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    miRCURY LNA miRNA PCR Assay
    Description:
    miRCURY LNA miRNA PCR Assays are individual miRNA PCR primer sets that enable extremely sensitive and specific miRNA quantification with themiRCURY LNA miRNA PCR System Both forward and reverse PCR amplification primers are miRNA specific and are optimized with LNA technology Assays are available in tube or plate format with 200 reactions per tube or well
    Catalog Number:
    339306
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    Category:
    qPCR Arrays and Assays
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    Structured Review

    Qiagen total rna
    miRCURY LNA miRNA PCR Assay
    miRCURY LNA miRNA PCR Assays are individual miRNA PCR primer sets that enable extremely sensitive and specific miRNA quantification with themiRCURY LNA miRNA PCR System Both forward and reverse PCR amplification primers are miRNA specific and are optimized with LNA technology Assays are available in tube or plate format with 200 reactions per tube or well
    https://www.bioz.com/result/total rna/product/Qiagen
    Average 99 stars, based on 21096 article reviews
    Price from $9.99 to $1999.99
    total rna - by Bioz Stars, 2020-04
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    Images

    1) Product Images from "Interplay between the Epigenetic Enzyme Lysine (K)-Specific Demethylase 2B and Epstein-Barr Virus Infection"

    Article Title: Interplay between the Epigenetic Enzyme Lysine (K)-Specific Demethylase 2B and Epstein-Barr Virus Infection

    Journal: Journal of Virology

    doi: 10.1128/JVI.00273-19

    EBV also uses LMP1-independent mechanisms to downregulate KDM2B. (A) B cells from two donors were infected with EBV or EBVΔLMP1 or mock infected (MI) and collected at 48 h after infection. Reverse-transcribed RNA samples were analyzed by qPCR for the KDM2B expression level. (B) B cells from two donors were infected with EBV and collected at 12, 24, and 48 h after infection. Cells were processed for RNA extraction and analyzed by qPCR for the expression levels of EBNA1, EBNA2, EBER, LMP1, and KDM2B transcripts. The expression levels of viral gene transcripts were measured relative to their levels in B cells collected 12 h (EBNA2) or 48 h (EBNA1, EBER, LMP1) postinfection. The levels of KDM2B mRNA were measured relative to its levels in mock-infected cells. (C) Different EBV(+) cell lines in latency phase I (BL110, I100, MUTU) or in latency phase III (LCL, Raji, B95) and primary B cells (BC) were collected, processed for RNA extraction and reverse transcription, and analyzed by qPCR for LMP1 (top), EBNA1 (middle), and KDM2B (bottom) mRNA levels. (D and E) RPMI (D) and Louckes (E) cells were transiently transfected with different constructs carrying individual EBV genes (TC) in three independent experiments. At 48 h after transfection, cells were processed for RNA and protein extraction and analyzed for the KDM2B expression level by RT-qPCR analysis. The levels of KDM2B mRNA were measured relative to its levels in mock-infected cells. Western blotting (LMP1, EBNA1, EBNA3A/3B, and EBNALP) or RT-PCR analysis (LMP2A, EBNA2, and EBNA3C) was performed to measure the expression of the different viral latent proteins. (F) RPMI cells were transfected with different EBV gene-carrying constructs. At 44 h after transfection, cells were exposed to the proteasome inhibitor MG132 for 4 h and then collected, processed for total protein extraction, and analyzed for the indicated proteins by immunoblotting. The histogram shows the average phosphorylated IκBα (P-IκBα) signal normalized to the levels of total IκBα. (G) B cells from different donors were untreated or treated with a miRNA-146a-5p inhibitor for 24 h before infection with EBV. At 48 h after infection, cells were collected and processed for RNA extraction and reverse transcription. cDNA samples were analyzed by qPCR for LMP1 and KDM2B expression levels. The levels of KDM2B mRNA were measured relative to its levels in mock-infected cells. The histograms show the average expression levels measured in 2 independent experiments (*, P  
    Figure Legend Snippet: EBV also uses LMP1-independent mechanisms to downregulate KDM2B. (A) B cells from two donors were infected with EBV or EBVΔLMP1 or mock infected (MI) and collected at 48 h after infection. Reverse-transcribed RNA samples were analyzed by qPCR for the KDM2B expression level. (B) B cells from two donors were infected with EBV and collected at 12, 24, and 48 h after infection. Cells were processed for RNA extraction and analyzed by qPCR for the expression levels of EBNA1, EBNA2, EBER, LMP1, and KDM2B transcripts. The expression levels of viral gene transcripts were measured relative to their levels in B cells collected 12 h (EBNA2) or 48 h (EBNA1, EBER, LMP1) postinfection. The levels of KDM2B mRNA were measured relative to its levels in mock-infected cells. (C) Different EBV(+) cell lines in latency phase I (BL110, I100, MUTU) or in latency phase III (LCL, Raji, B95) and primary B cells (BC) were collected, processed for RNA extraction and reverse transcription, and analyzed by qPCR for LMP1 (top), EBNA1 (middle), and KDM2B (bottom) mRNA levels. (D and E) RPMI (D) and Louckes (E) cells were transiently transfected with different constructs carrying individual EBV genes (TC) in three independent experiments. At 48 h after transfection, cells were processed for RNA and protein extraction and analyzed for the KDM2B expression level by RT-qPCR analysis. The levels of KDM2B mRNA were measured relative to its levels in mock-infected cells. Western blotting (LMP1, EBNA1, EBNA3A/3B, and EBNALP) or RT-PCR analysis (LMP2A, EBNA2, and EBNA3C) was performed to measure the expression of the different viral latent proteins. (F) RPMI cells were transfected with different EBV gene-carrying constructs. At 44 h after transfection, cells were exposed to the proteasome inhibitor MG132 for 4 h and then collected, processed for total protein extraction, and analyzed for the indicated proteins by immunoblotting. The histogram shows the average phosphorylated IκBα (P-IκBα) signal normalized to the levels of total IκBα. (G) B cells from different donors were untreated or treated with a miRNA-146a-5p inhibitor for 24 h before infection with EBV. At 48 h after infection, cells were collected and processed for RNA extraction and reverse transcription. cDNA samples were analyzed by qPCR for LMP1 and KDM2B expression levels. The levels of KDM2B mRNA were measured relative to its levels in mock-infected cells. The histograms show the average expression levels measured in 2 independent experiments (*, P  

