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DaAn Gene hcv rna
Hcv Rna, supplied by DaAn Gene, used in various techniques. Bioz Stars score: 92/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 92 stars, based on 2 article reviews
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Real-time Polymerase Chain Reaction:

Article Title: Anti-HCV reactive volunteer blood donors distribution character and genotypes switch in Xi'an, China
Article Snippet: .. After that, real time PCR was used to detect HCV RNA according the manufacture's manual (Daan Gene, Shenzhen, China) with standard controls [ , ]. .. Briefly, extracted RNA was measured with fluorescence labeled and self-quench probe and Perkin Elme PCR analyzer (PTC-200, Perkin Elmer, Covina, USA).

Polymerase Chain Reaction:

Article Title: Expansion of myeloid-derived suppressor cells from peripheral blood decreases after 4-week antiviral treatment in patients with chronic hepatitis C
Article Snippet: .. HCV RNA was tested by reverse transcriptase polymerase chain reaction assay (DAAN Gene, Sun Yat-Sen University, China). ..

Article Title: Prevalence and Determinants of Cryptosporidium Infection in an Underdeveloped Rural Region of Southwestern China
Article Snippet: .. Plasma viral loads of HBV, HCV, and HIV were measured by using the diagnostic kit for quantification of HBV DNA, HCV RNA, and HIV-1 RNA (PCR-Fluorescence Probing) (DaAn Gene Group Inc., Zhongshan, China) in the Center for Tropical Disease Research at Fudan University. ..

Diagnostic Assay:

Article Title: Prevalence and Determinants of Cryptosporidium Infection in an Underdeveloped Rural Region of Southwestern China
Article Snippet: .. Plasma viral loads of HBV, HCV, and HIV were measured by using the diagnostic kit for quantification of HBV DNA, HCV RNA, and HIV-1 RNA (PCR-Fluorescence Probing) (DaAn Gene Group Inc., Zhongshan, China) in the Center for Tropical Disease Research at Fudan University. ..

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    DaAn Gene hcv rna level
    Combinations of mutations promoted CH6a Core-NS5A recombinants to replicate efficiently in cultured cells. (A) Schematic diagrams of CH6a C-5A recombinants with different mutations (7m, 11m, 15m, and 18m). 7m was combinations of F1469L/A1677S (LS) ( Li et al., 2012a ), V1555L/I1720F/L1795M from HK6a 5-5A virus ( Li Y.P. et al., 2014 ), and K1303R/K1696R from the consensus 6a subgenomic replicon ( Yu et al., 2014 ). 4m (I355M/N416S/I831V/L881F; in green color) was identified from the Core-NS2 recombinant (Table 1 ). Mutations in red color were identified from the passaged C-5A_11m viruses (Table 2 ). (B) <t>RNA</t> transcripts of C-5A recombinants with indicated mutations were transfected into Huh7.5.1-VISI-mCherry cells ( Zhao et al., 2017 ), and the percentage of mCherry-positive cells was estimated (left y -axis; lines). <t>HCV</t> infectivity titers in culture supernatants at peak infection were determined (mean of triplicate infections ± SD, right y -axis; bars). Transfections were done in different experiments, and the data were combined in the graph. (C) C-5A_11m virus collected from days 33, 35, and 37 (in panel B ) were serially passaged to naïve cells. Virus spread and peak infectivity titers for passages 1, 3, 5, and 8 are shown for one of two independent infections performed in parallel; similar virus spread and infectivity titers were recorded for the other infected culture. ORF sequence analyses of the passaged viruses collected from peak infections are shown in Table 2 .
    Hcv Rna Level, supplied by DaAn Gene, used in various techniques. Bioz Stars score: 93/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hcv rna level/product/DaAn Gene
    Average 93 stars, based on 5 article reviews
    Price from $9.99 to $1999.99
    hcv rna level - by Bioz Stars, 2020-05
    93/100 stars
      Buy from Supplier

