Structured Review

Promega rt pcr reaction mixture
Circulating DENV genome (RNAemia) in serum post-challenge with either DENV-1 or DENV-2. Serum samples from monkeys ( N = 5 per group) who received an adjuvanted TDENV PIV formulation (0.5 μg per type per dose) or PBS were obtained daily post-challenge with either ( A ) DENV-1 or ( B ) DENV-2 and tested for the presence of viral <t>RNA</t> (RNAemia) by <t>RT-PCR</t> assay. The dashed line represents the limit of quantitation (i.e., 360 GEQ/mL; therefore, values below this limit are approximations).
Rt Pcr Reaction Mixture, supplied by Promega, used in various techniques. Bioz Stars score: 91/100, based on 17 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/rt pcr reaction mixture/product/Promega
Average 91 stars, based on 17 article reviews
Price from $9.99 to $1999.99
rt pcr reaction mixture - by Bioz Stars, 2020-05
91/100 stars

Images

1) Product Images from "An Adjuvanted, Tetravalent Dengue Virus Purified Inactivated Vaccine Candidate Induces Long-Lasting and Protective Antibody Responses Against Dengue Challenge in Rhesus Macaques"

Article Title: An Adjuvanted, Tetravalent Dengue Virus Purified Inactivated Vaccine Candidate Induces Long-Lasting and Protective Antibody Responses Against Dengue Challenge in Rhesus Macaques

Journal: The American Journal of Tropical Medicine and Hygiene

doi: 10.4269/ajtmh.14-0268

Circulating DENV genome (RNAemia) in serum post-challenge with either DENV-1 or DENV-2. Serum samples from monkeys ( N = 5 per group) who received an adjuvanted TDENV PIV formulation (0.5 μg per type per dose) or PBS were obtained daily post-challenge with either ( A ) DENV-1 or ( B ) DENV-2 and tested for the presence of viral RNA (RNAemia) by RT-PCR assay. The dashed line represents the limit of quantitation (i.e., 360 GEQ/mL; therefore, values below this limit are approximations).
Figure Legend Snippet: Circulating DENV genome (RNAemia) in serum post-challenge with either DENV-1 or DENV-2. Serum samples from monkeys ( N = 5 per group) who received an adjuvanted TDENV PIV formulation (0.5 μg per type per dose) or PBS were obtained daily post-challenge with either ( A ) DENV-1 or ( B ) DENV-2 and tested for the presence of viral RNA (RNAemia) by RT-PCR assay. The dashed line represents the limit of quantitation (i.e., 360 GEQ/mL; therefore, values below this limit are approximations).

Techniques Used: Reverse Transcription Polymerase Chain Reaction, Quantitation Assay

Related Articles

Reverse Transcription Polymerase Chain Reaction:

Article Title: Generation of HIV-1 and Internal Control Transcripts as Standards for an In-House Quantitative Competitive RT-PCR Assay to Determine HIV-1 Viral Load
Article Snippet: .. We added 5 μ L of the isolated RNA to 25 μ L of an RT-PCR reaction mixture that contained 50 pmol of the HIV1-Q3 and HIV1-Biot-Q4 primers, Go Taq buffer, 1.5 mM MgCl2 , 250 μ M each dNTPs, 3.6 u of AMV-RT, 9 u of RNasin ribonuclease inhibitor, and 1.05 u of Go Taq Flexi DNA polymerase (all from Promega, USA). .. These reactions were carried out in a thermocycler (Eppendorf AG 22331, Hamburg, Germany) at 45°C for 30 min followed by 95°C for 3 min and 50 cycles at 95°C for 30 s, 58°C for 30 s, and 72°C for 30 s. The hybridization and detection of the amplified products were done in the ultramicroenzyme-linked oligosorbent assay (UMELOSA) format [ ].

