plasmid pires hmga1  (Addgene inc)

Journal: Experimental & Molecular Medicine

Article Title: Crucial role of HMGA1 in the self-renewal and drug resistance of ovarian cancer stem cells

doi: 10.1038/emm.2016.73

Figure Lengend Snippet: HMGA1 is highly expressed in spheroids from A2780 ovarian cancer cells. ( a ) RT-PCR analysis and western blotting analysis of A2780 adherent cells and spheroid cells are shown in the upper panels with HMGA1 probes along with GAPDH probes. Quantification of HMGA expression by western blotting after normalization using GAPDH expression is shown in the lower panel. * P <0.05. ( b , c ) Flow cytometric analysis and sorting of A2780 spheroid cells after incubation with an ALDH substrate is shown. Western blotting analysis of whole spheroid cells, ALDH − cells and ALDH + cells after sorting is shown with the indicated antibodies.

Article Snippet: pIRES-puro3 vectors overexpressing HMGA1 or non-target controls were purchased from Addgene (Plasmid #13466).

Techniques: Reverse Transcription Polymerase Chain Reaction, Western Blot, Expressing, Incubation

Journal: Experimental & Molecular Medicine

Article Title: Crucial role of HMGA1 in the self-renewal and drug resistance of ovarian cancer stem cells

doi: 10.1038/emm.2016.73

Figure Lengend Snippet: Altering HMGA1 expression changes the expression of stemness-related gene expression. ( a ) Western blotting analysis of A2780 spheroid cells with (sh-HMGA1) or without (sh-Control) knockdown of HMGA1 is shown with the indicated probes. ( b ) Western blotting analysis of HMGA1-overexpressing A2780 cells (HMGA1) or control A2780 cells (Control) is shown with the indicated probes. ( c ) The results of flow cytometric analysis of A2780 spheroid cells with HMGA1 knockdown in comparison with control A2780 spheroid cells (upper panel) or HMGA1-overexpressing A2780 adherent cells in comparison with control A2780 adherent cells (lower panel) after incubation with ALDH substrate are shown.

Article Snippet: pIRES-puro3 vectors overexpressing HMGA1 or non-target controls were purchased from Addgene (Plasmid #13466).

Techniques: Expressing, Gene Expression, Western Blot, Control, Knockdown, Comparison, Incubation

Journal: Experimental & Molecular Medicine

Article Title: Crucial role of HMGA1 in the self-renewal and drug resistance of ovarian cancer stem cells

doi: 10.1038/emm.2016.73

Figure Lengend Snippet: HMGA1 expression regulates the CSC characteristics of A2780 ovarian cancer cells. ( a ) Time course of the cell numbers counted after plating the same number of cells (1 × 10 4 cells per well). A2780 spheroid cells with HMGA1 knockdown are compared with control A2780 spheroid cells (upper panel), and HMGA1-overexpressing A2780 adherent cells are compared with control A2780 adherent cells (lower panel). * P <0.05 ( b ) Bright-field images of spheroids generated from A2780 spheroid cells with or without HMGA1 knockdown (upper panels) and counts of the spheroid numbers (lower panel) are shown. Scale bar, 100 μm. * P <0.05 ( c ) Bright-field images of spheroids generated from HMGA1-overexpressing A2780 adherent cells in comparison with control A2780 adherent cells (upper panels) and counts of the spheroid numbers (lower panel) are shown. Scale bar, 100 μm. * P <0.05.

Article Snippet: pIRES-puro3 vectors overexpressing HMGA1 or non-target controls were purchased from Addgene (Plasmid #13466).

Techniques: Expressing, Knockdown, Control, Generated, Comparison

Journal: Experimental & Molecular Medicine

Article Title: Crucial role of HMGA1 in the self-renewal and drug resistance of ovarian cancer stem cells

doi: 10.1038/emm.2016.73

Figure Lengend Snippet: HMGA1 regulates ABCG2 promoter activity and drug resistance. ( a ) Schematic representations of the −1285 and −628 5′ upstream regions of the ABCG2 promoter are shown. HMGA1-binding sites are marked by boxes. ( b ) The results of a transcriptional activity assay after co-transfecting an ABCG2 promoter construct (−1285 or −628) with an HMGA1-overexpression construct into A2780 cells are shown (upper panel). * P <0.05. The results of western blotting analysis are shown in the lower panel. ( c ) The results of a transcriptional activity assay after co-transfecting an ABCG2 promoter construct (−1285, ΔHMGA1-1, ΔHMGA1-2 or ΔHMGA1-3) with an HMGA1-overexpression construct into A2780 cells are shown (upper panel). * P <0.05. The results of western blotting analysis are shown in the lower panel. ( d ) The results from a time course MTT assay with A2780 spheroid cells with HMGA1 knockdown in comparison with control A2780 spheroid cells (left panel), or with HMGA1-overexpressing A280 cells in comparison with control A2780 adherent cells (right panel) in the presence of paclitaxel (0.1 μ M ), are shown. * P <0.05. ( e ) The results from a time course MTT assay with A2780 spheroid cells with HMGA1 knockdown in comparison with control A2780 spheroid cells (left panel), or with HMGA1-overexpressing A280 adherent cells in comparison with control A2780 adherent cells (right panel) in the presence of doxorubicin (1 μ M ), are shown. * P <0.05.

Article Snippet: pIRES-puro3 vectors overexpressing HMGA1 or non-target controls were purchased from Addgene (Plasmid #13466).

Techniques: Activity Assay, Binding Assay, Construct, Over Expression, Western Blot, MTT Assay, Knockdown, Comparison, Control

Journal: Experimental & Molecular Medicine

Article Title: Crucial role of HMGA1 in the self-renewal and drug resistance of ovarian cancer stem cells

doi: 10.1038/emm.2016.73

Figure Lengend Snippet: HMGA1 overexpression enhances the drug resistance of ovarian cancer cells in a xenograft tumor model. ( a ) Tumor size was measured on the indicated day after the subcutaneous injection of HMGA1-overexpressing A2780 or control A2780 cells into nude mice (1 × 10 5 per injection) with HBSS or paclitaxel treatment ( n =6). Paclitaxel was injected at each data point. * P <0.05. ( b ) Immunohistochemistry results of tumors harvested on day 45. DAPI and the indicated antibodies are shown. Scale bar, 50 μm.

Article Snippet: pIRES-puro3 vectors overexpressing HMGA1 or non-target controls were purchased from Addgene (Plasmid #13466).

Techniques: Over Expression, Injection, Control, Immunohistochemistry