    Techniques Used: Infection, Real-time Polymerase Chain Reaction, Expressing, RNA Extraction, Transfection, Construct, Protein Extraction, Quantitative RT-PCR, Western Blot, Reverse Transcription Polymerase Chain Reaction

    KDM2B regulates EBV gene expression in latently infected cells. (A to C) LCL were transfected with 1.5 μg of pCDNA3-KDM2B or with pCDNA as a control in three independent experiments. Cells were collected at 24 h after transfection and processed for RNA/DNA and total protein extraction. (A) KDM2B mRNA and protein levels were shown by qPCR (bottom) and immunoblotting (top; the lines corresponding to conditions with 0.5 μg and 1 μg of KDM2B, originally present in the Western blot, are not shown because they were excluded from all the analyses). (B) DNA samples were analyzed by TaqMan PCR to assess the number of EBV genome copies per cell (ns, not significant). (C) The mRNA expression levels of EBV latent and early genes were assessed by qPCR (*, P  
    Figure Legend Snippet: KDM2B regulates EBV gene expression in latently infected cells. (A to C) LCL were transfected with 1.5 μg of pCDNA3-KDM2B or with pCDNA as a control in three independent experiments. Cells were collected at 24 h after transfection and processed for RNA/DNA and total protein extraction. (A) KDM2B mRNA and protein levels were shown by qPCR (bottom) and immunoblotting (top; the lines corresponding to conditions with 0.5 μg and 1 μg of KDM2B, originally present in the Western blot, are not shown because they were excluded from all the analyses). (B) DNA samples were analyzed by TaqMan PCR to assess the number of EBV genome copies per cell (ns, not significant). (C) The mRNA expression levels of EBV latent and early genes were assessed by qPCR (*, P  

    Techniques Used: Expressing, Infection, Transfection, Protein Extraction, Real-time Polymerase Chain Reaction, Western Blot, Polymerase Chain Reaction

    KDM2B deregulation alters EBV gene expression. (A) Louckes cells were transiently transfected with increasing concentrations of the KDM2B expression vector in three independent experiments, collected at 24 h after transfection, and processed for protein and RNA extraction to assess KDM2B levels by immunoblotting (top) or qPCR (bottom). (B) Cells were then infected with EBV, and at 24 h after infection they were collected and processed for FACS analysis and for RNA/DNA extraction. DNA samples were used to measure EBV genome copy number by TaqMan PCR (ns, not significant). (C) Live cells were analyzed by FACS to measure the GFP mean fluorescence intensity (MFI). (D) cDNA samples were analyzed for the expression level of EBV early and late genes by qPCR. The levels of EBV genes transcripts were normalized to the mRNA levels of the housekeeping gene β-globin and calculated relative to their levels in cells transfected with the empty vector (KDM2B, 0 μg). The values shown in the histogram are the average from 3 independent experiments (*, P  
    Figure Legend Snippet: KDM2B deregulation alters EBV gene expression. (A) Louckes cells were transiently transfected with increasing concentrations of the KDM2B expression vector in three independent experiments, collected at 24 h after transfection, and processed for protein and RNA extraction to assess KDM2B levels by immunoblotting (top) or qPCR (bottom). (B) Cells were then infected with EBV, and at 24 h after infection they were collected and processed for FACS analysis and for RNA/DNA extraction. DNA samples were used to measure EBV genome copy number by TaqMan PCR (ns, not significant). (C) Live cells were analyzed by FACS to measure the GFP mean fluorescence intensity (MFI). (D) cDNA samples were analyzed for the expression level of EBV early and late genes by qPCR. The levels of EBV genes transcripts were normalized to the mRNA levels of the housekeeping gene β-globin and calculated relative to their levels in cells transfected with the empty vector (KDM2B, 0 μg). The values shown in the histogram are the average from 3 independent experiments (*, P  

