orc5 Search Results


gene exp orc5 mm00457242 m1  (Thermo Fisher)
85
Thermo Fisher gene exp orc5 mm00457242 m1
Gene Exp Orc5 Mm00457242 M1, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 85/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/gene exp orc5 mm00457242 m1/product/Thermo Fisher
Average 85 stars, based on 1 article reviews
gene exp orc5 mm00457242 m1 - by Bioz Stars, 2026-04
85/100 stars
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orc5  (Proteintech)
93
Proteintech orc5
ETV4 transcriptionally controls key genes involved in DNA replication of NSCLC cells. A) Functional analysis showing the top KEGG pathways that are significantly associated with down‐regulated genes in H1299, H1703, and H358T si‐ETV4 groups normalized to negative control (NC) siRNA groups by Human Microarray ( GSE137445 ). B) Clustering analysis showing the gene signature is enriched in the DNA replication pathway. C) IGV tracks showing the enrichment of ETV4 at the promoter region of MCM2, ‐4, ‐5, ‐10, and ORC1 from ChIP‐seq signals of A549‐shETV4 cells transfected with Flag‐ETV4 plasmids. D) ChIP‐PCR analysis for endogenous or exogenous ETV4 binding to the promoter region of MCM2, ‐4, ‐5, ‐10, and ORC1 genes in H1299 cells using anti‐ETV4 antibody or H358‐ETV4 cells using anti‐Flag antibody. E) Schematic depicting the core motif of ETV4 binding with its target genes, the wild‐type luciferase reporter constructs, and the mutations in the putative ETV4‐binding sites (mut‐1 or mut‐2 type) of MCMs/ORC1 promoter region. F–J) The wild‐type luciferase reporter plasmids of MCMs (ORC1) were co‐transfected along with ETV4, ETV4‐DBD deletion plasmid, or the empty vector into HEK293T cells. The constructed luc‐MCMs (ORC1)‐mut‐1 or ‐mut‐2 type plasmids were co‐transfected along with ETV4 plasmid or the empty vector into HEK293T cells. Relative luciferase activity was normalized against Renilla luciferase activity, respectively (mean ± SD; n = 4; ordinary one‐way ANOVA with Tukey's multiple comparisons test). **** p < 0.0001; ### p < 0.001, #### p < 0.0001. K) RT‐qPCR analysis of MCM2, ‐4, ‐5, ‐10, and ORC1 mRNA expression in H1299 cells transfected with ETV4 or NC siRNA, Transcript levels were normalized to ACTB gene expression (mean ± SD, n = 3; two‐tailed unpaired t ‐test). ** p < 0.01; *** p < 0.001; **** p < 0.0001. L) Immunoblots showing MCM2, ‐4, ‐5, ‐10, and <t>ORC1</t> <t>protein</t> levels in NC and ETV4‐knockdown H1299 cells. M,N) Immunoblots showing the Chromatin‐bound proteins (Chrom.) and unbound proteins (Sol.) levels of MCM2, ‐4, ‐5, ‐10, and ORC1 protein in control and sh‐ETV4 A549 cells, or control and ETV4‐overexpression H358 cells.
Orc5, supplied by Proteintech, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/orc5/product/Proteintech
Average 93 stars, based on 1 article reviews
orc5 - by Bioz Stars, 2026-04
93/100 stars
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gene exp orc5 dm01842615 g1  (Thermo Fisher)
86
Thermo Fisher gene exp orc5 dm01842615 g1
ETV4 transcriptionally controls key genes involved in DNA replication of NSCLC cells. A) Functional analysis showing the top KEGG pathways that are significantly associated with down‐regulated genes in H1299, H1703, and H358T si‐ETV4 groups normalized to negative control (NC) siRNA groups by Human Microarray ( GSE137445 ). B) Clustering analysis showing the gene signature is enriched in the DNA replication pathway. C) IGV tracks showing the enrichment of ETV4 at the promoter region of MCM2, ‐4, ‐5, ‐10, and ORC1 from ChIP‐seq signals of A549‐shETV4 cells transfected with Flag‐ETV4 plasmids. D) ChIP‐PCR analysis for endogenous or exogenous ETV4 binding to the promoter region of MCM2, ‐4, ‐5, ‐10, and ORC1 genes in H1299 cells using anti‐ETV4 antibody or H358‐ETV4 cells using anti‐Flag antibody. E) Schematic depicting the core motif of ETV4 binding with its target genes, the wild‐type luciferase reporter constructs, and the mutations in the putative ETV4‐binding sites (mut‐1 or mut‐2 type) of MCMs/ORC1 promoter region. F–J) The wild‐type luciferase reporter plasmids of MCMs (ORC1) were co‐transfected along with ETV4, ETV4‐DBD deletion plasmid, or the empty vector into HEK293T cells. The constructed luc‐MCMs (ORC1)‐mut‐1 or ‐mut‐2 