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OriGene primer sequences
Primer Sequences, supplied by OriGene, used in various techniques. Bioz Stars score: 92/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/primer sequences/product/OriGene
Average 92 stars, based on 2 article reviews
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primer sequences - by Bioz Stars, 2020-07
92/100 stars

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Clone Assay:

Article Title: The RNA processing factors THRAP3 and BCLAF1 promote the DNA damage response through selective mRNA splicing and nuclear export
Article Snippet: .. The sequence encoding the ATM exon20-intron20-exon21 was PCR amplified from 293T derived genomic DNA using the following primer sequences: Forward 5′- GTGAAGCTTTCCAATGTGTGTTCTTTGTATC-3′ and Reverse 5′- TACGTCGACCTCAAGCAAAGTTTTAAG-3′, and cloned into the pRFP2GFP vector’s ORF between RFP and GFP tags using novel HindIII and SalI sites. pCMV6-AN-Myc-DDK was purchased from Origene (PS100016). .. The ORF of THRAP3 was inserted in this plasmid using SgfI and MluI sites.

Amplification:

Article Title: The RNA processing factors THRAP3 and BCLAF1 promote the DNA damage response through selective mRNA splicing and nuclear export
Article Snippet: .. The sequence encoding the ATM exon20-intron20-exon21 was PCR amplified from 293T derived genomic DNA using the following primer sequences: Forward 5′- GTGAAGCTTTCCAATGTGTGTTCTTTGTATC-3′ and Reverse 5′- TACGTCGACCTCAAGCAAAGTTTTAAG-3′, and cloned into the pRFP2GFP vector’s ORF between RFP and GFP tags using novel HindIII and SalI sites. pCMV6-AN-Myc-DDK was purchased from Origene (PS100016). .. The ORF of THRAP3 was inserted in this plasmid using SgfI and MluI sites.

Real-time Polymerase Chain Reaction:

Article Title: Transcriptional Profiling of Newly Generated Dentate Granule Cells Using TU Tagging Reveals Pattern Shifts in Gene Expression during Circuit Integration 1Transcriptional Profiling of Newly Generated Dentate Granule Cells Using TU Tagging Reveals Pattern Shifts in Gene Expression during Circuit Integration 1 , 2
Article Snippet: .. Primer sequences were obtained from the Origene validated quantitative PCR (qPCR) primer collection (Origene Technologies; ). .. Statistical analysis was performed with Prism software (GraphPad Software).

Polymerase Chain Reaction:

Article Title: The RNA processing factors THRAP3 and BCLAF1 promote the DNA damage response through selective mRNA splicing and nuclear export
Article Snippet: .. The sequence encoding the ATM exon20-intron20-exon21 was PCR amplified from 293T derived genomic DNA using the following primer sequences: Forward 5′- GTGAAGCTTTCCAATGTGTGTTCTTTGTATC-3′ and Reverse 5′- TACGTCGACCTCAAGCAAAGTTTTAAG-3′, and cloned into the pRFP2GFP vector’s ORF between RFP and GFP tags using novel HindIII and SalI sites. pCMV6-AN-Myc-DDK was purchased from Origene (PS100016). .. The ORF of THRAP3 was inserted in this plasmid using SgfI and MluI sites.

Plasmid Preparation:

Article Title: The RNA processing factors THRAP3 and BCLAF1 promote the DNA damage response through selective mRNA splicing and nuclear export
Article Snippet: .. The sequence encoding the ATM exon20-intron20-exon21 was PCR amplified from 293T derived genomic DNA using the following primer sequences: Forward 5′- GTGAAGCTTTCCAATGTGTGTTCTTTGTATC-3′ and Reverse 5′- TACGTCGACCTCAAGCAAAGTTTTAAG-3′, and cloned into the pRFP2GFP vector’s ORF between RFP and GFP tags using novel HindIII and SalI sites. pCMV6-AN-Myc-DDK was purchased from Origene (PS100016). .. The ORF of THRAP3 was inserted in this plasmid using SgfI and MluI sites.