    Techniques Used: Expressing, Transfection, Plasmid Preparation, RNA Extraction, Real-time Polymerase Chain Reaction, Infection, FACS, DNA Extraction, Polymerase Chain Reaction, Fluorescence

    2) Product Images from "Evaluation of extraction kits and RT-qPCR systems adapted to high-throughput platform for circulating miRNAs"

    Article Title: Evaluation of extraction kits and RT-qPCR systems adapted to high-throughput platform for circulating miRNAs

    Journal: Scientific Reports

    doi: 10.1038/srep09430

    miRNA recovery of different extraction kits were evaluated by the C q values for spike-in controls (cel-miR-39 and cel-miR-54) assayed in the (a) TaqMan and (b) miScript RT-qPCR system. Box plots with whiskers show 5–95 percentile of the C q values for spike-in controls and the corresponding CVs are shown. The extent of extraction bias across samples was reflected in the range and CV of C q values for spike-in controls. Higher median C q compared to other extraction kits indicate poorer performance in miRNA recovery. Q, miRNeasy Serum/Plasma kit; E, miRCURY™ RNA Isolation Kit - Biofluids; A, mirVana™ PARIS™ Kit; MN, NucleoSpin® miRNA Plasma; NB, Plasma/Serum Circulating RNA Purification Kit.
    Figure Legend Snippet: miRNA recovery of different extraction kits were evaluated by the C q values for spike-in controls (cel-miR-39 and cel-miR-54) assayed in the (a) TaqMan and (b) miScript RT-qPCR system. Box plots with whiskers show 5–95 percentile of the C q values for spike-in controls and the corresponding CVs are shown. The extent of extraction bias across samples was reflected in the range and CV of C q values for spike-in controls. Higher median C q compared to other extraction kits indicate poorer performance in miRNA recovery. Q, miRNeasy Serum/Plasma kit; E, miRCURY™ RNA Isolation Kit - Biofluids; A, mirVana™ PARIS™ Kit; MN, NucleoSpin® miRNA Plasma; NB, Plasma/Serum Circulating RNA Purification Kit.

    Techniques Used: Quantitative RT-PCR, Isolation, Purification

    Comparison of TaqMan and miScript RT-qPCR systems adapted to the high-throughput platform by performance parameters which include reproducibility, accuracy and sensitivity. Transformation of data to z-score was carried out to facilitate direct comparison. A1, accuracy when measuring fold change of abundant miRNAs; A2, accuracy when measuring fold change of less abundant miRNAs, R1, reproducibility of RT replicates; R2, reproducibility of RT replicates after normalisation; R3, reproducibility of qPCR replicates; S1, sensitivity to detect the presence of plasma miRNAs and S2, sensitivity to detect the presence of miRNAs in titration points.
    Figure Legend Snippet: Comparison of TaqMan and miScript RT-qPCR systems adapted to the high-throughput platform by performance parameters which include reproducibility, accuracy and sensitivity. Transformation of data to z-score was carried out to facilitate direct comparison. A1, accuracy when measuring fold change of abundant miRNAs; A2, accuracy when measuring fold change of less abundant miRNAs, R1, reproducibility of RT replicates; R2, reproducibility of RT replicates after normalisation; R3, reproducibility of qPCR replicates; S1, sensitivity to detect the presence of plasma miRNAs and S2, sensitivity to detect the presence of miRNAs in titration points.

    Techniques Used: Quantitative RT-PCR, High Throughput Screening Assay, Transformation Assay, Real-time Polymerase Chain Reaction, Titration

    Evaluation of consistency between technical replicatesin RT-qPCR. Consistencies among (a) RT replicates in TaqMan system, (b) RT replicates in miScript system, (c) qPCR replicates in TaqMan system and (d) qPCR replicates in miScript system were evaluated by measuring the levels of 11–16 miRNAs in a same set of RNA samples (n = 5–14). Each data point in scatter plot represents the average C q value obtained from the duplicate or triplicate wells of qPCR. Overall, significant correlations (ICC = 0.712–0.996, p
    Figure Legend Snippet: Evaluation of consistency between technical replicatesin RT-qPCR. Consistencies among (a) RT replicates in TaqMan system, (b) RT replicates in miScript system, (c) qPCR replicates in TaqMan system and (d) qPCR replicates in miScript system were evaluated by measuring the levels of 11–16 miRNAs in a same set of RNA samples (n = 5–14). Each data point in scatter plot represents the average C q value obtained from the duplicate or triplicate wells of qPCR. Overall, significant correlations (ICC = 0.712–0.996, p

    Techniques Used: Quantitative RT-PCR, Real-time Polymerase Chain Reaction, Immunocytochemistry

