Journal: Frontiers in Genetics

Article Title: Integrated Dissection of lncRNA-miRNA-mRNA Pairs and Potential Regulatory Role of lncRNA PCAT19 in Lung Adenocarcinoma

doi: 10.3389/fgene.2021.765275

Figure Lengend Snippet: Prognosis values of differentially expressed long noncoding RNAs in lung adenocarcinoma.

Article Snippet: A549 cells were seeded into 96-well plates and were co-transfected with 60 ng of the dual luciferase reporter construct pmiR-RB-Report ™ -PCAT19 3′UTR (RiboBio, Guangdong) and miR-143-3p mimic.

Techniques:

Journal: Frontiers in Genetics

Article Title: Integrated Dissection of lncRNA-miRNA-mRNA Pairs and Potential Regulatory Role of lncRNA PCAT19 in Lung Adenocarcinoma

doi: 10.3389/fgene.2021.765275

Figure Lengend Snippet: Key lncRNAs, miRNAs, and mRNAs in the ceRNA network of lung adenocarcinoma.

Article Snippet: A549 cells were seeded into 96-well plates and were co-transfected with 60 ng of the dual luciferase reporter construct pmiR-RB-Report ™ -PCAT19 3′UTR (RiboBio, Guangdong) and miR-143-3p mimic.

Techniques: Binding Assay

Journal: Frontiers in Genetics

Article Title: Integrated Dissection of lncRNA-miRNA-mRNA Pairs and Potential Regulatory Role of lncRNA PCAT19 in Lung Adenocarcinoma

doi: 10.3389/fgene.2021.765275

Figure Lengend Snippet: (A) Univariate Cox proportional hazard regression analysis of 14 lncRNAs related to the prognosis of patients with lung adenocarcinoma. (B) Risk score distribution, (C) patient survival status distribution, and (D) heatmap of PCAT19 and SMIM25 expression by the risk score. (E) Kaplan–Meier curves for high-risk and low-risk groups. (F) ROC curves for predicting the survival in lung adenocarcinoma patients by the risk score. (G,H) RT-qPCR analysis was conducted to assess the expression levels of PCAT19 and SMIM25 in (G) lung adenocarcinoma tissues (the control was advanced normal tissues) and (H) A549 and SPC-A1 cells (the control was 16HBE cells). The expression level of GAPDH was used as an internal reference, and the data are shown as the mean ± standard deviation. *** p < 0.001.

Article Snippet: A549 cells were seeded into 96-well plates and were co-transfected with 60 ng of the dual luciferase reporter construct pmiR-RB-Report ™ -PCAT19 3′UTR (RiboBio, Guangdong) and miR-143-3p mimic.

Techniques: Expressing, Quantitative RT-PCR, Standard Deviation

Journal: Frontiers in Genetics

Article Title: Integrated Dissection of lncRNA-miRNA-mRNA Pairs and Potential Regulatory Role of lncRNA PCAT19 in Lung Adenocarcinoma

doi: 10.3389/fgene.2021.765275

Figure Lengend Snippet: (A) Correlation between overall survival and multivariable characteristics via multivariate survival analysis of PCAT19 and (B) multivariate survival analysis of SMIM25 and PCAT19.

Article Snippet: A549 cells were seeded into 96-well plates and were co-transfected with 60 ng of the dual luciferase reporter construct pmiR-RB-Report ™ -PCAT19 3′UTR (RiboBio, Guangdong) and miR-143-3p mimic.

Techniques:

Journal: Frontiers in Genetics

Article Title: Integrated Dissection of lncRNA-miRNA-mRNA Pairs and Potential Regulatory Role of lncRNA PCAT19 in Lung Adenocarcinoma

doi: 10.3389/fgene.2021.765275

Figure Lengend Snippet: (A) Correlation analysis of the expression of PCAT19 with clinicopathological parameters of lung adenocarcinoma. (B) Radar chart shows the correlation between the expression of PCAT19 and the tumor mutational burden of each tumor. The red font represents lung adenocarcinoma; (C,D) KEGG pathway analysis showed (C) positively correlated groups and (D) negatively correlated groups; (E,F) GO term analysis revealed (E) positively correlated groups and (F) negatively correlated groups.

Article Snippet: A549 cells were seeded into 96-well plates and were co-transfected with 60 ng of the dual luciferase reporter construct pmiR-RB-Report ™ -PCAT19 3′UTR (RiboBio, Guangdong) and miR-143-3p mimic.