Sequencing:

Article Title: Nerve growth factor modulates the tumor cells migration in ovarian cancer through the WNT/β-catenin pathway
Article Snippet: .. The primer sequences of GAPDH, NGF, C-myc, CD44, MMP2, MMP7, TIMP2 and BCL9-2 were from http://pga.mgh.harvard.edu/primerbank/; The primer sequences of P75 were from http://www.origene.com/qPCR/Primers.aspx; The primer sequences of β-catenin were from http://medgen.ugent.be/rtprimerdb/assay; The primer sequences of TrkA were designed using Primer-Blast from sequence submitted to the Genbank. ..

Article Title: The RNA processing factors THRAP3 and BCLAF1 promote the DNA damage response through selective mRNA splicing and nuclear export
Article Snippet: .. The sequence encoding the ATM exon20-intron20-exon21 was PCR amplified from 293T derived genomic DNA using the following primer sequences: Forward 5′- GTGAAGCTTTCCAATGTGTGTTCTTTGTATC-3′ and Reverse 5′- TACGTCGACCTCAAGCAAAGTTTTAAG-3′, and cloned into the pRFP2GFP vector’s ORF between RFP and GFP tags using novel HindIII and SalI sites. pCMV6-AN-Myc-DDK was purchased from Origene (PS100016). .. The ORF of THRAP3 was inserted in this plasmid using SgfI and MluI sites.

Derivative Assay:

Article Title: The RNA processing factors THRAP3 and BCLAF1 promote the DNA damage response through selective mRNA splicing and nuclear export
Article Snippet: .. The sequence encoding the ATM exon20-intron20-exon21 was PCR amplified from 293T derived genomic DNA using the following primer sequences: Forward 5′- GTGAAGCTTTCCAATGTGTGTTCTTTGTATC-3′ and Reverse 5′- TACGTCGACCTCAAGCAAAGTTTTAAG-3′, and cloned into the pRFP2GFP vector’s ORF between RFP and GFP tags using novel HindIII and SalI sites. pCMV6-AN-Myc-DDK was purchased from Origene (PS100016). .. The ORF of THRAP3 was inserted in this plasmid using SgfI and MluI sites.

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    OriGene nqo1 dna sequence
    3.3. Influence of <t>NQO1</t> on E2 -3,4-Q metabolism and <t>DNA</t> adduct formation
    Nqo1 Dna Sequence, supplied by OriGene, 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/nqo1 dna sequence/product/OriGene
    Average 85 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    nqo1 dna sequence - by Bioz Stars, 2020-07
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    93
    OriGene cdk2ap1
    Observed phenotype following <t>CDK2AP1</t> knockdown is p53 dependent. Panel A: Primary HDFs were transduced with EV or CDK2AP1-shRNA or CDK2AP1-shRNA and a p53 specific shRNA and were analyzed by qPCR for mRNA levels of p53 , p21 , BAX and PUMA . Knockdown of p53 with CDK2AP1 prevented the increase in p21, BAX and PUMA. Panel B: The knockdown of p53 with CDK2AP1 increased the percentage of cells in the S and reduced the cells in G1 when compared to CDK2AP1 only knockdown cells (p-value
    Cdk2ap1, supplied by OriGene, used in various techniques. Bioz Stars score: 93/100, based on 12 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    OriGene socs5
    miR-432 targets <t>SOCS5.</t> For validation of SOCS5 UTR as target of miR-432, 100 pmol of miR-432 mimic was overexpressed in CHME3 cells. (A) Western blot showing downregulation of SOCS5 upon miR-432 overexpression in CHME3 cells. The average fold change values with respect to control have been mentioned. (B) Overexpression of miR-432 mimic was confirmed by Real time PCR using miR-432 specific TaqMan probe. RNU24 was used for normalization. (C) Blot showing upregulation of SOCS5 upon antimiR-432 transfection. The average fold change values with respect to control have been mentioned . (D) Silencing of miR-432 by antimiR-432 transfection was confirmed by real time PCR. (E) SOCS5 UTR clone (pEZX-MT06) for microRNA targeting was transfected along with miR-432 mimic along with β-galactosidase vector. Graph shows decreased luciferase activity of the vector in presence of miR-432 mimic. A mutant clone was generated deleting the targeting region present in SOCS5 UTR (shown in Fig. 2) and transfected along with miR-432 mimic. The mutant clone did not display any reduction in luciferase activity. Non-targeting mimic miR-146a was also used as negative control. β-galactosidase activity was used for normalization. All experiments were performed in triplicates. The data are shown as mean ± S.E from three independent experiments. The fold change is significant where *denotes P
    Socs5, supplied by OriGene, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    OriGene human mir 141 primer pairs
    <t>miR-141</t> binding to solute carrier family 25 member 3 (Slc25a3). A : sequence homology between Slc25a3 3′-UTR ( top ) and miR-141 ( bottom ). Black lines represent perfect match, and grey lines represent G-U wobbles. B : miR-141 recognition sequence
    Human Mir 141 Primer Pairs, supplied by OriGene, used in various techniques. Bioz Stars score: 85/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    3.3. Influence of NQO1 on E2 -3,4-Q metabolism and DNA adduct formation