Article Title: Differential Expression of Glypican-3 and Insulin–Like Growth Factor-II mRNAs and Alpha-Fetoprotein and Ki-67 Markers in HCV Related Hepatocellular Carcinomas In Egyptian Patients
Article Snippet: .. The gene-specific primers were synthesized by (Sigma Scientific Services Corporation, Cairo, Egypt) in the following sequence: GPC-3 mRNA primer sequence (5′-GATACAGCCAAAAGGCAG-3′-5′-ATCATTCCATCACCAGAG-3′) fragment size 250 (bp), IGF ll mRNA primer sequence (5′-CTTGGACTTTGAGTCAAATTGG-3′-5′-GGTCGTGCCAATTACATTTCA-3′) fragment size 292 (bp), and β actin mRNA primer sequence (5′-ACCATGGATGATGATATCGC-3′- 5′-ACATGGCTGGGGTGTTGAAG-3′), fragment size 381(bp), Five µl RNA was added to 20 µl RT-PCR reaction mix containing 4 µl of 10Mm dNTPs (Promega Corporation, Madison, WI 53711 USA), 1µl primers (Sigma Scientific Services Corporation, Cairo, Egypt), 20mM each and 5 µl of 10x buffer, 3µl of 25mM MgCl2 and 2.5U of Taq DNA polymerase (Promega Corporation, Madison, WI 53711 USA). .. The amplification was conducted using PCR thermal cycler (BioRAD-T100 ™ programmable thermal controller, Singapore) for 40 cycles (950C for 30 sec, 550C for 30 sec and 720C for 1min).

Article Title: The Inhibitory Effects of Low-Dose Ionizing Radiation in IgE-Mediated Allergic Responses
Article Snippet: .. The RT-PCR reaction mix was as follows: 10 μl cDNA (40 ng/μl), 1 μl forward primer, 1 μl reverse primer, 12 μl GoTaq qPCR master mix (Promega, WI, USA). .. PCR reactions were performed for 35 to 40 cycles in a 7500 Real-Time PCR System (Applied Biosystems, CA, USA).

Article Title: Haemophilus ducreyi Secretes a Filamentous Hemagglutinin-Like Protein
Article Snippet: .. Each RT-PCR reaction mixture (50 μl total volume) contained 1× RT-PCR buffer, a 150 μM concentration of each dNTP, 100 ng of each primer, 1.5 mM MgCl2 , 5 mM dithiothreitol, 8 U of RNasin (Promega), and 1 μl of RT-PCR enzyme mixture. ..

Article Title: An Adjuvanted, Tetravalent Dengue Virus Purified Inactivated Vaccine Candidate Induces Long-Lasting and Protective Antibody Responses Against Dengue Challenge in Rhesus Macaques
Article Snippet: .. Each RT-PCR reaction mixture contained 2.5 μL RNA template (corresponding to approximately 400 ng RNA per sample), 10 pmol either DENV-1 or DENV-2 forward/reverse primers, 5 pmol either DENV-1 or DENV-2 probes, 1.67 μL Detection Enhancer (from the kit), 20 U RNAse inhibitor (RNAsin; Promega, Madison, WI), 1× RT-PCR Buffer, and 1× RT-PCR Enzyme Mix (from the kit) in a total volume of 25 μL. ..

Article Title: Inhibition of cyclin E1 sensitizes hepatocellular carcinoma cells to regorafenib by mcl-1 suppression
Article Snippet: .. The RT-PCR reaction mixture contained 1 μg RNA and reverse transcriptase (Promega) with β-actin as the internal control. .. List of 5′ and 3′ primers for RT-PCR: β-actin: 5′-CTTAATGTCACGCACGATTTC-3′.

Article Title: Advancing age alters the expression of the ryanodine receptor 3 isoform in adult rat superior cervical ganglia
Article Snippet: .. RT-PCR reactions (50 μl) using RT-PCR reaction mix (Promega, Madison, WI) were performed with samples containing RNA ranging from 10 ng to 1 μg for optimization. .. To test for DNA contamination we performed negative controls, which consisted of the same RT-PCR reaction mix contained in the biological samples minus reverse transcriptase.