    Comparison of different RT-qPCR systems adapted to the BioMark high-throughput platform. (a) Average C q values for endogeneous plasma miRNAs were depicted in box plot with whiskers showing the 5–95 percentile. Irrespective of extraction kits being used, within the same set of samples, more undetermined C q values were obtained in the miScript system. (b) Correlation of qPCR results from TaqMan and miScript systems may vary among different miRNA assays. Each data point represents the normalised C q value transformed into z-score. Q, miRNeasy Serum/Plasma kit; E, miRCURY™ RNA Isolation Kit - Biofluids; A, mirVana™ PARIS™ Kit; MN, NucleoSpin® miRNA Plasma; NB, Plasma/Serum Circulating RNA Purification Kit.
    Figure Legend Snippet: Comparison of different RT-qPCR systems adapted to the BioMark high-throughput platform. (a) Average C q values for endogeneous plasma miRNAs were depicted in box plot with whiskers showing the 5–95 percentile. Irrespective of extraction kits being used, within the same set of samples, more undetermined C q values were obtained in the miScript system. (b) Correlation of qPCR results from TaqMan and miScript systems may vary among different miRNA assays. Each data point represents the normalised C q value transformed into z-score. Q, miRNeasy Serum/Plasma kit; E, miRCURY™ RNA Isolation Kit - Biofluids; A, mirVana™ PARIS™ Kit; MN, NucleoSpin® miRNA Plasma; NB, Plasma/Serum Circulating RNA Purification Kit.

    Techniques Used: Quantitative RT-PCR, High Throughput Screening Assay, Real-time Polymerase Chain Reaction, Transformation Assay, Isolation, Purification

    3) Product Images from "Stemness, Pluripotentiality, and Wnt Antagonism: sFRP4, a Wnt antagonist Mediates Pluripotency and Stemness in Glioblastoma"

    Article Title: Stemness, Pluripotentiality, and Wnt Antagonism: sFRP4, a Wnt antagonist Mediates Pluripotency and Stemness in Glioblastoma

    Journal: Cancers

    doi: 10.3390/cancers11010025

    ( A – F ). Apoptosis related genes identified in sFRP4 chromatin immunoprecipitation (ChIP) and pull-down sequencing analysis. Schematic 1 represents the sequence of steps of ChIP and downstream sequencing. ChIP DNA resolved on agarose gel indicated a 150 bp DNA band ( A ), ChIP mapping statistics by Burrow-Wheeler Aligner software indicated 5,581,398 mapped reads ( B ); peak calling analysis output using MACS2 software revealed 34,711 peaks related to secreted frizzled-related protein 4 ( sFRP4 ) ( C , left panel), categorization of peak identities represented by pie chart and table, analysis gene list present within 5′UTR (enlarged) indicated the presence of miRNA 885 ( C , right panel), RNA sequencing data of 5′UTR revealed an upregulation of three 5′UTR genes in sFRP4 OE (OE) and downregulation in sFRP4 SI (SI) cells as indicated in the box ( D ), upregulation of active miR-885 in sFRP4 OE cells using an miR-885 5′LNA probe was detected as red fluorescence using a fluorescence microscope (scale bar = 10 μm) ( E ), quantitative RT-PCR indicated an over expression of miR885 in sFRP4 OE and downregulation in sFRP4 SI cells ( F ). Schematic 2 represents a model indicating the mode of action of miR-885 through its target genes CDK2 and MCM5 in cellular homeostasis via activation of p53 . Results are mean ± SD of three independent experiments performed in triplicates (* p value
    Figure Legend Snippet: ( A – F ). Apoptosis related genes identified in sFRP4 chromatin immunoprecipitation (ChIP) and pull-down sequencing analysis. Schematic 1 represents the sequence of steps of ChIP and downstream sequencing. ChIP DNA resolved on agarose gel indicated a 150 bp DNA band ( A ), ChIP mapping statistics by Burrow-Wheeler Aligner software indicated 5,581,398 mapped reads ( B ); peak calling analysis output using MACS2 software revealed 34,711 peaks related to secreted frizzled-related protein 4 ( sFRP4 ) ( C , left panel), categorization of peak identities represented by pie chart and table, analysis gene list present within 5′UTR (enlarged) indicated the presence of miRNA 885 ( C , right panel), RNA sequencing data of 5′UTR revealed an upregulation of three 5′UTR genes in sFRP4 OE (OE) and downregulation in sFRP4 SI (SI) cells as indicated in the box ( D ), upregulation of active miR-885 in sFRP4 OE cells using an miR-885 5′LNA probe was detected as red fluorescence using a fluorescence microscope (scale bar = 10 μm) ( E ), quantitative RT-PCR indicated an over expression of miR885 in sFRP4 OE and downregulation in sFRP4 SI cells ( F ). Schematic 2 represents a model indicating the mode of action of miR-885 through its target genes CDK2 and MCM5 in cellular homeostasis via activation of p53 . Results are mean ± SD of three independent experiments performed in triplicates (* p value

    Techniques Used: Chromatin Immunoprecipitation, Sequencing, Agarose Gel Electrophoresis, Software, RNA Sequencing Assay, Fluorescence, Microscopy, Quantitative RT-PCR, Over Expression, Activation Assay