type plasmids were co‐transfected along with ETV4 plasmid or the empty vector into HEK293T cells. Relative luciferase activity was normalized against Renilla luciferase activity, respectively (mean ± SD; n = 4; ordinary one‐way ANOVA with Tukey's multiple comparisons test). **** p < 0.0001; ### p < 0.001, #### p < 0.0001. K) RT‐qPCR analysis of MCM2, ‐4, ‐5, ‐10, and ORC1 mRNA expression in H1299 cells transfected with ETV4 or NC siRNA, Transcript levels were normalized to ACTB gene expression (mean ± SD, n = 3; two‐tailed unpaired t ‐test). ** p < 0.01; *** p < 0.001; **** p < 0.0001. L) Immunoblots showing MCM2, ‐4, ‐5, ‐10, and <t>ORC1</t> <t>protein</t> levels in NC and ETV4‐knockdown H1299 cells. M,N) Immunoblots showing the Chromatin‐bound proteins (Chrom.) and unbound proteins (Sol.) levels of MCM2, ‐4, ‐5, ‐10, and ORC1 protein in control and sh‐ETV4 A549 cells, or control and ETV4‐overexpression H358 cells.
Gene Exp Orc5 Dm01842615 G1, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/gene exp orc5 dm01842615 g1/product/Thermo Fisher
Average 86 stars, based on 1 article reviews
gene exp orc5 dm01842615 g1 - by Bioz Stars, 2026-04
86/100 stars
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orc5  (Boster Bio)
93
Boster Bio orc5
( A ) Scheme of introduced loxP sites in Orc 2 locus. ( B ) Representative picture of genotyping of offspring coming from Orc2 f/+ crossed with Orc2 f/+ . ( C ) The ratio of observed to expected animals coming from Orc2 f/+ crossed with Orc2 f/+ . ( D ) Schematic of the <t>ORC2</t> <t>protein</t> and the DeltaORC2 protein produced after deletion of exons 6 and 7. A110 is mutated to V110 and then the protein goes out of frame. ( E ) Validation of Orc2 deletion 3 d after Adeno cre transduction. ( F ) Western blot of ORC2 protein 5 d after Adeno cre transduction. 10 or indicated μl of lysate loaded/lane as written on the top. ( G ) MTT assay of WT and Orc2 f/f MEFs without and with Adeno cre transduction. ( H ) Western blot of ORC2 protein 5 and 15 d after Adeno Cre transduction. Figure 1—source data 1. PDF file containing original DNA gel picture corresponding to , panel B, indicating the relevant bands and individual animals. Figure 1—source data 2. Original image for , panel B. Figure 1—source data 3. PDF file containing original DNA gel picture corresponding to , panel E, indicating the relevant bands and increasing Adeno-Cre. Figure 1—source data 4. Original image for , panel E. Figure 1—source data 5. PDF file containing original Western blot membrane picture corresponding to , panel F, indicating the relevant bands and addition of Adeno-Cre. Figure 1—source data 6. Original image for , panel F. Figure 1—source data 7. PDF file containing original Western blot membrane picture corresponding to , panel H, indicating the relevant bands and ORC2 protein expression. Figure 1—source data 8. Original image for , panel H.
Orc5, supplied by Boster Bio, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/orc5/product/Boster Bio
Average 93 stars, based on 1 article reviews
orc5 - by Bioz Stars, 2026-04
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orc5  (Becton Dickinson)
90
Becton Dickinson orc5
( A ) Scheme of introduced loxP sites in Orc 2 locus. ( B ) Representative picture of genotyping of offspring coming from Orc2 f/+ crossed with Orc2 f/+ . ( C ) The ratio of observed to expected animals coming from Orc2 f/+ crossed with Orc2 f/+ . ( D ) Schematic of the <t>ORC2</t> <t>protein</t> and the DeltaORC2 protein produced after deletion of exons 6 and 7. A110 is mutated to V110 and then the protein goes out of frame. ( E ) Validation of Orc2 deletion 3 d after Adeno cre transduction. ( F ) Western blot of ORC2 protein 5 d after Adeno cre transduction. 10 or indicated μl of lysate loaded/lane as written on the top. ( G ) MTT assay of WT and Orc2 f/f MEFs without and with Adeno cre transduction. ( H ) Western blot of ORC2 protein 5 and 15 d after Adeno Cre transduction. Figure 1—source data 1. PDF file containing original DNA gel picture corresponding to , panel B, indicating the relevant bands and individual animals. Figure 1—source data 2. Original image for , panel B. Figure 1—source data 3. PDF file containing original DNA gel picture corresponding to , panel E, indicating the relevant bands and increasing Adeno-Cre. Figure 1—source data 4. Original image for , panel E. Figure 1—source data 5. PDF file containing original Western blot membrane picture corresponding to , panel F, indicating the relevant bands and addition of Adeno-Cre. Figure 1—source data 6. Original image for , panel F. Figure 1—source data 7. PDF file containing original Western blot membrane picture corresponding to , panel H, indicating the relevant bands and ORC2 protein expression. Figure 1—source data 8. Original image for , panel H.