Article Title: The production of fibroblast growth factor 23 is controlled by TGF-β2
Article Snippet: .. For qRT-PCR analysis, the final volume of the RT-PCR reaction mixture was 20 µl and contained: 2 µl cDNA, 1 µM of each primer, 10 µl GoTaq® qPCR Master Mix (Promega), and sterile water up to 20 µl. .. PCR conditions were 95 °C for 3 min, followed by 40 cycles of 95 °C for 10 s, 58 °C for 30 s and 72 °C for 45 s. Quantitative RT-PCR was performed on a Rotor-Gene Q (QIAGEN, Hilden, Germany).

Synthesized:

Article Title: Differential Expression of Glypican-3 and Insulin–Like Growth Factor-II mRNAs and Alpha-Fetoprotein and Ki-67 Markers in HCV Related Hepatocellular Carcinomas In Egyptian Patients
Article Snippet: .. The gene-specific primers were synthesized by (Sigma Scientific Services Corporation, Cairo, Egypt) in the following sequence: GPC-3 mRNA primer sequence (5′-GATACAGCCAAAAGGCAG-3′-5′-ATCATTCCATCACCAGAG-3′) fragment size 250 (bp), IGF ll mRNA primer sequence (5′-CTTGGACTTTGAGTCAAATTGG-3′-5′-GGTCGTGCCAATTACATTTCA-3′) fragment size 292 (bp), and β actin mRNA primer sequence (5′-ACCATGGATGATGATATCGC-3′- 5′-ACATGGCTGGGGTGTTGAAG-3′), fragment size 381(bp), Five µl RNA was added to 20 µl RT-PCR reaction mix containing 4 µl of 10Mm dNTPs (Promega Corporation, Madison, WI 53711 USA), 1µl primers (Sigma Scientific Services Corporation, Cairo, Egypt), 20mM each and 5 µl of 10x buffer, 3µl of 25mM MgCl2 and 2.5U of Taq DNA polymerase (Promega Corporation, Madison, WI 53711 USA). .. The amplification was conducted using PCR thermal cycler (BioRAD-T100 ™ programmable thermal controller, Singapore) for 40 cycles (950C for 30 sec, 550C for 30 sec and 720C for 1min).

Isolation:

Article Title: Generation of HIV-1 and Internal Control Transcripts as Standards for an In-House Quantitative Competitive RT-PCR Assay to Determine HIV-1 Viral Load
Article Snippet: .. We added 5 μ L of the isolated RNA to 25 μ L of an RT-PCR reaction mixture that contained 50 pmol of the HIV1-Q3 and HIV1-Biot-Q4 primers, Go Taq buffer, 1.5 mM MgCl2 , 250 μ M each dNTPs, 3.6 u of AMV-RT, 9 u of RNasin ribonuclease inhibitor, and 1.05 u of Go Taq Flexi DNA polymerase (all from Promega, USA). .. These reactions were carried out in a thermocycler (Eppendorf AG 22331, Hamburg, Germany) at 45°C for 30 min followed by 95°C for 3 min and 50 cycles at 95°C for 30 s, 58°C for 30 s, and 72°C for 30 s. The hybridization and detection of the amplified products were done in the ultramicroenzyme-linked oligosorbent assay (UMELOSA) format [ ].

Gel Permeation Chromatography:

Article Title: Differential Expression of Glypican-3 and Insulin–Like Growth Factor-II mRNAs and Alpha-Fetoprotein and Ki-67 Markers in HCV Related Hepatocellular Carcinomas In Egyptian Patients
Article Snippet: .. The gene-specific primers were synthesized by (Sigma Scientific Services Corporation, Cairo, Egypt) in the following sequence: GPC-3 mRNA primer sequence (5′-GATACAGCCAAAAGGCAG-3′-5′-ATCATTCCATCACCAGAG-3′) fragment size 250 (bp), IGF ll mRNA primer sequence (5′-CTTGGACTTTGAGTCAAATTGG-3′-5′-GGTCGTGCCAATTACATTTCA-3′) fragment size 292 (bp), and β actin mRNA primer sequence (5′-ACCATGGATGATGATATCGC-3′- 5′-ACATGGCTGGGGTGTTGAAG-3′), fragment size 381(bp), Five µl RNA was added to 20 µl RT-PCR reaction mix containing 4 µl of 10Mm dNTPs (Promega Corporation, Madison, WI 53711 USA), 1µl primers (Sigma Scientific Services Corporation, Cairo, Egypt), 20mM each and 5 µl of 10x buffer, 3µl of 25mM MgCl2 and 2.5U of Taq DNA polymerase (Promega Corporation, Madison, WI 53711 USA). .. The amplification was conducted using PCR thermal cycler (BioRAD-T100 ™ programmable thermal controller, Singapore) for 40 cycles (950C for 30 sec, 550C for 30 sec and 720C for 1min).

Quantitative RT-PCR:

Article Title: The production of fibroblast growth factor 23 is controlled by TGF-β2
Article Snippet: .. For qRT-PCR analysis, the final volume of the RT-PCR reaction mixture was 20 µl and contained: 2 µl cDNA, 1 µM of each primer, 10 µl GoTaq® qPCR Master Mix (Promega), and sterile water up to 20 µl. .. PCR conditions were 95 °C for 3 min, followed by 40 cycles of 95 °C for 10 s, 58 °C for 30 s and 72 °C for 45 s. Quantitative RT-PCR was performed on a Rotor-Gene Q (QIAGEN, Hilden, Germany).

Real-time Polymerase Chain Reaction:

Article Title: The Inhibitory Effects of Low-Dose Ionizing Radiation in IgE-Mediated Allergic Responses
Article Snippet: .. The RT-PCR reaction mix was as follows: 10 μl cDNA (40 ng/μl), 1 μl forward primer, 1 μl reverse primer, 12 μl GoTaq qPCR master mix (Promega, WI, USA). .. PCR reactions were performed for 35 to 40 cycles in a 7500 Real-Time PCR System (Applied Biosystems, CA, USA).

Article Title: The production of fibroblast growth factor 23 is controlled by TGF-β2
Article Snippet: .. For qRT-PCR analysis, the final volume of the RT-PCR reaction mixture was 20 µl and contained: 2 µl cDNA, 1 µM of each primer, 10 µl GoTaq® qPCR Master Mix (Promega), and sterile water up to 20 µl. .. PCR conditions were 95 °C for 3 min, followed by 40 cycles of 95 °C for 10 s, 58 °C for 30 s and 72 °C for 45 s. Quantitative RT-PCR was performed on a Rotor-Gene Q (QIAGEN, Hilden, Germany).

Concentration Assay:

Article Title: Haemophilus ducreyi Secretes a Filamentous Hemagglutinin-Like Protein
Article Snippet: .. Each RT-PCR reaction mixture (50 μl total volume) contained 1× RT-PCR buffer, a 150 μM concentration of each dNTP, 100 ng of each primer, 1.5 mM MgCl2 , 5 mM dithiothreitol, 8 U of RNasin (Promega), and 1 μl of RT-PCR enzyme mixture. ..

Sequencing:

Article Title: Differential Expression of Glypican-3 and Insulin–Like Growth Factor-II mRNAs and Alpha-Fetoprotein and Ki-67 Markers in HCV Related Hepatocellular Carcinomas In Egyptian Patients
Article Snippet: .. The gene-specific primers were synthesized by (Sigma Scientific Services Corporation, Cairo, Egypt) in the following sequence: GPC-3 mRNA primer sequence (5′-GATACAGCCAAAAGGCAG-3′-5′-ATCATTCCATCACCAGAG-3′) fragment size 250 (bp), IGF ll mRNA primer sequence (5′-CTTGGACTTTGAGTCAAATTGG-3′-5′-GGTCGTGCCAATTACATTTCA-3′) fragment size 292 (bp), and β actin mRNA primer sequence (5′-ACCATGGATGATGATATCGC-3′- 5′-ACATGGCTGGGGTGTTGAAG-3′), fragment size 381(bp), Five µl RNA was added to 20 µl RT-PCR reaction mix containing 4 µl of 10Mm dNTPs (Promega Corporation, Madison, WI 53711 USA), 1µl primers (Sigma Scientific Services Corporation, Cairo, Egypt), 20mM each and 5 µl of 10x buffer, 3µl of 25mM MgCl2 and 2.5U of Taq DNA polymerase (Promega Corporation, Madison, WI 53711 USA). .. The amplification was conducted using PCR thermal cycler (BioRAD-T100 ™ programmable thermal controller, Singapore) for 40 cycles (950C for 30 sec, 550C for 30 sec and 720C for 1min).

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  • 91
    Promega rt pcr reaction mixture
    Circulating DENV genome (RNAemia) in serum post-challenge with either DENV-1 or DENV-2. Serum samples from monkeys ( N = 5 per group) who received an adjuvanted TDENV PIV formulation (0.5 μg per type per dose) or PBS were obtained daily post-challenge with either ( A ) DENV-1 or ( B ) DENV-2 and tested for the presence of viral <t>RNA</t> (RNAemia) by <t>RT-PCR</t> assay. The dashed line represents the limit of quantitation (i.e., 360 GEQ/mL; therefore, values below this limit are approximations).
    Rt Pcr Reaction Mixture, supplied by Promega, used in various techniques. Bioz Stars score: 91/100, based on 17 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rt pcr reaction mixture/product/Promega
    Average 91 stars, based on 17 article reviews
    Price from $9.99 to $1999.99
    rt pcr reaction mixture - by Bioz Stars, 2020-05
    91/100 stars
      Buy from Supplier

    85
    Promega firefly pgl4 10 luciferase reporter vectors
    Functional analysis of IRF-1-dependent TG promoter activity. a , constructs. 2.5-kb sequence, including the rs180195 A allele at the 5′ upstream TG gene, was cloned into <t>pGL4.10</t> vector to generate pGL4.10-TG(A). The rs180195 G allele as well as
    Firefly Pgl4 10 Luciferase Reporter Vectors, supplied by Promega, 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/firefly pgl4 10 luciferase reporter vectors/product/Promega
    Average 85 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    firefly pgl4 10 luciferase reporter vectors - by Bioz Stars, 2020-05
    85/100 stars
      Buy from Supplier

    92
    Promega rnase h
    R-Loops Display In Vitro Promoter Activity (A) R-loop plasmid construction: 32 P labeled (star) RNA (red line) was annealed to plasmid containing the G-rich β-actin gene 3′ end termination region (blue lines) flanked by S. cerevisiae URA3 reporter gene (green lines). An ethidium bromide stained gel (left panel) and autoradiograph (right panel) show R-loops as slower migrating species, sensitive to <t>RNase</t> H but not A or T1 treatments. D, plasmid DNA; R, RNA. (B) PCR amplification and sequencing of ten cloned plasmids following bisulfite treatment of S or AS R-loop plasmid. The upper reference line (in black) depicts every potential C to T (upper panel) or G to A (lower panel) conversion. Gray lines show individual clones with C-to-T or G-to-A changes, respectively. (C) Diagram showing S9.6 antibody immobilized on Dynabeads (black bar) and R-loop containing plasmids selected for transcription using HeLa nuclear extracts (NE). These were immuno-depleted for RNase H1 and H2A, as shown by western blot analysis. (D) Diagram and quantitation of qRT-PCR analysis on in vitro transcribed RNA. Strand-specific RT primers were used to distinguish transcript orientation, as indicated by arrows (see Table S2 ). Thick arrows denote more abundant transcripts. Data are represented as means ± SEM (n = 3; ∗ p
    Rnase H, supplied by Promega, used in various techniques. Bioz Stars score: 92/100, based on 71 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rnase h/product/Promega
    Average 92 stars, based on 71 article reviews
    Price from $9.99 to $1999.99
    rnase h - by Bioz Stars, 2020-05
    92/100 stars
      Buy from Supplier