    4) Product Images from "The miR-30 miRNA family regulates Xenopus pronephros development and targets the transcription factor Xlim1/Lhx1"

    Article Title: The miR-30 miRNA family regulates Xenopus pronephros development and targets the transcription factor Xlim1/Lhx1

    Journal:

    doi: 10.1242/dev.037432

    Kidney expression of miRNAs. ( A ) In situ hybridizations of mouse kidney sections at E14.5 and P0 using LNA-modified oligonucleotide probes. miRNA families were studied using degenerate probes that recognize all family members. ( B ) Whole-mount in situ
    Figure Legend Snippet: Kidney expression of miRNAs. ( A ) In situ hybridizations of mouse kidney sections at E14.5 and P0 using LNA-modified oligonucleotide probes. miRNA families were studied using degenerate probes that recognize all family members. ( B ) Whole-mount in situ

    Techniques Used: Expressing, In Situ, Modification

    5) Product Images from "Interplay between the Epigenetic Enzyme Lysine (K)-Specific Demethylase 2B and Epstein-Barr Virus Infection"

    Article Title: Interplay between the Epigenetic Enzyme Lysine (K)-Specific Demethylase 2B and Epstein-Barr Virus Infection

    Journal: Journal of Virology

    doi: 10.1128/JVI.00273-19

    EBV also uses LMP1-independent mechanisms to downregulate KDM2B. (A) B cells from two donors were infected with EBV or EBVΔLMP1 or mock infected (MI) and collected at 48 h after infection. Reverse-transcribed RNA samples were analyzed by qPCR for the KDM2B expression level. (B) B cells from two donors were infected with EBV and collected at 12, 24, and 48 h after infection. Cells were processed for RNA extraction and analyzed by qPCR for the expression levels of EBNA1, EBNA2, EBER, LMP1, and KDM2B transcripts. The expression levels of viral gene transcripts were measured relative to their levels in B cells collected 12 h (EBNA2) or 48 h (EBNA1, EBER, LMP1) postinfection. The levels of KDM2B mRNA were measured relative to its levels in mock-infected cells. (C) Different EBV(+) cell lines in latency phase I (BL110, I100, MUTU) or in latency phase III (LCL, Raji, B95) and primary B cells (BC) were collected, processed for RNA extraction and reverse transcription, and analyzed by qPCR for LMP1 (top), EBNA1 (middle), and KDM2B (bottom) mRNA levels. (D and E) RPMI (D) and Louckes (E) cells were transiently transfected with different constructs carrying individual EBV genes (TC) in three independent experiments. At 48 h after transfection, cells were processed for RNA and protein extraction and analyzed for the KDM2B expression level by RT-qPCR analysis. The levels of KDM2B mRNA were measured relative to its levels in mock-infected cells. Western blotting (LMP1, EBNA1, EBNA3A/3B, and EBNALP) or RT-PCR analysis (LMP2A, EBNA2, and EBNA3C) was performed to measure the expression of the different viral latent proteins. (F) RPMI cells were transfected with different EBV gene-carrying constructs. At 44 h after transfection, cells were exposed to the proteasome inhibitor MG132 for 4 h and then collected, processed for total protein extraction, and analyzed for the indicated proteins by immunoblotting. The histogram shows the average phosphorylated IκBα (P-IκBα) signal normalized to the levels of total IκBα. (G) B cells from different donors were untreated or treated with a miRNA-146a-5p inhibitor for 24 h before infection with EBV. At 48 h after infection, cells were collected and processed for RNA extraction and reverse transcription. cDNA samples were analyzed by qPCR for LMP1 and KDM2B expression levels. The levels of KDM2B mRNA were measured relative to its levels in mock-infected cells. The histograms show the average expression levels measured in 2 independent experiments (*, P
    Figure Legend Snippet: EBV also uses LMP1-independent mechanisms to downregulate KDM2B. (A) B cells from two donors were infected with EBV or EBVΔLMP1 or mock infected (MI) and collected at 48 h after infection. Reverse-transcribed RNA samples were analyzed by qPCR for the KDM2B expression level. (B) B cells from two donors were infected with EBV and collected at 12, 24, and 48 h after infection. Cells were processed for RNA extraction and analyzed by qPCR for the expression levels of EBNA1, EBNA2, EBER, LMP1, and KDM2B transcripts. The expression levels of viral gene transcripts were measured relative to their levels in B cells collected 12 h (EBNA2) or 48 h (EBNA1, EBER, LMP1) postinfection. The levels of KDM2B mRNA were measured relative to its levels in mock-infected cells. (C) Different EBV(+) cell lines in latency phase I (BL110, I100, MUTU) or in latency phase III (LCL, Raji, B95) and primary B cells (BC) were collected, processed for RNA extraction and reverse transcription, and analyzed by qPCR for LMP1 (top), EBNA1 (middle), and KDM2B (bottom) mRNA levels. (D and E) RPMI (D) and Louckes (E) cells were transiently transfected with different constructs carrying individual EBV genes (TC) in three independent experiments. At 48 h after transfection, cells were processed for RNA and protein extraction and analyzed for the KDM2B expression level by RT-qPCR analysis. The levels of KDM2B mRNA were measured relative to its levels in mock-infected cells. Western blotting (LMP1, EBNA1, EBNA3A/3B, and EBNALP) or RT-PCR analysis (LMP2A, EBNA2, and EBNA3C) was performed to measure the expression of the different viral latent proteins. (F) RPMI cells were transfected with different EBV gene-carrying constructs. At 44 h after transfection, cells were exposed to the proteasome inhibitor MG132 for 4 h and then collected, processed for total protein extraction, and analyzed for the indicated proteins by immunoblotting. The histogram shows the average phosphorylated IκBα (P-IκBα) signal normalized to the levels of total IκBα. (G) B cells from different donors were untreated or treated with a miRNA-146a-5p inhibitor for 24 h before infection with EBV. At 48 h after infection, cells were collected and processed for RNA extraction and reverse transcription. cDNA samples were analyzed by qPCR for LMP1 and KDM2B expression levels. The levels of KDM2B mRNA were measured relative to its levels in mock-infected cells. The histograms show the average expression levels measured in 2 independent experiments (*, P