    Journal: The Journal of steroid biochemistry and molecular biology

    Article Title: NAD(P)H:quinone oxidoreductase 1 Arg139Trp and Pro187Ser polymorphisms imbalance estrogen metabolism towards DNA adduct formation in human mammary epithelial cells

    doi: 10.1016/j.jsbmb.2009.07.003

    Figure Lengend Snippet: 3.3. Influence of NQO1 on E2 -3,4-Q metabolism and DNA adduct formation

    Article Snippet: Following validation of the NQO1 DNA sequence (with vector primer v1.5 and vector primer XL39, Origene), this plasmid was used as a template for generating three mutant clones; one contained an AAALys 135 TAAStop mutation in exon 4 of the NQO1 gene (Entrez GeneID: 1728), a second contained the polymorphic CGGArg 139 TGGTrp mutation in exon 4, and a third contained the polymorphic CCTPro 187 TCTSer mutation in exon 6.

    Techniques:

    Analysis of estrogen metabolism and DNA adduct formation in MCF-10F cells expressing wild-type and polymorphic NQO1 variants.

    Journal: The Journal of steroid biochemistry and molecular biology

    Article Title: NAD(P)H:quinone oxidoreductase 1 Arg139Trp and Pro187Ser polymorphisms imbalance estrogen metabolism towards DNA adduct formation in human mammary epithelial cells

    doi: 10.1016/j.jsbmb.2009.07.003

    Figure Lengend Snippet: Analysis of estrogen metabolism and DNA adduct formation in MCF-10F cells expressing wild-type and polymorphic NQO1 variants.

    Article Snippet: Following validation of the NQO1 DNA sequence (with vector primer v1.5 and vector primer XL39, Origene), this plasmid was used as a template for generating three mutant clones; one contained an AAALys 135 TAAStop mutation in exon 4 of the NQO1 gene (Entrez GeneID: 1728), a second contained the polymorphic CGGArg 139 TGGTrp mutation in exon 4, and a third contained the polymorphic CCTPro 187 TCTSer mutation in exon 6.

    Techniques: Expressing

    Observed phenotype following CDK2AP1 knockdown is p53 dependent. Panel A: Primary HDFs were transduced with EV or CDK2AP1-shRNA or CDK2AP1-shRNA and a p53 specific shRNA and were analyzed by qPCR for mRNA levels of p53 , p21 , BAX and PUMA . Knockdown of p53 with CDK2AP1 prevented the increase in p21, BAX and PUMA. Panel B: The knockdown of p53 with CDK2AP1 increased the percentage of cells in the S and reduced the cells in G1 when compared to CDK2AP1 only knockdown cells (p-value

    Journal: PLoS ONE

    Article Title: Knockdown of CDK2AP1 in Primary Human Fibroblasts Induces p53 Dependent Senescence

    doi: 10.1371/journal.pone.0120782

    Figure Lengend Snippet: Observed phenotype following CDK2AP1 knockdown is p53 dependent. Panel A: Primary HDFs were transduced with EV or CDK2AP1-shRNA or CDK2AP1-shRNA and a p53 specific shRNA and were analyzed by qPCR for mRNA levels of p53 , p21 , BAX and PUMA . Knockdown of p53 with CDK2AP1 prevented the increase in p21, BAX and PUMA. Panel B: The knockdown of p53 with CDK2AP1 increased the percentage of cells in the S and reduced the cells in G1 when compared to CDK2AP1 only knockdown cells (p-value

    Article Snippet: Commercially available primers for CDK2AP1 were obtained from Origene, MD, while other primers summarized in were obtained from Integrated DNA Technologies, IA.