    rna  (Promega)
    93
    Promega rna
    <t>Anti-DV2</t> activity of WSS45. (A) Cytotoxic effect of WSS45. Cells were exposed to compounds for 48 h and the cytotoxicity was evaluated by MTT method. It indicated that WSS45 slightly affected cell livability at 3 mg/mL. (B) Antiviral activity of WSS45 in BHK cells. DV2 (MOI=0.1) virus was added to confluent cells in the presence of compounds indicated. Supernatant viral <t>RNA</t> was extracted and quantified by qRT-PCR at 48 h post-infection. WSS45 dramatically reduced supernatant viral yield in a dose dependent manner. The results were expressed as Mean±SD of three independent experiments. b P
    Rna, supplied by Promega, used in various techniques. Bioz Stars score: 93/100, based on 205 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rna/product/Promega
    Average 93 stars, based on 205 article reviews
    Price from $9.99 to $1999.99
    rna - by Bioz Stars, 2020-05
    93/100 stars
      Buy from Supplier

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    Circulating DENV genome (RNAemia) in serum post-challenge with either DENV-1 or DENV-2. Serum samples from monkeys ( N = 5 per group) who received an adjuvanted TDENV PIV formulation (0.5 μg per type per dose) or PBS were obtained daily post-challenge with either ( A ) DENV-1 or ( B ) DENV-2 and tested for the presence of viral RNA (RNAemia) by RT-PCR assay. The dashed line represents the limit of quantitation (i.e., 360 GEQ/mL; therefore, values below this limit are approximations).

    Journal: The American Journal of Tropical Medicine and Hygiene

    Article Title: An Adjuvanted, Tetravalent Dengue Virus Purified Inactivated Vaccine Candidate Induces Long-Lasting and Protective Antibody Responses Against Dengue Challenge in Rhesus Macaques

    doi: 10.4269/ajtmh.14-0268

    Figure Lengend Snippet: Circulating DENV genome (RNAemia) in serum post-challenge with either DENV-1 or DENV-2. Serum samples from monkeys ( N = 5 per group) who received an adjuvanted TDENV PIV formulation (0.5 μg per type per dose) or PBS were obtained daily post-challenge with either ( A ) DENV-1 or ( B ) DENV-2 and tested for the presence of viral RNA (RNAemia) by RT-PCR assay. The dashed line represents the limit of quantitation (i.e., 360 GEQ/mL; therefore, values below this limit are approximations).

    Article Snippet: Each RT-PCR reaction mixture contained 2.5 μL RNA template (corresponding to approximately 400 ng RNA per sample), 10 pmol either DENV-1 or DENV-2 forward/reverse primers, 5 pmol either DENV-1 or DENV-2 probes, 1.67 μL Detection Enhancer (from the kit), 20 U RNAse inhibitor (RNAsin; Promega, Madison, WI), 1× RT-PCR Buffer, and 1× RT-PCR Enzyme Mix (from the kit) in a total volume of 25 μL.

    Techniques: Reverse Transcription Polymerase Chain Reaction, Quantitation Assay

    Functional analysis of IRF-1-dependent TG promoter activity. a , constructs. 2.5-kb sequence, including the rs180195 A allele at the 5′ upstream TG gene, was cloned into pGL4.10 vector to generate pGL4.10-TG(A). The rs180195 G allele as well as

    Journal: The Journal of Biological Chemistry

    Article Title: Novel Variant of Thyroglobulin Promoter Triggers Thyroid Autoimmunity through an Epigenetic Interferon ?-modulated Mechanism *

    doi: 10.1074/jbc.M111.247510

    Figure Lengend Snippet: Functional analysis of IRF-1-dependent TG promoter activity. a , constructs. 2.5-kb sequence, including the rs180195 A allele at the 5′ upstream TG gene, was cloned into pGL4.10 vector to generate pGL4.10-TG(A). The rs180195 G allele as well as

    Article Snippet: Cells were co-transfected with a mix of 200 ng of firefly pGL4.10 luciferase reporter vectors and 4 ng of Renilla luciferase reporter pGL4.74[ hRluc /TK] (Promega).

