Ngn2 alone or Ngn2/E12 heterodimers can bind the Hes5(e1) element. Mixing Olig2 or Ngn2 with Id1 does not inhibit their homodimeric binding activities (lanes 2, 5, 8, and 10). In contrast, Id1 strongly inhibits binding of both E12/E12 and Ngn2/E12 complexes (lanes 6 and 10). The addition of E12 without and with Id1 does not affect Olig2 binding efficiency (lanes 2, 4, and 7).

Article Snippet: pCS2+ plasmid expression vectors for Olig2 , E12 , Ngn2 , and Id1 ( ) were transcribed and translated in vitro using the Promega TNT Coupled Wheat Germ Extract System.

Techniques: ChIP-sequencing, Binding Assay, Electrophoretic Mobility Shift Assay, Mutagenesis