    Techniques: Transduction, shRNA, Real-time Polymerase Chain Reaction, Significance Assay

    Knockdown of CDK2AP1 altered the morphology of HDFs and increased senescence-associated β-galactosidase activity. HDFs that were transduced with an empty vector (EV) or CDK2AP1-shRNA1 or CDK2AP1-shRNA2, were seeded and grown in a 6-well plate and senescence-associated β-galactosidase assay was performed. CDK2AP1 knockdown fibroblasts displayed a significantly higher senescence associated β-galactosidase activity. The percentage of cells exhibiting β-galactosidase activity is shown at the top of each panel in the right.

    Journal: PLoS ONE

    Article Title: Knockdown of CDK2AP1 in Primary Human Fibroblasts Induces p53 Dependent Senescence

    doi: 10.1371/journal.pone.0120782

    Figure Lengend Snippet: Knockdown of CDK2AP1 altered the morphology of HDFs and increased senescence-associated β-galactosidase activity. HDFs that were transduced with an empty vector (EV) or CDK2AP1-shRNA1 or CDK2AP1-shRNA2, were seeded and grown in a 6-well plate and senescence-associated β-galactosidase assay was performed. CDK2AP1 knockdown fibroblasts displayed a significantly higher senescence associated β-galactosidase activity. The percentage of cells exhibiting β-galactosidase activity is shown at the top of each panel in the right.

    Article Snippet: Commercially available primers for CDK2AP1 were obtained from Origene, MD, while other primers summarized in were obtained from Integrated DNA Technologies, IA.

    Techniques: Activity Assay, Transduction, Plasmid Preparation

    Knockdown of CDK2AP1 in HDFs reduces proliferation and induces G1 phase arrest. Panel A: Primary HDFs were transduced with an empty vector or with CDK2AP1 specific shRNA1 or 2. Following antibiotic selection, cells were examined for CDK2AP1 expression using qPCR. CDK2AP1 was successfully downregulated using both shRNA’s when compared to EV transduced cells. Panel B: HDFs were transduced with CDK2AP1 shRNA1 or a scrambled-sequence shRNA and stained with a CDK2AP1 specific antibody. Panel C: CDK2AP1 knockdown inhibited proliferation of HDFs. Cells were plated after selection, and counted at different time points by trypan blue exclusion method. Panel D: Asynchronized wild type and CDK2AP1 knockdown HDFs were harvested and stained with propidium iodide. DNA content was analyzed by flow cytometry. Results are presented together with standard deviation from experiments conducted in triplicate.

    Journal: PLoS ONE

    Article Title: Knockdown of CDK2AP1 in Primary Human Fibroblasts Induces p53 Dependent Senescence

    doi: 10.1371/journal.pone.0120782

    Figure Lengend Snippet: Knockdown of CDK2AP1 in HDFs reduces proliferation and induces G1 phase arrest. Panel A: Primary HDFs were transduced with an empty vector or with CDK2AP1 specific shRNA1 or 2. Following antibiotic selection, cells were examined for CDK2AP1 expression using qPCR. CDK2AP1 was successfully downregulated using both shRNA’s when compared to EV transduced cells. Panel B: HDFs were transduced with CDK2AP1 shRNA1 or a scrambled-sequence shRNA and stained with a CDK2AP1 specific antibody. Panel C: CDK2AP1 knockdown inhibited proliferation of HDFs. Cells were plated after selection, and counted at different time points by trypan blue exclusion method. Panel D: Asynchronized wild type and CDK2AP1 knockdown HDFs were harvested and stained with propidium iodide. DNA content was analyzed by flow cytometry. Results are presented together with standard deviation from experiments conducted in triplicate.

    Article Snippet: Commercially available primers for CDK2AP1 were obtained from Origene, MD, while other primers summarized in were obtained from Integrated DNA Technologies, IA.

    Techniques: Transduction, Plasmid Preparation, Selection, Expressing, Real-time Polymerase Chain Reaction, shRNA, Sequencing, Staining, Flow Cytometry, Cytometry, Standard Deviation

    Knockdown of CDK2AP1 increases the expression of p53, p21 and the Apoptotic Genes BAX and PUMA . Panel A: HDFs transduced with EV, CDK2AP1-shRNA1 or CDK2AP1-shRNA2 were harvested and analyzed for the mRNA expression of p53 , p21 , BAX and PUMA . Knockdown of CDK2AP1 in HDFs increased the expression of these genes. Panel B: Whole cell lysates were extracted from wild and knockdown HDFs and analyzed for p21 and p53 levels by Western Blot. Panel C: Whole cell lysates were extracted from wild and knockdown HDFs and analyzed for p14 ARF by Western Blot.