    Techniques: Functional Assay, Activity Assay, Construct, Sequencing, Clone Assay, Plasmid Preparation

    a , reduced IRF-1 mRNA expression in ML-1 cells treated with IRF-1 siRNA. mRNA expression of IRF-1 was assessed by regular RT-PCR and QRT-PCR. b , knockdown of IRF-1 by siRNA reduces the TG promoter activity only in the presence of rs180195 G allele. pGL4.10-TG(A)

    Journal: The Journal of Biological Chemistry

    Article Title: Novel Variant of Thyroglobulin Promoter Triggers Thyroid Autoimmunity through an Epigenetic Interferon ?-modulated Mechanism *

    doi: 10.1074/jbc.M111.247510

    Figure Lengend Snippet: a , reduced IRF-1 mRNA expression in ML-1 cells treated with IRF-1 siRNA. mRNA expression of IRF-1 was assessed by regular RT-PCR and QRT-PCR. b , knockdown of IRF-1 by siRNA reduces the TG promoter activity only in the presence of rs180195 G allele. pGL4.10-TG(A)

    Article Snippet: Cells were co-transfected with a mix of 200 ng of firefly pGL4.10 luciferase reporter vectors and 4 ng of Renilla luciferase reporter pGL4.74[ hRluc /TK] (Promega).

    Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Quantitative RT-PCR, Activity Assay

    INFα stimulates expression of TG in the presence of the disease-associated variant. a , specific effect of IFNα on pGL4.10-TG(G) construct activity. Treatment of ML-1 cells with IFNα increased the TG promoter activity when G allele

    Journal: The Journal of Biological Chemistry

    Article Title: Novel Variant of Thyroglobulin Promoter Triggers Thyroid Autoimmunity through an Epigenetic Interferon ?-modulated Mechanism *

    doi: 10.1074/jbc.M111.247510

    Figure Lengend Snippet: INFα stimulates expression of TG in the presence of the disease-associated variant. a , specific effect of IFNα on pGL4.10-TG(G) construct activity. Treatment of ML-1 cells with IFNα increased the TG promoter activity when G allele

    Article Snippet: Cells were co-transfected with a mix of 200 ng of firefly pGL4.10 luciferase reporter vectors and 4 ng of Renilla luciferase reporter pGL4.74[ hRluc /TK] (Promega).

    Techniques: Expressing, Variant Assay, Construct, Activity Assay

    R-Loops Display In Vitro Promoter Activity (A) R-loop plasmid construction: 32 P labeled (star) RNA (red line) was annealed to plasmid containing the G-rich β-actin gene 3′ end termination region (blue lines) flanked by S. cerevisiae URA3 reporter gene (green lines). An ethidium bromide stained gel (left panel) and autoradiograph (right panel) show R-loops as slower migrating species, sensitive to RNase H but not A or T1 treatments. D, plasmid DNA; R, RNA. (B) PCR amplification and sequencing of ten cloned plasmids following bisulfite treatment of S or AS R-loop plasmid. The upper reference line (in black) depicts every potential C to T (upper panel) or G to A (lower panel) conversion. Gray lines show individual clones with C-to-T or G-to-A changes, respectively. (C) Diagram showing S9.6 antibody immobilized on Dynabeads (black bar) and R-loop containing plasmids selected for transcription using HeLa nuclear extracts (NE). These were immuno-depleted for RNase H1 and H2A, as shown by western blot analysis. (D) Diagram and quantitation of qRT-PCR analysis on in vitro transcribed RNA. Strand-specific RT primers were used to distinguish transcript orientation, as indicated by arrows (see Table S2 ). Thick arrows denote more abundant transcripts. Data are represented as means ± SEM (n = 3; ∗ p