    90
    DaAn Gene hcv rna polymerase chain reaction pcr
    GADD45α plays a vital role in regulating FFA-mediated <t>HCV</t> replication. Huh7 cells were transfected with GADD45α siRNA, a negative control siRNA (scramble), or GADD45α over-expressed plasmid. Forty-eight hours later, GADD45α expression was identified ( A ), and cells were then infected with HCV and simultaneously treated with FFA (0.8 mM). After 72 h, the relative HCV-NS3 protein and <t>RNA</t> expression were detected by Western blotting and <t>RT-PCR</t> ( A ); intracellular lipid accumulation was detected by Bodipy-493/503 staining and visualized after 400× amplification ( B ). Results are shown as mean ±SD of three independent experiments.
    Hcv Rna Polymerase Chain Reaction Pcr, supplied by DaAn Gene, 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/hcv rna polymerase chain reaction pcr/product/DaAn Gene
    Average 90 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    hcv rna polymerase chain reaction pcr - by Bioz Stars, 2020-05
    90/100 stars
      Buy from Supplier

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    Combinations of mutations promoted CH6a Core-NS5A recombinants to replicate efficiently in cultured cells. (A) Schematic diagrams of CH6a C-5A recombinants with different mutations (7m, 11m, 15m, and 18m). 7m was combinations of F1469L/A1677S (LS) ( Li et al., 2012a ), V1555L/I1720F/L1795M from HK6a 5-5A virus ( Li Y.P. et al., 2014 ), and K1303R/K1696R from the consensus 6a subgenomic replicon ( Yu et al., 2014 ). 4m (I355M/N416S/I831V/L881F; in green color) was identified from the Core-NS2 recombinant (Table 1 ). Mutations in red color were identified from the passaged C-5A_11m viruses (Table 2 ). (B) RNA transcripts of C-5A recombinants with indicated mutations were transfected into Huh7.5.1-VISI-mCherry cells ( Zhao et al., 2017 ), and the percentage of mCherry-positive cells was estimated (left y -axis; lines). HCV infectivity titers in culture supernatants at peak infection were determined (mean of triplicate infections ± SD, right y -axis; bars). Transfections were done in different experiments, and the data were combined in the graph. (C) C-5A_11m virus collected from days 33, 35, and 37 (in panel B ) were serially passaged to naïve cells. Virus spread and peak infectivity titers for passages 1, 3, 5, and 8 are shown for one of two independent infections performed in parallel; similar virus spread and infectivity titers were recorded for the other infected culture. ORF sequence analyses of the passaged viruses collected from peak infections are shown in Table 2 .

    Journal: Frontiers in Microbiology

    Article Title: Development of an Infectious Cell Culture System for Hepatitis C Virus Genotype 6a Clinical Isolate Using a Novel Strategy and Its Sensitivity to Direct-Acting Antivirals

    doi: 10.3389/fmicb.2018.02950

    Figure Lengend Snippet: Combinations of mutations promoted CH6a Core-NS5A recombinants to replicate efficiently in cultured cells. (A) Schematic diagrams of CH6a C-5A recombinants with different mutations (7m, 11m, 15m, and 18m). 7m was combinations of F1469L/A1677S (LS) ( Li et al., 2012a ), V1555L/I1720F/L1795M from HK6a 5-5A virus ( Li Y.P. et al., 2014 ), and K1303R/K1696R from the consensus 6a subgenomic replicon ( Yu et al., 2014 ). 4m (I355M/N416S/I831V/L881F; in green color) was identified from the Core-NS2 recombinant (Table 1 ). Mutations in red color were identified from the passaged C-5A_11m viruses (Table 2 ). (B) RNA transcripts of C-5A recombinants with indicated mutations were transfected into Huh7.5.1-VISI-mCherry cells ( Zhao et al., 2017 ), and the percentage of mCherry-positive cells was estimated (left y -axis; lines). HCV infectivity titers in culture supernatants at peak infection were determined (mean of triplicate infections ± SD, right y -axis; bars). Transfections were done in different experiments, and the data were combined in the graph. (C) C-5A_11m virus collected from days 33, 35, and 37 (in panel B ) were serially passaged to naïve cells. Virus spread and peak infectivity titers for passages 1, 3, 5, and 8 are shown for one of two independent infections performed in parallel; similar virus spread and infectivity titers were recorded for the other infected culture. ORF sequence analyses of the passaged viruses collected from peak infections are shown in Table 2 .

    Article Snippet: HCV Infected Serum and Plasmids The HCV RNA level in the patient serum was determined by the HCV Nucleic Acid Detection Kit (TaqMan) (Daangene, China).