    Journal: PLoS ONE

    Article Title: Knockdown of CDK2AP1 in Primary Human Fibroblasts Induces p53 Dependent Senescence

    doi: 10.1371/journal.pone.0120782

    Figure Lengend Snippet: Knockdown of CDK2AP1 increases the expression of p53, p21 and the Apoptotic Genes BAX and PUMA . Panel A: HDFs transduced with EV, CDK2AP1-shRNA1 or CDK2AP1-shRNA2 were harvested and analyzed for the mRNA expression of p53 , p21 , BAX and PUMA . Knockdown of CDK2AP1 in HDFs increased the expression of these genes. Panel B: Whole cell lysates were extracted from wild and knockdown HDFs and analyzed for p21 and p53 levels by Western Blot. Panel C: Whole cell lysates were extracted from wild and knockdown HDFs and analyzed for p14 ARF by Western Blot.

    Article Snippet: Commercially available primers for CDK2AP1 were obtained from Origene, MD, while other primers summarized in were obtained from Integrated DNA Technologies, IA.

    Techniques: Expressing, Transduction, Western Blot

    Knockdown of CDK2AP1 in Primary HDFs Increases γ-H2AX Signal. Panel A: Primary HDFs were either transduced with empty vector (EV), CDK2AP1-shRNA1 or CDK2AP1-shRNA2. Following antibiotic selection, immunocytochemistry was carried out using a γ-H2AX specific antibody. Percentage of γ-H2AX was calculated in multiple (n = 28) randomly selected fields by dividing the number of γ-H2AX positive cells by total number of cells in the field. Results are presented together with standard deviation from multiple fields of view (*-p-value

    Journal: PLoS ONE

    Article Title: Knockdown of CDK2AP1 in Primary Human Fibroblasts Induces p53 Dependent Senescence

    doi: 10.1371/journal.pone.0120782

    Figure Lengend Snippet: Knockdown of CDK2AP1 in Primary HDFs Increases γ-H2AX Signal. Panel A: Primary HDFs were either transduced with empty vector (EV), CDK2AP1-shRNA1 or CDK2AP1-shRNA2. Following antibiotic selection, immunocytochemistry was carried out using a γ-H2AX specific antibody. Percentage of γ-H2AX was calculated in multiple (n = 28) randomly selected fields by dividing the number of γ-H2AX positive cells by total number of cells in the field. Results are presented together with standard deviation from multiple fields of view (*-p-value

    Article Snippet: Commercially available primers for CDK2AP1 were obtained from Origene, MD, while other primers summarized in were obtained from Integrated DNA Technologies, IA.

    Techniques: Transduction, Plasmid Preparation, Selection, Immunocytochemistry, Standard Deviation

    miR-432 targets SOCS5. For validation of SOCS5 UTR as target of miR-432, 100 pmol of miR-432 mimic was overexpressed in CHME3 cells. (A) Western blot showing downregulation of SOCS5 upon miR-432 overexpression in CHME3 cells. The average fold change values with respect to control have been mentioned. (B) Overexpression of miR-432 mimic was confirmed by Real time PCR using miR-432 specific TaqMan probe. RNU24 was used for normalization. (C) Blot showing upregulation of SOCS5 upon antimiR-432 transfection. The average fold change values with respect to control have been mentioned . (D) Silencing of miR-432 by antimiR-432 transfection was confirmed by real time PCR. (E) SOCS5 UTR clone (pEZX-MT06) for microRNA targeting was transfected along with miR-432 mimic along with β-galactosidase vector. Graph shows decreased luciferase activity of the vector in presence of miR-432 mimic. A mutant clone was generated deleting the targeting region present in SOCS5 UTR (shown in Fig. 2) and transfected along with miR-432 mimic. The mutant clone did not display any reduction in luciferase activity. Non-targeting mimic miR-146a was also used as negative control. β-galactosidase activity was used for normalization. All experiments were performed in triplicates. The data are shown as mean ± S.E from three independent experiments. The fold change is significant where *denotes P