    Techniques Used: Infection, Real-time Polymerase Chain Reaction, Expressing, RNA Extraction, Transfection, Construct, Protein Extraction, Quantitative RT-PCR, Western Blot, Reverse Transcription Polymerase Chain Reaction

    KDM2B regulates EBV gene expression in latently infected cells. (A to C) LCL were transfected with 1.5 μg of pCDNA3-KDM2B or with pCDNA as a control in three independent experiments. Cells were collected at 24 h after transfection and processed for RNA/DNA and total protein extraction. (A) KDM2B mRNA and protein levels were shown by qPCR (bottom) and immunoblotting (top; the lines corresponding to conditions with 0.5 μg and 1 μg of KDM2B, originally present in the Western blot, are not shown because they were excluded from all the analyses). (B) DNA samples were analyzed by TaqMan PCR to assess the number of EBV genome copies per cell (ns, not significant). (C) The mRNA expression levels of EBV latent and early genes were assessed by qPCR (*, P
    Figure Legend Snippet: KDM2B regulates EBV gene expression in latently infected cells. (A to C) LCL were transfected with 1.5 μg of pCDNA3-KDM2B or with pCDNA as a control in three independent experiments. Cells were collected at 24 h after transfection and processed for RNA/DNA and total protein extraction. (A) KDM2B mRNA and protein levels were shown by qPCR (bottom) and immunoblotting (top; the lines corresponding to conditions with 0.5 μg and 1 μg of KDM2B, originally present in the Western blot, are not shown because they were excluded from all the analyses). (B) DNA samples were analyzed by TaqMan PCR to assess the number of EBV genome copies per cell (ns, not significant). (C) The mRNA expression levels of EBV latent and early genes were assessed by qPCR (*, P

    Techniques Used: Expressing, Infection, Transfection, Protein Extraction, Real-time Polymerase Chain Reaction, Western Blot, Polymerase Chain Reaction

    KDM2B deregulation alters EBV gene expression. (A) Louckes cells were transiently transfected with increasing concentrations of the KDM2B expression vector in three independent experiments, collected at 24 h after transfection, and processed for protein and RNA extraction to assess KDM2B levels by immunoblotting (top) or qPCR (bottom). (B) Cells were then infected with EBV, and at 24 h after infection they were collected and processed for FACS analysis and for RNA/DNA extraction. DNA samples were used to measure EBV genome copy number by TaqMan PCR (ns, not significant). (C) Live cells were analyzed by FACS to measure the GFP mean fluorescence intensity (MFI). (D) cDNA samples were analyzed for the expression level of EBV early and late genes by qPCR. The levels of EBV genes transcripts were normalized to the mRNA levels of the housekeeping gene β-globin and calculated relative to their levels in cells transfected with the empty vector (KDM2B, 0 μg). The values shown in the histogram are the average from 3 independent experiments (*, P
    Figure Legend Snippet: KDM2B deregulation alters EBV gene expression. (A) Louckes cells were transiently transfected with increasing concentrations of the KDM2B expression vector in three independent experiments, collected at 24 h after transfection, and processed for protein and RNA extraction to assess KDM2B levels by immunoblotting (top) or qPCR (bottom). (B) Cells were then infected with EBV, and at 24 h after infection they were collected and processed for FACS analysis and for RNA/DNA extraction. DNA samples were used to measure EBV genome copy number by TaqMan PCR (ns, not significant). (C) Live cells were analyzed by FACS to measure the GFP mean fluorescence intensity (MFI). (D) cDNA samples were analyzed for the expression level of EBV early and late genes by qPCR. The levels of EBV genes transcripts were normalized to the mRNA levels of the housekeeping gene β-globin and calculated relative to their levels in cells transfected with the empty vector (KDM2B, 0 μg). The values shown in the histogram are the average from 3 independent experiments (*, P