    Techniques: Cell Culture, Recombinant, Transfection, Infection, Sequencing

    Culture adaptation of CH6a Core-NS2 recombinant identified mutations that enhanced virus production. RNA transcripts of CH6a C-NS2 recombinants (wild-type or with mutations as indicated in panel A ) were transfected into Huh7.5.1 cells, either HCV Core or NS5A-EGFP antigens were detected by immunostaining or directly visualizing under fluorescence microscope, respectively. Percentage of HCV-positive cells was estimated (left y -axis; shown as line plots). HCV infectivity titers in the supernatant at peak of infection (≥80% HCV-positive cells) were determined by focus-forming-unit (FFU) assay [mean of triplicate infections ± standard deviation (SD), right y-axis; shown as bar graphs]. (B) Transfection of wild-type CH6a C-NS2 recombinant. The supernatant of C-NS2 virus collected from days 25 and 27 were inoculated to naïve cells to generate passage virus, and consecutively we generated the fifth- and ninth-passage viruses for sequence analysis (Table 1 ). (C) Transfection of CH6a C-NS2_4m, which contained four mutations (4m, I355M/N416S/I831V/L881F) identified in the C-NS2 virus of the fifth-passage (Table 1 ).

    Journal: Frontiers in Microbiology

    Article Title: Development of an Infectious Cell Culture System for Hepatitis C Virus Genotype 6a Clinical Isolate Using a Novel Strategy and Its Sensitivity to Direct-Acting Antivirals

    doi: 10.3389/fmicb.2018.02950

    Figure Lengend Snippet: Culture adaptation of CH6a Core-NS2 recombinant identified mutations that enhanced virus production. RNA transcripts of CH6a C-NS2 recombinants (wild-type or with mutations as indicated in panel A ) were transfected into Huh7.5.1 cells, either HCV Core or NS5A-EGFP antigens were detected by immunostaining or directly visualizing under fluorescence microscope, respectively. Percentage of HCV-positive cells was estimated (left y -axis; shown as line plots). HCV infectivity titers in the supernatant at peak of infection (≥80% HCV-positive cells) were determined by focus-forming-unit (FFU) assay [mean of triplicate infections ± standard deviation (SD), right y-axis; shown as bar graphs]. (B) Transfection of wild-type CH6a C-NS2 recombinant. The supernatant of C-NS2 virus collected from days 25 and 27 were inoculated to naïve cells to generate passage virus, and consecutively we generated the fifth- and ninth-passage viruses for sequence analysis (Table 1 ). (C) Transfection of CH6a C-NS2_4m, which contained four mutations (4m, I355M/N416S/I831V/L881F) identified in the C-NS2 virus of the fifth-passage (Table 1 ).

    Article Snippet: HCV Infected Serum and Plasmids The HCV RNA level in the patient serum was determined by the HCV Nucleic Acid Detection Kit (TaqMan) (Daangene, China).

    Techniques: Recombinant, Transfection, Immunostaining, Fluorescence, Microscopy, Infection, Standard Deviation, Generated, Sequencing

    Effects of newly identified adaptive mutations on the viability of CH6acc. (A) RNA transcripts of CH6acc and CH6acc with each of 12 putative adaptive mutations, which were newly identified in this study, mutated back to the wild-type sequence were transfected into Huh7.5 cells. Virus spread (% HCV antigen- positive cells) was determined by anti-HCV Core immunostaining (left y -axis; lines), and HCV infectivity titers (FFU/ml) in supernatants from cultures at day 5, 7, and 9 were determined and shown as the mean from triplicate infections ± SEM (the standard error of the mean). The transfection experiments were performed in two independent experiments, and the representative data from one experiment were shown. (B) Intracellular HCV Core antigen levels were determined by Western blotting. Cell lysates were separated through acrylamide gels, and proteins were transferred to PVDF membranes and immunoblotted with anti-HCV Core C7-50 antibody for detection of HCV Core and anti-β-actin for detection of host cellular actin.