    Journal: Scientific Reports

    Article Title: Japanese Encephalitis Virus exploits the microRNA-432 to regulate the expression of Suppressor of Cytokine Signaling (SOCS) 5

    doi: 10.1038/srep27685

    Figure Lengend Snippet: miR-432 targets SOCS5. For validation of SOCS5 UTR as target of miR-432, 100 pmol of miR-432 mimic was overexpressed in CHME3 cells. (A) Western blot showing downregulation of SOCS5 upon miR-432 overexpression in CHME3 cells. The average fold change values with respect to control have been mentioned. (B) Overexpression of miR-432 mimic was confirmed by Real time PCR using miR-432 specific TaqMan probe. RNU24 was used for normalization. (C) Blot showing upregulation of SOCS5 upon antimiR-432 transfection. The average fold change values with respect to control have been mentioned . (D) Silencing of miR-432 by antimiR-432 transfection was confirmed by real time PCR. (E) SOCS5 UTR clone (pEZX-MT06) for microRNA targeting was transfected along with miR-432 mimic along with β-galactosidase vector. Graph shows decreased luciferase activity of the vector in presence of miR-432 mimic. A mutant clone was generated deleting the targeting region present in SOCS5 UTR (shown in Fig. 2) and transfected along with miR-432 mimic. The mutant clone did not display any reduction in luciferase activity. Non-targeting mimic miR-146a was also used as negative control. β-galactosidase activity was used for normalization. All experiments were performed in triplicates. The data are shown as mean ± S.E from three independent experiments. The fold change is significant where *denotes P

    Article Snippet: The primers used to amplify SOCS5 from human genomic DNA are listed in .

    Techniques: Western Blot, Over Expression, Real-time Polymerase Chain Reaction, Transfection, Plasmid Preparation, Luciferase, Activity Assay, Mutagenesis, Generated, Negative Control

    JEV infection downregulates miR-432 levels and leads to SOCS5 upregulation. Human brain microglial cells were infected by JEV (MOI 5) and cells were harvested at various time points to determine miR-432 levels. (A) Graph showing downregulation of miR-432 levels at 24 and 48 hours post infection as compared to 12 hours sample. TaqMan probe specific to miR-432 were used to determine fold change. The C t values were normalized by RNU-24 levels. (B) Graph showing reduced levels of miR-432 in JEV infected mice brain. The RNA was isolated from brain tissue of 2 day and 4 day infected mice and mock infected mice brain was used as control. The C t values were normalized by RNU-6 levels. (C) Western blots depicting upregulation of SOCS5 upon JEV infection. CHME3 cells were infected by JEV (MOI 5) and harvested after 24 and 48 hours. The average fold change values with respect to control have been mentioned. (D) Western blots showing upregulation of SOCS5 in JEV infected brain tissue. Mock infected mice brain tissue was used as control. Infected mice were harvested 2 day and 4 day post infection. β-tubulin was used for normalization. The average fold change values with respect to control have been mentioned. (E) Immunohistochemistry image showing increased levels of SOCS5 protein in brain of JEV infected BALB/c mice. Mock and JEV infected mice brain sections were collected 2 days post JEV infection. Enhanced levels of Alexa 488 florescence was observed in JEV infected brain section (Magnification 20x, Scale bar 100 μm). All experiments were performed in triplicates. The data are shown as mean ± S.E from three independent experiments. The fold change is significant where *denotes P

    Journal: Scientific Reports

    Article Title: Japanese Encephalitis Virus exploits the microRNA-432 to regulate the expression of Suppressor of Cytokine Signaling (SOCS) 5

    doi: 10.1038/srep27685

    Figure Lengend Snippet: JEV infection downregulates miR-432 levels and leads to SOCS5 upregulation. Human brain microglial cells were infected by JEV (MOI 5) and cells were harvested at various time points to determine miR-432 levels. (A) Graph showing downregulation of miR-432 levels at 24 and 48 hours post infection as compared to 12 hours sample. TaqMan probe specific to miR-432 were used to determine fold change. The C t values were normalized by RNU-24 levels. (B) Graph showing reduced levels of miR-432 in JEV infected mice brain. The RNA was isolated from brain tissue of 2 day and 4 day infected mice and mock infected mice brain was used as control. The C t values were normalized by RNU-6 levels. (C) Western blots depicting upregulation of SOCS5 upon JEV infection. CHME3 cells were infected by JEV (MOI 5) and harvested after 24 and 48 hours. The average fold change values with respect to control have been mentioned. (D) Western blots showing upregulation of SOCS5 in JEV infected brain tissue. Mock infected mice brain tissue was used as control. Infected mice were harvested 2 day and 4 day post infection. β-tubulin was used for normalization. The average fold change values with respect to control have been mentioned. (E) Immunohistochemistry image showing increased levels of SOCS5 protein in brain of JEV infected BALB/c mice. Mock and JEV infected mice brain sections were collected 2 days post JEV infection. Enhanced levels of Alexa 488 florescence was observed in JEV infected brain section (Magnification 20x, Scale bar 100 μm). All experiments were performed in triplicates. The data are shown as mean ± S.E from three independent experiments. The fold change is significant where *denotes P