Orc5, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/orc5/product/Becton Dickinson
Average 90 stars, based on 1 article reviews
orc5 - by Bioz Stars, 2026-04
90/100 stars
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certified reference material, roasted coffee with a level of ochratoxin a1⁄4 6.6 ppb 1.00  (Biopharm GmbH)
90
Biopharm GmbH certified reference material, roasted coffee with a level of ochratoxin a1⁄4 6.6 ppb 1.00
( A ) Scheme of introduced loxP sites in Orc 2 locus. ( B ) Representative picture of genotyping of offspring coming from Orc2 f/+ crossed with Orc2 f/+ . ( C ) The ratio of observed to expected animals coming from Orc2 f/+ crossed with Orc2 f/+ . ( D ) Schematic of the <t>ORC2</t> <t>protein</t> and the DeltaORC2 protein produced after deletion of exons 6 and 7. A110 is mutated to V110 and then the protein goes out of frame. ( E ) Validation of Orc2 deletion 3 d after Adeno cre transduction. ( F ) Western blot of ORC2 protein 5 d after Adeno cre transduction. 10 or indicated μl of lysate loaded/lane as written on the top. ( G ) MTT assay of WT and Orc2 f/f MEFs without and with Adeno cre transduction. ( H ) Western blot of ORC2 protein 5 and 15 d after Adeno Cre transduction. Figure 1—source data 1. PDF file containing original DNA gel picture corresponding to , panel B, indicating the relevant bands and individual animals. Figure 1—source data 2. Original image for , panel B. Figure 1—source data 3. PDF file containing original DNA gel picture corresponding to , panel E, indicating the relevant bands and increasing Adeno-Cre. Figure 1—source data 4. Original image for , panel E. Figure 1—source data 5. PDF file containing original Western blot membrane picture corresponding to , panel F, indicating the relevant bands and addition of Adeno-Cre. Figure 1—source data 6. Original image for , panel F. Figure 1—source data 7. PDF file containing original Western blot membrane picture corresponding to , panel H, indicating the relevant bands and ORC2 protein expression. Figure 1—source data 8. Original image for , panel H.
Certified Reference Material, Roasted Coffee With A Level Of Ochratoxin A1⁄4 6.6 Ppb 1.00, supplied by Biopharm GmbH, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/certified reference material, roasted coffee with a level of ochratoxin a1⁄4 6.6 ppb 1.00/product/Biopharm GmbH
Average 90 stars, based on 1 article reviews
certified reference material, roasted coffee with a level of ochratoxin a1⁄4 6.6 ppb 1.00 - by Bioz Stars, 2026-04
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orc5l orc5 nm 002553 human tagged orf clone  (OriGene)
N/A
ORC5 Myc DDK tagged Human origin recognition complex subunit 5 ORC5 transcript variant 1
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orc5 nm 001014186 rat tagged orf clone lentiviral particle  (OriGene)
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Lenti ORF particles Orc5 GFP tagged ORF Rat origin recognition complex subunit 5 like yeast Orc5l 200ul 10 7 TU mL
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orc5 mouse shrna plasmid  (OriGene)
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Orc5l Mouse 4 unique 29mer shRNA constructs in lentiviral GFP vector
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orc5 mouse shrna lentiviral particle  (OriGene)
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Orc5l Mouse shRNA lentiviral particles 4 unique 29mer target specific shRNA 1 scramble control 0 5 ml each 10 7 TU ml
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orc5l orc5 nm 181747 human tagged orf clone  (OriGene)
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Image Search Results