    Journal: Frontiers in Microbiology

    Article Title: Development of an Infectious Cell Culture System for Hepatitis C Virus Genotype 6a Clinical Isolate Using a Novel Strategy and Its Sensitivity to Direct-Acting Antivirals

    doi: 10.3389/fmicb.2018.02950

    Figure Lengend Snippet: Effects of newly identified adaptive mutations on the viability of CH6acc. (A) RNA transcripts of CH6acc and CH6acc with each of 12 putative adaptive mutations, which were newly identified in this study, mutated back to the wild-type sequence were transfected into Huh7.5 cells. Virus spread (% HCV antigen- positive cells) was determined by anti-HCV Core immunostaining (left y -axis; lines), and HCV infectivity titers (FFU/ml) in supernatants from cultures at day 5, 7, and 9 were determined and shown as the mean from triplicate infections ± SEM (the standard error of the mean). The transfection experiments were performed in two independent experiments, and the representative data from one experiment were shown. (B) Intracellular HCV Core antigen levels were determined by Western blotting. Cell lysates were separated through acrylamide gels, and proteins were transferred to PVDF membranes and immunoblotted with anti-HCV Core C7-50 antibody for detection of HCV Core and anti-β-actin for detection of host cellular actin.

    Article Snippet: HCV Infected Serum and Plasmids The HCV RNA level in the patient serum was determined by the HCV Nucleic Acid Detection Kit (TaqMan) (Daangene, China).

    Techniques: Sequencing, Transfection, Immunostaining, Infection, Western Blot

    Functional analysis of the role of newly identified CH6acc adaptive mutations in the HCV life cycle. Equal amount of RNA transcripts (5 μg) from CH6acc and twelve CH6acc mutants with single mutation changed back to original sequence were transfected into HCV entry-deficient S29 cells ( Russell et al., 2008 ). Cell lysates (intracellular) and culture supernatant (extracellular) were collected 48 h post transfection. HCV infectivity titers, RNA copies, and Core levels were determined and normalized to replication-independent genome, J6/JFH1-GND. (A) Intracellular and extracellular HCV infectivity titers. ∗ , no FFU was detected by manual count. Values are expressed as log 10 FFU/ml (45 μl loaded) for extracellular titers and as log 10 FFU/well (1/8 cell lysate of a well of 6-well-plate) for intracellular infectivity titers. (B) HCV RNA levels in the cells and supernatant. The method for the determination of RNA levels were as described previously ( Boson et al., 2011 ). (C) HCV Core levels. Western blotting was performed on the cell lysates and supernatant harvested 48 h post transfection.

    Journal: Frontiers in Microbiology

    Article Title: Development of an Infectious Cell Culture System for Hepatitis C Virus Genotype 6a Clinical Isolate Using a Novel Strategy and Its Sensitivity to Direct-Acting Antivirals

    doi: 10.3389/fmicb.2018.02950

    Figure Lengend Snippet: Functional analysis of the role of newly identified CH6acc adaptive mutations in the HCV life cycle. Equal amount of RNA transcripts (5 μg) from CH6acc and twelve CH6acc mutants with single mutation changed back to original sequence were transfected into HCV entry-deficient S29 cells ( Russell et al., 2008 ). Cell lysates (intracellular) and culture supernatant (extracellular) were collected 48 h post transfection. HCV infectivity titers, RNA copies, and Core levels were determined and normalized to replication-independent genome, J6/JFH1-GND. (A) Intracellular and extracellular HCV infectivity titers. ∗ , no FFU was detected by manual count. Values are expressed as log 10 FFU/ml (45 μl loaded) for extracellular titers and as log 10 FFU/well (1/8 cell lysate of a well of 6-well-plate) for intracellular infectivity titers. (B) HCV RNA levels in the cells and supernatant. The method for the determination of RNA levels were as described previously ( Boson et al., 2011 ). (C) HCV Core levels. Western blotting was performed on the cell lysates and supernatant harvested 48 h post transfection.

    Article Snippet: HCV Infected Serum and Plasmids The HCV RNA level in the patient serum was determined by the HCV Nucleic Acid Detection Kit (TaqMan) (Daangene, China).

    Techniques: Functional Assay, Mutagenesis, Sequencing, Transfection, Infection, Western Blot

    CH6a full-length clone assembled from the consensus-like PCR fragments was more efficient in virus production. Two amino acids, S2362G in NS5A and N2738D in NS5B of CH6acc genome were mutated to the consensus sequence, designated CH6acc/Cons. RNA transcripts were transfected into Huh7.5 cells, and CH6acc was included in parallel. CH6acc/Cons was delayed in virus spread and attenuated in infectivity titers. HCV-positive cells were visualized by secure anti-HCV Core immunostaining (left y -axis; lines), and HCV infectivity titers were done by FFU assay (mean of triplicate infections ± SEM, right y -axis; bars). In other transfections, CH6aFL/Cons recombinant was non-viable.