    Article Snippet: The primers used to amplify SOCS5 from human genomic DNA are listed in .

    Techniques: Infection, Mouse Assay, Isolation, Western Blot, Immunohistochemistry

    SOCS5 knock down restricts JEV replication. To decipher regulatory role of SOCS5 on JEV replication, siRNA mediated knock down of SOCS5 was done in CHME3 cells and cells were given JEV infection after 30 hours post transfection. (A) SOCS5 knock down was confirmed by western blotting. Scramble negative control sequence was transfected as control. The average fold change values with respect to control have been mentioned. (B) Western blot depicting enhanced STAT1 phosphorylation in SOCS5 knock down cells. The average fold change values with respect to control have been mentioned. (C) Viral NS1 protein blot showing downregulation of NS1 protein in SOCS5 knock down cells. The average fold change values with respect to control have been mentioned. (D) Real-time graph showing downregulation of viral RNA in SOCS5 knock down cells. RNA was isolated from the cells and viral RNA was determined by using viral NS3 specific primers. GAPDH was used for normalization. Fold change was calculated by 2 −∆∆C t method. JEV infected sample was used as control for statistical analysis. (E) Viral RNA was isolated from culture supernatant to determine copies of viral RNA released. The number of viral copies was calculated by running standard series of viral RNA and viral copies were determined by absolute quantification. Fold change in viral copies was calculated by assuming viral copies in supernatant of mock transfected (JEV infected) cells as control. (F) Graph showing enhanced ISRE activity in SOCS5 knock down cells upon JEV infection. All experiments were performed in triplicates. The data are shown as mean ± S.E from three independent experiments. The fold change is significant where *denotes P

    Journal: Scientific Reports

    Article Title: Japanese Encephalitis Virus exploits the microRNA-432 to regulate the expression of Suppressor of Cytokine Signaling (SOCS) 5

    doi: 10.1038/srep27685

    Figure Lengend Snippet: SOCS5 knock down restricts JEV replication. To decipher regulatory role of SOCS5 on JEV replication, siRNA mediated knock down of SOCS5 was done in CHME3 cells and cells were given JEV infection after 30 hours post transfection. (A) SOCS5 knock down was confirmed by western blotting. Scramble negative control sequence was transfected as control. The average fold change values with respect to control have been mentioned. (B) Western blot depicting enhanced STAT1 phosphorylation in SOCS5 knock down cells. The average fold change values with respect to control have been mentioned. (C) Viral NS1 protein blot showing downregulation of NS1 protein in SOCS5 knock down cells. The average fold change values with respect to control have been mentioned. (D) Real-time graph showing downregulation of viral RNA in SOCS5 knock down cells. RNA was isolated from the cells and viral RNA was determined by using viral NS3 specific primers. GAPDH was used for normalization. Fold change was calculated by 2 −∆∆C t method. JEV infected sample was used as control for statistical analysis. (E) Viral RNA was isolated from culture supernatant to determine copies of viral RNA released. The number of viral copies was calculated by running standard series of viral RNA and viral copies were determined by absolute quantification. Fold change in viral copies was calculated by assuming viral copies in supernatant of mock transfected (JEV infected) cells as control. (F) Graph showing enhanced ISRE activity in SOCS5 knock down cells upon JEV infection. All experiments were performed in triplicates. The data are shown as mean ± S.E from three independent experiments. The fold change is significant where *denotes P

    Article Snippet: The primers used to amplify SOCS5 from human genomic DNA are listed in .