ETV4 transcriptionally controls key genes involved in DNA replication of NSCLC cells. A) Functional analysis showing the top KEGG pathways that are significantly associated with down‐regulated genes in H1299, H1703, and H358T si‐ETV4 groups normalized to negative control (NC) siRNA groups by Human Microarray ( GSE137445 ). B) Clustering analysis showing the gene signature is enriched in the DNA replication pathway. C) IGV tracks showing the enrichment of ETV4 at the promoter region of MCM2, ‐4, ‐5, ‐10, and ORC1 from ChIP‐seq signals of A549‐shETV4 cells transfected with Flag‐ETV4 plasmids. D) ChIP‐PCR analysis for endogenous or exogenous ETV4 binding to the promoter region of MCM2, ‐4, ‐5, ‐10, and ORC1 genes in H1299 cells using anti‐ETV4 antibody or H358‐ETV4 cells using anti‐Flag antibody. E) Schematic depicting the core motif of ETV4 binding with its target genes, the wild‐type luciferase reporter constructs, and the mutations in the putative ETV4‐binding sites (mut‐1 or mut‐2 type) of MCMs/ORC1 promoter region. F–J) The wild‐type luciferase reporter plasmids of MCMs (ORC1) were co‐transfected along with ETV4, ETV4‐DBD deletion plasmid, or the empty vector into HEK293T cells. The constructed luc‐MCMs (ORC1)‐mut‐1 or ‐mut‐2 type plasmids were co‐transfected along with ETV4 plasmid or the empty vector into HEK293T cells. Relative luciferase activity was normalized against Renilla luciferase activity, respectively (mean ± SD; n = 4; ordinary one‐way ANOVA with Tukey's multiple comparisons test). **** p < 0.0001; ### p < 0.001, #### p < 0.0001. K) RT‐qPCR analysis of MCM2, ‐4, ‐5, ‐10, and ORC1 mRNA expression in H1299 cells transfected with ETV4 or NC siRNA, Transcript levels were normalized to ACTB gene expression (mean ± SD, n = 3; two‐tailed unpaired t ‐test). ** p < 0.01; *** p < 0.001; **** p < 0.0001. L) Immunoblots showing MCM2, ‐4, ‐5, ‐10, and ORC1 protein levels in NC and ETV4‐knockdown H1299 cells. M,N) Immunoblots showing the Chromatin‐bound proteins (Chrom.) and unbound proteins (Sol.) levels of MCM2, ‐4, ‐5, ‐10, and ORC1 protein in control and sh‐ETV4 A549 cells, or control and ETV4‐overexpression H358 cells.