    Journal: Frontiers in Microbiology

    Article Title: Development of an Infectious Cell Culture System for Hepatitis C Virus Genotype 6a Clinical Isolate Using a Novel Strategy and Its Sensitivity to Direct-Acting Antivirals

    doi: 10.3389/fmicb.2018.02950

    Figure Lengend Snippet: CH6a full-length clone assembled from the consensus-like PCR fragments was more efficient in virus production. Two amino acids, S2362G in NS5A and N2738D in NS5B of CH6acc genome were mutated to the consensus sequence, designated CH6acc/Cons. RNA transcripts were transfected into Huh7.5 cells, and CH6acc was included in parallel. CH6acc/Cons was delayed in virus spread and attenuated in infectivity titers. HCV-positive cells were visualized by secure anti-HCV Core immunostaining (left y -axis; lines), and HCV infectivity titers were done by FFU assay (mean of triplicate infections ± SEM, right y -axis; bars). In other transfections, CH6aFL/Cons recombinant was non-viable.

    Article Snippet: HCV Infected Serum and Plasmids The HCV RNA level in the patient serum was determined by the HCV Nucleic Acid Detection Kit (TaqMan) (Daangene, China).

    Techniques: Polymerase Chain Reaction, Sequencing, Transfection, Infection, Immunostaining, Recombinant

    Effects of newly identified adaptive mutations on the viability of CH6acc. (A) RNA transcripts of CH6acc and CH6acc with each of 12 putative adaptive mutations, which were newly identified in this study, mutated back to the wild-type sequence were transfected into Huh7.5 cells. Virus spread (% HCV antigen- positive cells) was determined by anti-HCV Core immunostaining (left y -axis; lines), and HCV infectivity titers (FFU/ml) in supernatants from cultures at day 5, 7, and 9 were determined and shown as the mean from triplicate infections ± SEM (the standard error of the mean). The transfection experiments were performed in two independent experiments, and the representative data from one experiment were shown. (B) Intracellular HCV Core antigen levels were determined by Western blotting. Cell lysates were separated through acrylamide gels, and proteins were transferred to PVDF membranes and immunoblotted with anti-HCV Core C7-50 antibody for detection of HCV Core and anti-β-actin for detection of host cellular actin.

    Journal: Frontiers in Microbiology

    Article Title: Development of an Infectious Cell Culture System for Hepatitis C Virus Genotype 6a Clinical Isolate Using a Novel Strategy and Its Sensitivity to Direct-Acting Antivirals

    doi: 10.3389/fmicb.2018.02950

    Figure Lengend Snippet: Effects of newly identified adaptive mutations on the viability of CH6acc. (A) RNA transcripts of CH6acc and CH6acc with each of 12 putative adaptive mutations, which were newly identified in this study, mutated back to the wild-type sequence were transfected into Huh7.5 cells. Virus spread (% HCV antigen- positive cells) was determined by anti-HCV Core immunostaining (left y -axis; lines), and HCV infectivity titers (FFU/ml) in supernatants from cultures at day 5, 7, and 9 were determined and shown as the mean from triplicate infections ± SEM (the standard error of the mean). The transfection experiments were performed in two independent experiments, and the representative data from one experiment were shown. (B) Intracellular HCV Core antigen levels were determined by Western blotting. Cell lysates were separated through acrylamide gels, and proteins were transferred to PVDF membranes and immunoblotted with anti-HCV Core C7-50 antibody for detection of HCV Core and anti-β-actin for detection of host cellular actin.

    Article Snippet: The HCV RNA level in the patient serum was determined by the HCV Nucleic Acid Detection Kit (TaqMan) (Daangene, China).

    Techniques: Sequencing, Transfection, Immunostaining, Infection, Western Blot

    ). Cell lysates (intracellular) and culture supernatant (extracellular) were collected 48 h post transfection. HCV infectivity titers, RNA copies, and Core levels were determined and normalized to replication-independent genome, J6/JFH1-GND. (A) Intracellular and extracellular HCV infectivity titers. ∗ , no FFU was detected by manual count. Values are expressed as log 10 FFU/ml (45 μl loaded) for extracellular titers and as log 10 FFU/well (1/8 cell lysate of a well of 6-well-plate) for intracellular infectivity titers. (B) ). (C) HCV Core levels. Western blotting was performed on the cell lysates and supernatant harvested 48 h post transfection.