    Techniques Used: Expressing, Transfection, Plasmid Preparation, RNA Extraction, Real-time Polymerase Chain Reaction, Infection, FACS, DNA Extraction, Polymerase Chain Reaction, Fluorescence

    6) Product Images from "Microarray and deep sequencing cross-platform analysis of the mirRNome and isomiR variation in response to epidermal growth factor"

    Article Title: Microarray and deep sequencing cross-platform analysis of the mirRNome and isomiR variation in response to epidermal growth factor

    Journal: BMC Genomics

    doi: 10.1186/1471-2164-14-371

    miRNA transcriptome profiles of EGF-treated HeLa cells using Exiqon and Agilent miRNA arrays. A . Shared miRNA genes between Exiqon and Agilent microRNA arrays. Venn diagram showing the unique and named miRNAs genes shared between Agilent and Exiqon microarray platforms. The pool of 346 shared genes was used for all subsequent cross-platform analyses. Numbers outside diagrams indicate the total amount of miRNA genes contained in each platform. In brackets, percentage of miRNA genes contained in each platform over the total miRNA genes contained in all platforms. B . Regulated miRNAs after 6 hours EGF treatment using Exiqon and Agilent miRNA arrays. HeLa cells were serum-starved for 24 h and treated with EGF for 6 h. Total RNAs prepared from cells lysates were hybridized to Exiqon miRCURY LNA microRNA Array V9.2 and Agilent Human miRNA V2 Oligo Microarray. List shows regulated miRNAs identified by SAM analysis (FDR = 0.05) for miRNAs with a minimal expression change of 1.2 fold.
    Figure Legend Snippet: miRNA transcriptome profiles of EGF-treated HeLa cells using Exiqon and Agilent miRNA arrays. A . Shared miRNA genes between Exiqon and Agilent microRNA arrays. Venn diagram showing the unique and named miRNAs genes shared between Agilent and Exiqon microarray platforms. The pool of 346 shared genes was used for all subsequent cross-platform analyses. Numbers outside diagrams indicate the total amount of miRNA genes contained in each platform. In brackets, percentage of miRNA genes contained in each platform over the total miRNA genes contained in all platforms. B . Regulated miRNAs after 6 hours EGF treatment using Exiqon and Agilent miRNA arrays. HeLa cells were serum-starved for 24 h and treated with EGF for 6 h. Total RNAs prepared from cells lysates were hybridized to Exiqon miRCURY LNA microRNA Array V9.2 and Agilent Human miRNA V2 Oligo Microarray. List shows regulated miRNAs identified by SAM analysis (FDR = 0.05) for miRNAs with a minimal expression change of 1.2 fold.

    Techniques Used: Microarray, Expressing

    RT-qPCR validation of EGF-regulated miRNAs. Total RNA prepared from cells lysates were analyzed by quantitative real time PCR using the miRCURY LNA™ microRNA PCR System (Exiqon) for each of the miRNAs as indicated. A . Control treatment without inhibitors: HeLa cells were serum-starved for 24 hours and treated with EGF for 1 and 6 hours. B . Inhibitor treatment. HeLa cells were serum-starved for 24 hours and treated with EGF for 6 hours in the presence or absence of protein kinase inhibitors: AG1470 (EGFR inhibitor), U0126 (MEK inhibitor) and Wortmannin (PI3K inhibitor). In addition, HeLa cells were transfected with a constitutively active form of Ras (RasV12). Effective pathway inhibition was verified by western blotting in parallel samples from the same experiment (see Additional file 8 ).
    Figure Legend Snippet: RT-qPCR validation of EGF-regulated miRNAs. Total RNA prepared from cells lysates were analyzed by quantitative real time PCR using the miRCURY LNA™ microRNA PCR System (Exiqon) for each of the miRNAs as indicated. A . Control treatment without inhibitors: HeLa cells were serum-starved for 24 hours and treated with EGF for 1 and 6 hours. B . Inhibitor treatment. HeLa cells were serum-starved for 24 hours and treated with EGF for 6 hours in the presence or absence of protein kinase inhibitors: AG1470 (EGFR inhibitor), U0126 (MEK inhibitor) and Wortmannin (PI3K inhibitor). In addition, HeLa cells were transfected with a constitutively active form of Ras (RasV12). Effective pathway inhibition was verified by western blotting in parallel samples from the same experiment (see Additional file 8 ).