Journal: Advanced Science

Article Title: Topoisomerase I Inhibition in ETV4‐overexpressed Non‐Small Cell Lung Cancer Promotes Replication and Transcription Mediated R‐Loop Accumulation and DNA Damage

doi: 10.1002/advs.202409307

Figure Lengend Snippet: ETV4 transcriptionally controls key genes involved in DNA replication of NSCLC cells. A) Functional analysis showing the top KEGG pathways that are significantly associated with down‐regulated genes in H1299, H1703, and H358T si‐ETV4 groups normalized to negative control (NC) siRNA groups by Human Microarray ( GSE137445 ). B) Clustering analysis showing the gene signature is enriched in the DNA replication pathway. C) IGV tracks showing the enrichment of ETV4 at the promoter region of MCM2, ‐4, ‐5, ‐10, and ORC1 from ChIP‐seq signals of A549‐shETV4 cells transfected with Flag‐ETV4 plasmids. D) ChIP‐PCR analysis for endogenous or exogenous ETV4 binding to the promoter region of MCM2, ‐4, ‐5, ‐10, and ORC1 genes in H1299 cells using anti‐ETV4 antibody or H358‐ETV4 cells using anti‐Flag antibody. E) Schematic depicting the core motif of ETV4 binding with its target genes, the wild‐type luciferase reporter constructs, and the mutations in the putative ETV4‐binding sites (mut‐1 or mut‐2 type) of MCMs/ORC1 promoter region. F–J) The wild‐type luciferase reporter plasmids of MCMs (ORC1) were co‐transfected along with ETV4, ETV4‐DBD deletion plasmid, or the empty vector into HEK293T cells. The constructed luc‐MCMs (ORC1)‐mut‐1 or ‐mut‐2 type plasmids were co‐transfected along with ETV4 plasmid or the empty vector into HEK293T cells. Relative luciferase activity was normalized against Renilla luciferase activity, respectively (mean ± SD; n = 4; ordinary one‐way ANOVA with Tukey's multiple comparisons test). **** p < 0.0001; ### p < 0.001, #### p < 0.0001. K) RT‐qPCR analysis of MCM2, ‐4, ‐5, ‐10, and ORC1 mRNA expression in H1299 cells transfected with ETV4 or NC siRNA, Transcript levels were normalized to ACTB gene expression (mean ± SD, n = 3; two‐tailed unpaired t ‐test). ** p < 0.01; *** p < 0.001; **** p < 0.0001. L) Immunoblots showing MCM2, ‐4, ‐5, ‐10, and ORC1 protein levels in NC and ETV4‐knockdown H1299 cells. M,N) Immunoblots showing the Chromatin‐bound proteins (Chrom.) and unbound proteins (Sol.) levels of MCM2, ‐4, ‐5, ‐10, and ORC1 protein in control and sh‐ETV4 A549 cells, or control and ETV4‐overexpression H358 cells.

Article Snippet: Blots were blocked with 5% nonfat milk in Tris‐Buffered Saline and Tween 20 (TBST), and incubated with antibodies specific for ETV4 (10684‐1‐AP, Proteintech), MCM2 (3619, CST), MCM3 (15597‐1‐AP, Proteintech), MCM4 (13043‐1‐AP, Proteintech), MCM5 (11703‐1‐AP, Proteintech), MCM6 (13347‐2‐AP, Proteintech), MCM7 (11225‐1‐AP, Proteintech), MCM10 (12251‐1‐AP, Proteintech), ORC1 (NBP100‐121, Novus), ORC2 (12739‐1‐AP, Proteintech), ORC3 (sc‐374231, Santa Cruz), ORC4 (13026‐1‐AP, Proteintech), ORC5 (11542‐1‐AP, Proteintech), ORC6 (17784‐1‐AP, Proteintech), Histone‐H3 (17168‐1‐AP, Proteintech), PCNA (13110, CST), phospho‐MCM2 S40 (ab133243, Abcam), CDC45 (15678‐1‐AP, Proteintech), SUPT16H (28598‐1‐AP, Proteintech), SSRP1 (15696‐1‐AP, Proteintech), γ‐H2AX (ET‐1602, HUABio), Flag (14793, CST), His (66005‐1‐Ig, Proteintech), HA (sc‐7392, Santa Cruz), GST (66001‐2‐lg, Proteintech), GAPDH (60004‐1‐Ig, Proteintech) and β‐actin (66009‐1‐Ig, Proteintech) was used as a loading control for Western blot.

Techniques: Functional Assay, Negative Control, Microarray, ChIP-sequencing, Transfection, Binding Assay, Luciferase, Construct, Plasmid Preparation, Activity Assay, Quantitative RT-PCR, Expressing, Gene Expression, Two Tailed Test, Western Blot, Knockdown, Control, Over Expression