    Journal: Frontiers in Microbiology

    Article Title: Development of an Infectious Cell Culture System for Hepatitis C Virus Genotype 6a Clinical Isolate Using a Novel Strategy and Its Sensitivity to Direct-Acting Antivirals

    doi: 10.3389/fmicb.2018.02950

    Figure Lengend Snippet: ). Cell lysates (intracellular) and culture supernatant (extracellular) were collected 48 h post transfection. HCV infectivity titers, RNA copies, and Core levels were determined and normalized to replication-independent genome, J6/JFH1-GND. (A) Intracellular and extracellular HCV infectivity titers. ∗ , no FFU was detected by manual count. Values are expressed as log 10 FFU/ml (45 μl loaded) for extracellular titers and as log 10 FFU/well (1/8 cell lysate of a well of 6-well-plate) for intracellular infectivity titers. (B) ). (C) HCV Core levels. Western blotting was performed on the cell lysates and supernatant harvested 48 h post transfection.

    Article Snippet: The HCV RNA level in the patient serum was determined by the HCV Nucleic Acid Detection Kit (TaqMan) (Daangene, China).

    Techniques: Transfection, Infection, Western Blot

    CH6a full-length clone assembled from the consensus-like PCR fragments was more efficient in virus production. Two amino acids, S2362G in NS5A and N2738D in NS5B of CH6acc genome were mutated to the consensus sequence, designated CH6acc/Cons. RNA transcripts were transfected into Huh7.5 cells, and CH6acc was included in parallel. CH6acc/Cons was delayed in virus spread and attenuated in infectivity titers. HCV-positive cells were visualized by secure anti-HCV Core immunostaining (left y -axis; lines), and HCV infectivity titers were done by FFU assay (mean of triplicate infections ± SEM, right y -axis; bars). In other transfections, CH6aFL/Cons recombinant was non-viable.

    Journal: Frontiers in Microbiology

    Article Title: Development of an Infectious Cell Culture System for Hepatitis C Virus Genotype 6a Clinical Isolate Using a Novel Strategy and Its Sensitivity to Direct-Acting Antivirals

    doi: 10.3389/fmicb.2018.02950

    Figure Lengend Snippet: CH6a full-length clone assembled from the consensus-like PCR fragments was more efficient in virus production. Two amino acids, S2362G in NS5A and N2738D in NS5B of CH6acc genome were mutated to the consensus sequence, designated CH6acc/Cons. RNA transcripts were transfected into Huh7.5 cells, and CH6acc was included in parallel. CH6acc/Cons was delayed in virus spread and attenuated in infectivity titers. HCV-positive cells were visualized by secure anti-HCV Core immunostaining (left y -axis; lines), and HCV infectivity titers were done by FFU assay (mean of triplicate infections ± SEM, right y -axis; bars). In other transfections, CH6aFL/Cons recombinant was non-viable.

    Article Snippet: The HCV RNA level in the patient serum was determined by the HCV Nucleic Acid Detection Kit (TaqMan) (Daangene, China).

    Techniques: Polymerase Chain Reaction, Sequencing, Transfection, Infection, Immunostaining, Recombinant

    GADD45α plays a vital role in regulating FFA-mediated HCV replication. Huh7 cells were transfected with GADD45α siRNA, a negative control siRNA (scramble), or GADD45α over-expressed plasmid. Forty-eight hours later, GADD45α expression was identified ( A ), and cells were then infected with HCV and simultaneously treated with FFA (0.8 mM). After 72 h, the relative HCV-NS3 protein and RNA expression were detected by Western blotting and RT-PCR ( A ); intracellular lipid accumulation was detected by Bodipy-493/503 staining and visualized after 400× amplification ( B ). Results are shown as mean ±SD of three independent experiments.

    Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

    Article Title: Hepatitis C Virus Increases Free Fatty Acids Absorption and Promotes its Replication Via Down-Regulating GADD45α Expression

    doi: 10.12659/MSM.899591

    Figure Lengend Snippet: GADD45α plays a vital role in regulating FFA-mediated HCV replication. Huh7 cells were transfected with GADD45α siRNA, a negative control siRNA (scramble), or GADD45α over-expressed plasmid. Forty-eight hours later, GADD45α expression was identified ( A ), and cells were then infected with HCV and simultaneously treated with FFA (0.8 mM). After 72 h, the relative HCV-NS3 protein and RNA expression were detected by Western blotting and RT-PCR ( A ); intracellular lipid accumulation was detected by Bodipy-493/503 staining and visualized after 400× amplification ( B ). Results are shown as mean ±SD of three independent experiments.