Techniques: Expressing

Journal: Frontiers in Genetics

Article Title: Integrated Dissection of lncRNA-miRNA-mRNA Pairs and Potential Regulatory Role of lncRNA PCAT19 in Lung Adenocarcinoma

doi: 10.3389/fgene.2021.765275

Figure Lengend Snippet: (A) Overexpression of PCAT19 in A549 cells analyzed by fluorescence observation; (B) transfection efficiency of pCDH-PCAT19 and si-PCAT19 was detected by qPCR; (C) effects of PCAT19 knockdown or overexpression on the vitality of A549 cells measured using the CCK-8 assay; (D) effects of PCAT19 knockdown or overexpression on the vitality of SPC-A1 cells measured using the CCK-8 assay; (E–G) effect of PCAT19 knockdown or overexpression on A549 and SPC-A1 cell proliferation measured using the soft agar assay. (H,I) effects of PCAT19 knockdown or overexpression on A549 and SPC-A1 cell migration measured using the migration assay. (J–L) effects of PCAT19 knockdown or overexpression on A549 and SPC-A1 cells invasion using transwell assays. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: A549 cells were seeded into 96-well plates and were co-transfected with 60 ng of the dual luciferase reporter construct pmiR-RB-Report ™ -PCAT19 3′UTR (RiboBio, Guangdong) and miR-143-3p mimic.

Techniques: Over Expression, Fluorescence, Transfection, CCK-8 Assay, Soft Agar Assay, Migration

Journal: Frontiers in Genetics

Article Title: Integrated Dissection of lncRNA-miRNA-mRNA Pairs and Potential Regulatory Role of lncRNA PCAT19 in Lung Adenocarcinoma

doi: 10.3389/fgene.2021.765275

Figure Lengend Snippet: (A) Schematic diagram of the binding site between miR-143-3p and WT or Mut PCAT19. (B) Luciferase activity of WT or Mut PCAT19 after co-transfection of miR-143-3p mimic and WT or Mut PCAT19 dual fluorescent vector into A549 cells. (C) Correlation between the expression levels of PCAT19 and miR-143-3p. (D) Expression change of miR-143-3p after PCAT19 knockdown or overexpression in A549 and SPC-A1 cells. The expression level of U6 was used as internal reference. (E) CCK-8 analysis of the proliferation activity of A549 and SPC-A1 cells co-transfected with si-PCAT19 and miR-143-3p mimic. The data shown are the mean ± standard deviation of three experiments. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: A549 cells were seeded into 96-well plates and were co-transfected with 60 ng of the dual luciferase reporter construct pmiR-RB-Report ™ -PCAT19 3′UTR (RiboBio, Guangdong) and miR-143-3p mimic.

Techniques: Binding Assay, Luciferase, Activity Assay, Cotransfection, Plasmid Preparation, Expressing, Over Expression, CCK-8 Assay, Transfection, Standard Deviation

Journal: Apoptosis

Article Title: Identification and characterization of the human inhibitor of caspase-activated DNase gene promoter

doi: 10.1007/s10495-008-0223-8

Figure Lengend Snippet: ( a ) Mapping of the transcription start sites by 5′ RACE analysis. Following nested PCR reaction, a PCR product of 100-bp was detected by 1.5% agarose gel electrophoresis (lane 2). Lane 1: no template control; lane M: 100-bp DNA ladder marker. ( b ) The 5′ flanking region of the human ICAD gene. The nucleotide sequence is numbered from the major transcription site, which is indicated by an arrow. Putative binding sites for transcription factors analyzed by the TRANSFAC database and search program were shown with underlines

Article Snippet: DNA transfection of cells with each ICAD -promoter-luciferase construct (1 μg) with an internal control vector pRL-TK (0.1 μg) (Promega) was performed with Trans IT LT-1 (Mirus) during 6-h of incubation.

Techniques: Nested PCR, Agarose Gel Electrophoresis, Control, Marker, Sequencing, Binding Assay

Journal: Apoptosis

Article Title: Identification and characterization of the human inhibitor of caspase-activated DNase gene promoter

doi: 10.1007/s10495-008-0223-8

Figure Lengend Snippet: Human ICAD promoter drives reporter gene expression in transiently transfected Huh-7 cells. Deletion constructs of the upstream region of the ICAD gene linked to the firefly luciferase reporter gene (LUC) are illustrated in the left. Huh-7 cells were cotransfected with a firefly luciferase reporter and a Renilla luciferase internal control (pRL-TK). The relative luciferase activity (RLU) was obtained by normalizing the firefly luciferase activity to the Renilla luciferase activity, and is expressed as a percentage of RLU of pLuc(−936/+71). The average values are shown with standard deviation (SD) ( n = 4 per construct)

Article Snippet: DNA transfection of cells with each ICAD -promoter-luciferase construct (1 μg) with an internal control vector pRL-TK (0.1 μg) (Promega) was performed with Trans IT LT-1 (Mirus) during 6-h of incubation.