( A ) Scheme of introduced loxP sites in Orc 2 locus. ( B ) Representative picture of genotyping of offspring coming from Orc2 f/+ crossed with Orc2 f/+ . ( C ) The ratio of observed to expected animals coming from Orc2 f/+ crossed with Orc2 f/+ . ( D ) Schematic of the ORC2 protein and the DeltaORC2 protein produced after deletion of exons 6 and 7. A110 is mutated to V110 and then the protein goes out of frame. ( E ) Validation of Orc2 deletion 3 d after Adeno cre transduction. ( F ) Western blot of ORC2 protein 5 d after Adeno cre transduction. 10 or indicated μl of lysate loaded/lane as written on the top. ( G ) MTT assay of WT and Orc2 f/f MEFs without and with Adeno cre transduction. ( H ) Western blot of ORC2 protein 5 and 15 d after Adeno Cre transduction. Figure 1—source data 1. PDF file containing original DNA gel picture corresponding to , panel B, indicating the relevant bands and individual animals. Figure 1—source data 2. Original image for , panel B. Figure 1—source data 3. PDF file containing original DNA gel picture corresponding to , panel E, indicating the relevant bands and increasing Adeno-Cre. Figure 1—source data 4. Original image for , panel E. Figure 1—source data 5. PDF file containing original Western blot membrane picture corresponding to , panel F, indicating the relevant bands and addition of Adeno-Cre. Figure 1—source data 6. Original image for , panel F. Figure 1—source data 7. PDF file containing original Western blot membrane picture corresponding to , panel H, indicating the relevant bands and ORC2 protein expression. Figure 1—source data 8. Original image for , panel H.

Journal: eLife

Article Title: DNA replication in primary hepatocytes without the six-subunit ORC

doi: 10.7554/eLife.102915

Figure Lengend Snippet: ( A ) Scheme of introduced loxP sites in Orc 2 locus. ( B ) Representative picture of genotyping of offspring coming from Orc2 f/+ crossed with Orc2 f/+ . ( C ) The ratio of observed to expected animals coming from Orc2 f/+ crossed with Orc2 f/+ . ( D ) Schematic of the ORC2 protein and the DeltaORC2 protein produced after deletion of exons 6 and 7. A110 is mutated to V110 and then the protein goes out of frame. ( E ) Validation of Orc2 deletion 3 d after Adeno cre transduction. ( F ) Western blot of ORC2 protein 5 d after Adeno cre transduction. 10 or indicated μl of lysate loaded/lane as written on the top. ( G ) MTT assay of WT and Orc2 f/f MEFs without and with Adeno cre transduction. ( H ) Western blot of ORC2 protein 5 and 15 d after Adeno Cre transduction. Figure 1—source data 1. PDF file containing original DNA gel picture corresponding to , panel B, indicating the relevant bands and individual animals. Figure 1—source data 2. Original image for , panel B. Figure 1—source data 3. PDF file containing original DNA gel picture corresponding to , panel E, indicating the relevant bands and increasing Adeno-Cre. Figure 1—source data 4. Original image for , panel E. Figure 1—source data 5. PDF file containing original Western blot membrane picture corresponding to , panel F, indicating the relevant bands and addition of Adeno-Cre. Figure 1—source data 6. Original image for , panel F. Figure 1—source data 7. PDF file containing original Western blot membrane picture corresponding to , panel H, indicating the relevant bands and ORC2 protein expression. Figure 1—source data 8. Original image for , panel H.

Article Snippet: The antibodies used in this study are listed: ORC1 (Santa Cruz; sc-28741); ORC3 (Santa Cruz; sc-374231); ORC5 (Boster Biological technology; A03676-1); ORC6 (Santa Cruz; sc-390490); CDC6 (Santa Cruz; sc-9964); MCM2 (Abcam; ab4461); MCM3 (Santa Cruz; sc-9850).

Techniques: Produced, Biomarker Discovery, Transduction, Western Blot, MTT Assay, Membrane, Expressing