    Techniques: Infection, Transfection, Western Blot, Negative Control, Sequencing, Isolation, Activity Assay

    miR-141 binding to solute carrier family 25 member 3 (Slc25a3). A : sequence homology between Slc25a3 3′-UTR ( top ) and miR-141 ( bottom ). Black lines represent perfect match, and grey lines represent G-U wobbles. B : miR-141 recognition sequence

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: miR-141 as a regulator of the mitochondrial phosphate carrier (Slc25a3) in the type 1 diabetic heart

    doi: 10.1152/ajpcell.00137.2012

    Figure Lengend Snippet: miR-141 binding to solute carrier family 25 member 3 (Slc25a3). A : sequence homology between Slc25a3 3′-UTR ( top ) and miR-141 ( bottom ). Black lines represent perfect match, and grey lines represent G-U wobbles. B : miR-141 recognition sequence

    Article Snippet: Briefly, 15 ng of cDNA sample, 0.5 μM of human miR-141 primer pairs (Origene), and 2× SYBR Green I qPCR Master Mix (Origene) were used to perform RTQ-PCR analysis using a Taqman 7900HT system (Applied Biosystems).

    Techniques: Binding Assay, Sequencing

    miRNA-141 overexpression and ATP synthase activity. A : ATP synthase activity was assessed in HL-1 control cells and miR-141 cells using an assay coupled with pyruvate kinase which converts ADP to ATP and produces pyruvate from phosphoenolpyruvate. Final

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: miR-141 as a regulator of the mitochondrial phosphate carrier (Slc25a3) in the type 1 diabetic heart

    doi: 10.1152/ajpcell.00137.2012

    Figure Lengend Snippet: miRNA-141 overexpression and ATP synthase activity. A : ATP synthase activity was assessed in HL-1 control cells and miR-141 cells using an assay coupled with pyruvate kinase which converts ADP to ATP and produces pyruvate from phosphoenolpyruvate. Final

    Article Snippet: Briefly, 15 ng of cDNA sample, 0.5 μM of human miR-141 primer pairs (Origene), and 2× SYBR Green I qPCR Master Mix (Origene) were used to perform RTQ-PCR analysis using a Taqman 7900HT system (Applied Biosystems).

    Techniques: Over Expression, Activity Assay

    miR-141 binding to the Slc25a3 3′-UTR target sequence. A : sequence homology between miR-141 and family member miR-200c. Bold underlined characters indicate differences in miR-200c nucleotide sequence compared with miR-141. B : relative luciferase

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: miR-141 as a regulator of the mitochondrial phosphate carrier (Slc25a3) in the type 1 diabetic heart

    doi: 10.1152/ajpcell.00137.2012

    Figure Lengend Snippet: miR-141 binding to the Slc25a3 3′-UTR target sequence. A : sequence homology between miR-141 and family member miR-200c. Bold underlined characters indicate differences in miR-200c nucleotide sequence compared with miR-141. B : relative luciferase

    Article Snippet: Briefly, 15 ng of cDNA sample, 0.5 μM of human miR-141 primer pairs (Origene), and 2× SYBR Green I qPCR Master Mix (Origene) were used to perform RTQ-PCR analysis using a Taqman 7900HT system (Applied Biosystems).

    Techniques: Binding Assay, Sequencing, Luciferase

    miR-141 overexpression. A : fold increase of miR-141 in HL-1 cells 72 h posttransfection with control (pCMV-MIR) or miR-141 expression plasmids. B : relative mRNA levels of Slc25a3 in HL-1 control cells and miR-141 cells. GAPDH was used as an internal control.

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: miR-141 as a regulator of the mitochondrial phosphate carrier (Slc25a3) in the type 1 diabetic heart

    doi: 10.1152/ajpcell.00137.2012

    Figure Lengend Snippet: miR-141 overexpression. A : fold increase of miR-141 in HL-1 cells 72 h posttransfection with control (pCMV-MIR) or miR-141 expression plasmids. B : relative mRNA levels of Slc25a3 in HL-1 control cells and miR-141 cells. GAPDH was used as an internal control.

    Article Snippet: Briefly, 15 ng of cDNA sample, 0.5 μM of human miR-141 primer pairs (Origene), and 2× SYBR Green I qPCR Master Mix (Origene) were used to perform RTQ-PCR analysis using a Taqman 7900HT system (Applied Biosystems).

    Techniques: Over Expression, Expressing