Techniques: Gene Expression, Transfection, Construct, Luciferase, Control, Activity Assay, Standard Deviation

Journal: Apoptosis

Article Title: Identification and characterization of the human inhibitor of caspase-activated DNase gene promoter

doi: 10.1007/s10495-008-0223-8

Figure Lengend Snippet: ( a ) ICAD promoter activity after transient expression of c-Myc, N-Myc, USF1 and USF2. Huh-7 cells were cotransfected with either expression vector for c-Myc, N-Myc, USF1 or USF2 driven by the CMV promoter and pLuc(−145/+71), pLuc(−90/+71) or pGL3 basic. Each firefly luciferase reporter plasmid ( n = 4) was cotransfected in cells with pRL-TK for normalization of the reporter activity. RLU is expressed as a percentage of that of pLuc(−145/+71) in the absence of expressing plasmids for Myc and USF. ( b ) Effect of substitution mutation in the E-box element on the ICAD promoter activity. Cells were cotransfected with pLuc(−106/±71) or an E-box-mutant, pLuc(−106/±71)mt and either expression vector for c-Myc or N-Myc. pRL-TK was also used for normalization of the activity. RLU is expressed as a percentage of that of pLuc(−106/±71) without over-expression of Myc

Article Snippet: DNA transfection of cells with each ICAD -promoter-luciferase construct (1 μg) with an internal control vector pRL-TK (0.1 μg) (Promega) was performed with Trans IT LT-1 (Mirus) during 6-h of incubation.

Techniques: Activity Assay, Expressing, Plasmid Preparation, Luciferase, Mutagenesis, Over Expression

Journal: Apoptosis

Article Title: Identification and characterization of the human inhibitor of caspase-activated DNase gene promoter

doi: 10.1007/s10495-008-0223-8

Figure Lengend Snippet: ( a ) An alignment of the sequence around putative Myc binding site of human ICAD promoter and the corresponding part of mouse ICAD sequence. ( b ) Binding of Myc proteins in the endogenous ICAD promoter. Crosslinked chromatins isolated from Huh-7, IMR-32 and GOTO cells were immunoprecipitated with indicated antibodies (c-Myc, N-Myc) or an equivalent amount of mouse IgG. Recovered DNAs were purified and PCR-amplified with primers for nt −272/±71 region or for nt −789/−451 region. Input represents 1% of chromatin sample applied for immunoprecipitation

Article Snippet: DNA transfection of cells with each ICAD -promoter-luciferase construct (1 μg) with an internal control vector pRL-TK (0.1 μg) (Promega) was performed with Trans IT LT-1 (Mirus) during 6-h of incubation.

Techniques: Sequencing, Binding Assay, Isolation, Immunoprecipitation, Purification, Amplification

Journal: Apoptosis

Article Title: Identification and characterization of the human inhibitor of caspase-activated DNase gene promoter

doi: 10.1007/s10495-008-0223-8

Figure Lengend Snippet: Myc-dependent expression of human ICAD. ( a ) Suppression of Myc expression decreases expression of ICAD protein. Huh-7 cells were transiently transfected with Myc siRNA (c-Myc, N-Myc) or the control siRNA (cont). Three days later, the cells were harvested and subjected to Western blotting. ( b ) Ectopic expression of Myc increases expression of ICAD protein. Cells (Huh-7 and mouse 3T3) were transfected with the expression vector for c-Myc or N-Myc, and after 3 days the cells were harvested for Western blotting. ( c ) Comparison of the mRNA expression of c-Myc and ICAD in Huh-7 cells and human oral squamous carcinoma cells (Ca22-9, HSC-2, and HSC-3). Levels of mRNA expression of ICAD and c-Myc were measured by quantitative real-time RT-PCR based on Taq Man chemistry. Results, relative to β-actin mRNA, are depicted as averages with SD ( n = 3)

Article Snippet: DNA transfection of cells with each ICAD -promoter-luciferase construct (1 μg) with an internal control vector pRL-TK (0.1 μg) (Promega) was performed with Trans IT LT-1 (Mirus) during 6-h of incubation.

Techniques: Expressing, Transfection, Control, Western Blot, Plasmid Preparation, Comparison, Quantitative RT-PCR