( A ) Scheme of Alb +/- - Orc2 f/f ROSA26 stop-EYFP crossed with Alb +/- -Orc2 f/f ROSA26 stop-EYFP (All mice are with ROSA26 stop-EYFP and so we do not include this in the genotypes below). ( B ) The ratio of observed to expected animals coming from A. ( C ) Western blot of hepatocytes from Orc2 f/f and Alb +/- - Orc2 f/f animals. Tubulin was used as loading control. ( D ) Quantification of the Western blots of hepatocyte lysates from Orc2 f/f (without Alb-cre ) mice and the same genotype but with Alb-Cre to show the levels of other key replication initiation proteins in the ORC2 KO hepatocytes. ( E ) Average body weight of Orc2 f/f and Alb-Orc2 f/f animals. ( F ) Average liver weight of Orc2 f/f and Alb-Orc2 f/f animals. ( G ) Average liver-to-body weight ratio of Orc2 f/f and Alb-Orc2 f/f animals. ( H ) Representative H&E staining of liver tissue from Orc2 f/f (WT) and Alb-Orc2 f/f (KO) animals. Both panels at same scale. ( I ) Quantification of hepatocyte nuclear size in Orc2 f/f and Alb-Orc2 f/f animals. ( J ) Quantification of hepatocyte nuclear size in Orc2 f/f and Alb-Orc2 f/f female mice. ( K ) Quantification of hepatocytes nuclear size in Orc2 f/f and Alb-Orc2 f/f male mice. *p<0.05, **p<0.01, two-tailed Student’s t-test. Figure 2—source data 1. Original Western blot membrane picture corresponding to , panel C. Molecular weight markers are labeled on the left. The bands next to the arrow represent ORC2 protein. Figure 2—source data 2. Original image for , panel C. Figure 2—source data 3. Original Western blot membrane picture corresponding to , panel C. Molecular weight markers are labeled on the left. The bands next to the arrow represent Tubulin protein. Figure 2—source data 4. Original image for , panel C.

Journal: eLife

Article Title: DNA replication in primary hepatocytes without the six-subunit ORC

doi: 10.7554/eLife.102915

Figure Lengend Snippet: ( A ) Scheme of Alb +/- - Orc2 f/f ROSA26 stop-EYFP crossed with Alb +/- -Orc2 f/f ROSA26 stop-EYFP (All mice are with ROSA26 stop-EYFP and so we do not include this in the genotypes below). ( B ) The ratio of observed to expected animals coming from A. ( C ) Western blot of hepatocytes from Orc2 f/f and Alb +/- - Orc2 f/f animals. Tubulin was used as loading control. ( D ) Quantification of the Western blots of hepatocyte lysates from Orc2 f/f (without Alb-cre ) mice and the same genotype but with Alb-Cre to show the levels of other key replication initiation proteins in the ORC2 KO hepatocytes. ( E ) Average body weight of Orc2 f/f and Alb-Orc2 f/f animals. ( F ) Average liver weight of Orc2 f/f and Alb-Orc2 f/f animals. ( G ) Average liver-to-body weight ratio of Orc2 f/f and Alb-Orc2 f/f animals. ( H ) Representative H&E staining of liver tissue from Orc2 f/f (WT) and Alb-Orc2 f/f (KO) animals. Both panels at same scale. ( I ) Quantification of hepatocyte nuclear size in Orc2 f/f and Alb-Orc2 f/f animals. ( J ) Quantification of hepatocyte nuclear size in Orc2 f/f and Alb-Orc2 f/f female mice. ( K ) Quantification of hepatocytes nuclear size in Orc2 f/f and Alb-Orc2 f/f male mice. *p<0.05, **p<0.01, two-tailed Student’s t-test. Figure 2—source data 1. Original Western blot membrane picture corresponding to , panel C. Molecular weight markers are labeled on the left. The bands next to the arrow represent ORC2 protein. Figure 2—source data 2. Original image for , panel C. Figure 2—source data 3. Original Western blot membrane picture corresponding to , panel C. Molecular weight markers are labeled on the left. The bands next to the arrow represent Tubulin protein. Figure 2—source data 4. Original image for , panel C.

Article Snippet: The antibodies used in this study are listed: ORC1 (Santa Cruz; sc-28741); ORC3 (Santa Cruz; sc-374231); ORC5 (Boster Biological technology; A03676-1); ORC6 (Santa Cruz; sc-390490); CDC6 (Santa Cruz; sc-9964); MCM2 (Abcam; ab4461); MCM3 (Santa Cruz; sc-9850).