    Journal: Molecular Cell

    Article Title: R-Loops Promote Antisense Transcription across the Mammalian Genome

    doi: 10.1016/j.molcel.2019.10.002

    Figure Lengend Snippet: R-Loops Display In Vitro Promoter Activity (A) R-loop plasmid construction: 32 P labeled (star) RNA (red line) was annealed to plasmid containing the G-rich β-actin gene 3′ end termination region (blue lines) flanked by S. cerevisiae URA3 reporter gene (green lines). An ethidium bromide stained gel (left panel) and autoradiograph (right panel) show R-loops as slower migrating species, sensitive to RNase H but not A or T1 treatments. D, plasmid DNA; R, RNA. (B) PCR amplification and sequencing of ten cloned plasmids following bisulfite treatment of S or AS R-loop plasmid. The upper reference line (in black) depicts every potential C to T (upper panel) or G to A (lower panel) conversion. Gray lines show individual clones with C-to-T or G-to-A changes, respectively. (C) Diagram showing S9.6 antibody immobilized on Dynabeads (black bar) and R-loop containing plasmids selected for transcription using HeLa nuclear extracts (NE). These were immuno-depleted for RNase H1 and H2A, as shown by western blot analysis. (D) Diagram and quantitation of qRT-PCR analysis on in vitro transcribed RNA. Strand-specific RT primers were used to distinguish transcript orientation, as indicated by arrows (see Table S2 ). Thick arrows denote more abundant transcripts. Data are represented as means ± SEM (n = 3; ∗ p

    Article Snippet: Where indicated, the mixture was further digested with RNase H (Promega), RNase A (Sigma) or RNase T1 (Invitrogen).

    Techniques: In Vitro, Activity Assay, Plasmid Preparation, Labeling, Staining, Autoradiography, Polymerase Chain Reaction, Amplification, Sequencing, Clone Assay, Western Blot, Quantitation Assay, Quantitative RT-PCR

    Anti-DV2 activity of WSS45. (A) Cytotoxic effect of WSS45. Cells were exposed to compounds for 48 h and the cytotoxicity was evaluated by MTT method. It indicated that WSS45 slightly affected cell livability at 3 mg/mL. (B) Antiviral activity of WSS45 in BHK cells. DV2 (MOI=0.1) virus was added to confluent cells in the presence of compounds indicated. Supernatant viral RNA was extracted and quantified by qRT-PCR at 48 h post-infection. WSS45 dramatically reduced supernatant viral yield in a dose dependent manner. The results were expressed as Mean±SD of three independent experiments. b P

    Journal: Acta Pharmacologica Sinica

    Article Title: WSS45, a sulfated α-D-glucan, strongly interferes with Dengue 2 virus infection in vitro

    doi: 10.1038/aps.2010.29

    Figure Lengend Snippet: Anti-DV2 activity of WSS45. (A) Cytotoxic effect of WSS45. Cells were exposed to compounds for 48 h and the cytotoxicity was evaluated by MTT method. It indicated that WSS45 slightly affected cell livability at 3 mg/mL. (B) Antiviral activity of WSS45 in BHK cells. DV2 (MOI=0.1) virus was added to confluent cells in the presence of compounds indicated. Supernatant viral RNA was extracted and quantified by qRT-PCR at 48 h post-infection. WSS45 dramatically reduced supernatant viral yield in a dose dependent manner. The results were expressed as Mean±SD of three independent experiments. b P

    Article Snippet: A 20 μL reaction mixture contained 8 μL of extracted RNA, 1 μmol/L DV2.L1, 1×reaction buffer (Promega, USA), 2 U MMLV reverse transcriptase (Promega, USA), 300 μmol/L each deoxyribonucleoside (dNTP), and 5 mmol/L Mg2+ .

    Techniques: Activity Assay, MTT Assay, Quantitative RT-PCR, Infection