    Article Snippet: The HCV RNA polymerase chain reaction (PCR) commercial kit was from Daan Gene Co. Ltd., Guangzhou, China; FFA detection kits, HDL-C kit, and LDL-C kit were purchased from Sekisui Medical Technology (Japan).

    Techniques: Transfection, Negative Control, Plasmid Preparation, Expressing, Infection, RNA Expression, Western Blot, Reverse Transcription Polymerase Chain Reaction, Staining, Amplification

    FFA promotes HCV replication in Huh7 cells. Huh7 cells (2×10 5 ) were cultured in a 24-well plate to 80%–85% confluence and then co-cultured with different concentrations of FFA with or without HCV (MOI of 1.5) for 72 hours. ( A ) HCV RNA copies were quantified by RT-PCR, and NS3 protein level was detected by Western blotting. ( B ) Quantification of the GADD45α protein levels and mRNA levels in Huh7 cells. Results are shown as mean ± SD of three independent experiments.

    Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

    Article Title: Hepatitis C Virus Increases Free Fatty Acids Absorption and Promotes its Replication Via Down-Regulating GADD45α Expression

    doi: 10.12659/MSM.899591

    Figure Lengend Snippet: FFA promotes HCV replication in Huh7 cells. Huh7 cells (2×10 5 ) were cultured in a 24-well plate to 80%–85% confluence and then co-cultured with different concentrations of FFA with or without HCV (MOI of 1.5) for 72 hours. ( A ) HCV RNA copies were quantified by RT-PCR, and NS3 protein level was detected by Western blotting. ( B ) Quantification of the GADD45α protein levels and mRNA levels in Huh7 cells. Results are shown as mean ± SD of three independent experiments.

    Article Snippet: The HCV RNA polymerase chain reaction (PCR) commercial kit was from Daan Gene Co. Ltd., Guangzhou, China; FFA detection kits, HDL-C kit, and LDL-C kit were purchased from Sekisui Medical Technology (Japan).

    Techniques: Cell Culture, Reverse Transcription Polymerase Chain Reaction, Western Blot

    The role of FFA and HCV in GADD45α expression. ( A ) GADD45α protein expression and mRNA level were detected in tumor tissues from liver cancer patients coinciding with chronic HCV infection. Low, medium, and high mean different serum viral loads. ( B ) CCK-8 assay showing the effect of different concentrations of FFA (up to 2 mM) on cell viability of Huh7 cells. Cell viability was expressed as percentage of untreated cells. Huh7 cells (2×10 5 ) were cultured in a 24-well plate to 80%–85% confluence and then treated with different concentrations of FFA ( C ) or infected with HCV ( D ) for 72 hours. mRNA level of GADD45α was quantified by RT-PCR, and protein expression was detected by Western blotting. Results are shown as mean ±SD of three independent experiments.

    Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

    Article Title: Hepatitis C Virus Increases Free Fatty Acids Absorption and Promotes its Replication Via Down-Regulating GADD45α Expression

    doi: 10.12659/MSM.899591

    Figure Lengend Snippet: The role of FFA and HCV in GADD45α expression. ( A ) GADD45α protein expression and mRNA level were detected in tumor tissues from liver cancer patients coinciding with chronic HCV infection. Low, medium, and high mean different serum viral loads. ( B ) CCK-8 assay showing the effect of different concentrations of FFA (up to 2 mM) on cell viability of Huh7 cells. Cell viability was expressed as percentage of untreated cells. Huh7 cells (2×10 5 ) were cultured in a 24-well plate to 80%–85% confluence and then treated with different concentrations of FFA ( C ) or infected with HCV ( D ) for 72 hours. mRNA level of GADD45α was quantified by RT-PCR, and protein expression was detected by Western blotting. Results are shown as mean ±SD of three independent experiments.

    Article Snippet: The HCV RNA polymerase chain reaction (PCR) commercial kit was from Daan Gene Co. Ltd., Guangzhou, China; FFA detection kits, HDL-C kit, and LDL-C kit were purchased from Sekisui Medical Technology (Japan).

    Techniques: Expressing, Infection, CCK-8 Assay, Cell Culture, Reverse Transcription Polymerase Chain Reaction, Western Blot