Techniques: Western Blot, Control, Staining, Two Tailed Test, Membrane, Molecular Weight, Labeling

( A–B ) Breeding schemes to obtain conditional double flox animals. ( C ) The ratio of observed to expected animals coming from B. Orc1 =all animals with Orc1 f/f ROSA26 stop-EYFP , Orc2 =all animals with Orc2 f/f ROSA26 stop-EYFP , Orc1 Orc2 =all animals with Orc1 f/f Orc2 f/f ROSA26 stop-EYFP genotype. This was before the introduction of Alb-Cre . ( D ) Immunoblot of hepatocytes from WT ( Orc1 f/f Orc2 f/f ) and DKO ( Orc1 f/f Orc2 f/f Alb-cre +/- ) mice to show that ORC1 and ORC2 are depleted in the DKO cells. ( E ) Quantitation of immunoblots to show that levels of other key initiation protein subunits are not decreased in the DKO mice hepatocytes. ( F ) Average body, liver weight, and their ratio for WT and DKO animals. ( G ) Representative H&E staining of liver tissue from male WT and DKO animals. ( H ) Quantification of hepatocyte nuclear size in the WT and DKO animals. ( I ) Quantification of nuclei ploidy for EYFP low (includes negative) and high (positive) primary liver cells from DKO mice. Figure 6—source data 1. PDF file containing original Western blot membrane picture corresponding to , panel D, indicating the relevant bands, ORC1 protein expression, and individual animals. Figure 6—source data 2. Original image for panel D, ORC1 protein expression, and individual animals. Figure 6—source data 3. PDF file containing original Western blot membrane picture corresponding to , panel D, indicating the relevant bands, ORC2 protein expression, and individual animals. Figure 6—source data 4. Original image for panel D, ORC2 protein expression, and individual animals. Figure 6—source data 5. PDF file containing original Western blot membrane picture corresponding to , panel D, indicating the relevant bands, HSP90 protein expression, and individual animals. Figure 6—source data 6. Original image for panel D, HSP90 protein expression, and individual animals.

Journal: eLife

Article Title: DNA replication in primary hepatocytes without the six-subunit ORC

doi: 10.7554/eLife.102915

Figure Lengend Snippet: ( A–B ) Breeding schemes to obtain conditional double flox animals. ( C ) The ratio of observed to expected animals coming from B. Orc1 =all animals with Orc1 f/f ROSA26 stop-EYFP , Orc2 =all animals with Orc2 f/f ROSA26 stop-EYFP , Orc1 Orc2 =all animals with Orc1 f/f Orc2 f/f ROSA26 stop-EYFP genotype. This was before the introduction of Alb-Cre . ( D ) Immunoblot of hepatocytes from WT ( Orc1 f/f Orc2 f/f ) and DKO ( Orc1 f/f Orc2 f/f Alb-cre +/- ) mice to show that ORC1 and ORC2 are depleted in the DKO cells. ( E ) Quantitation of immunoblots to show that levels of other key initiation protein subunits are not decreased in the DKO mice hepatocytes. ( F ) Average body, liver weight, and their ratio for WT and DKO animals. ( G ) Representative H&E staining of liver tissue from male WT and DKO animals. ( H ) Quantification of hepatocyte nuclear size in the WT and DKO animals. ( I ) Quantification of nuclei ploidy for EYFP low (includes negative) and high (positive) primary liver cells from DKO mice. Figure 6—source data 1. PDF file containing original Western blot membrane picture corresponding to , panel D, indicating the relevant bands, ORC1 protein expression, and individual animals. Figure 6—source data 2. Original image for panel D, ORC1 protein expression, and individual animals. Figure 6—source data 3. PDF file containing original Western blot membrane picture corresponding to , panel D, indicating the relevant bands, ORC2 protein expression, and individual animals. Figure 6—source data 4. Original image for panel D, ORC2 protein expression, and individual animals. Figure 6—source data 5. PDF file containing original Western blot membrane picture corresponding to , panel D, indicating the relevant bands, HSP90 protein expression, and individual animals. Figure 6—source data 6. Original image for panel D, HSP90 protein expression, and individual animals.

Article Snippet: The antibodies used in this study are listed: ORC1 (Santa Cruz; sc-28741); ORC3 (Santa Cruz; sc-374231); ORC5 (Boster Biological technology; A03676-1); ORC6 (Santa Cruz; sc-390490); CDC6 (Santa Cruz; sc-9964); MCM2 (Abcam; ab4461); MCM3 (Santa Cruz; sc-9850).

Techniques: Western Blot, Quantitation Assay, Staining, Membrane, Expressing