    Techniques Used: Quantitative RT-PCR, Real-time Polymerase Chain Reaction, Polymerase Chain Reaction, Transfection, Inhibition, Western Blot

    7) Product Images from "MicroRNA-215 Regulates Fibroblast Function: Insights from a Human Fibrotic Disease"

    Article Title: MicroRNA-215 Regulates Fibroblast Function: Insights from a Human Fibrotic Disease

    Journal: Cell Cycle

    doi: 10.1080/15384101.2014.998077

    ( A ) Bar chart showing results of the Exiqon microRNA Array. Height of the bars shows the mean normalized log values of miR-215 levels in conjunctiva and pterygium tissues from different patients. ( B ) Bar chart showing individual and overall results from qRT-PCR assay performed on 3 separate pairs of human pterygium and conjunctival tissues. Height of the bars represents the relative fold change of miR-215 levels in pterygium with respect to conjunctival control. Error bars represent standard error of mean. Values are normalized to the 5S rRNA control values. ( C ) miR-215 in situ staining of human pterygium and conjunctival tissues. Diagram on the right showing conjunctival epithelium and some stroma. Pterygium and conjunctival sections hybridized with DIG-labeled miR-215 LNA probes (red). DNA was counterstained with DAPI (blue). Scale bar represents 100 μm. Dashed lines represent position of basement membrane separating epithelium from stroma.
    Figure Legend Snippet: ( A ) Bar chart showing results of the Exiqon microRNA Array. Height of the bars shows the mean normalized log values of miR-215 levels in conjunctiva and pterygium tissues from different patients. ( B ) Bar chart showing individual and overall results from qRT-PCR assay performed on 3 separate pairs of human pterygium and conjunctival tissues. Height of the bars represents the relative fold change of miR-215 levels in pterygium with respect to conjunctival control. Error bars represent standard error of mean. Values are normalized to the 5S rRNA control values. ( C ) miR-215 in situ staining of human pterygium and conjunctival tissues. Diagram on the right showing conjunctival epithelium and some stroma. Pterygium and conjunctival sections hybridized with DIG-labeled miR-215 LNA probes (red). DNA was counterstained with DAPI (blue). Scale bar represents 100 μm. Dashed lines represent position of basement membrane separating epithelium from stroma.

    Techniques Used: Quantitative RT-PCR, In Situ, Staining, Labeling

    8) Product Images from "Microarray and deep sequencing cross-platform analysis of the mirRNome and isomiR variation in response to epidermal growth factor"

    Article Title: Microarray and deep sequencing cross-platform analysis of the mirRNome and isomiR variation in response to epidermal growth factor

    Journal: BMC Genomics

    doi: 10.1186/1471-2164-14-371

    miRNA transcriptome profiles of EGF-treated HeLa cells using Exiqon and Agilent miRNA arrays. A . Shared miRNA genes between Exiqon and Agilent microRNA arrays. Venn diagram showing the unique and named miRNAs genes shared between Agilent and Exiqon microarray platforms. The pool of 346 shared genes was used for all subsequent cross-platform analyses. Numbers outside diagrams indicate the total amount of miRNA genes contained in each platform. In brackets, percentage of miRNA genes contained in each platform over the total miRNA genes contained in all platforms. B . Regulated miRNAs after 6 hours EGF treatment using Exiqon and Agilent miRNA arrays. HeLa cells were serum-starved for 24 h and treated with EGF for 6 h. Total RNAs prepared from cells lysates were hybridized to Exiqon miRCURY LNA microRNA Array V9.2 and Agilent Human miRNA V2 Oligo Microarray. List shows regulated miRNAs identified by SAM analysis (FDR = 0.05) for miRNAs with a minimal expression change of 1.2 fold.
    Figure Legend Snippet: miRNA transcriptome profiles of EGF-treated HeLa cells using Exiqon and Agilent miRNA arrays. A . Shared miRNA genes between Exiqon and Agilent microRNA arrays. Venn diagram showing the unique and named miRNAs genes shared between Agilent and Exiqon microarray platforms. The pool of 346 shared genes was used for all subsequent cross-platform analyses. Numbers outside diagrams indicate the total amount of miRNA genes contained in each platform. In brackets, percentage of miRNA genes contained in each platform over the total miRNA genes contained in all platforms. B . Regulated miRNAs after 6 hours EGF treatment using Exiqon and Agilent miRNA arrays. HeLa cells were serum-starved for 24 h and treated with EGF for 6 h. Total RNAs prepared from cells lysates were hybridized to Exiqon miRCURY LNA microRNA Array V9.2 and Agilent Human miRNA V2 Oligo Microarray. List shows regulated miRNAs identified by SAM analysis (FDR = 0.05) for miRNAs with a minimal expression change of 1.2 fold.

    Techniques Used: Microarray, Expressing

    RT-qPCR validation of EGF-regulated miRNAs. Total RNA prepared from cells lysates were analyzed by quantitative real time PCR using the miRCURY LNA™ microRNA PCR System (Exiqon) for each of the miRNAs as indicated. A . Control treatment without inhibitors: HeLa cells were serum-starved for 24 hours and treated with EGF for 1 and 6 hours. B . Inhibitor treatment. HeLa cells were serum-starved for 24 hours and treated with EGF for 6 hours in the presence or absence of protein kinase inhibitors: AG1470 (EGFR inhibitor), U0126 (MEK inhibitor) and Wortmannin (PI3K inhibitor). In addition, HeLa cells were transfected with a constitutively active form of Ras (RasV12). Effective pathway inhibition was verified by western blotting in parallel samples from the same experiment (see Additional file 8 ).
    Figure Legend Snippet: RT-qPCR validation of EGF-regulated miRNAs. Total RNA prepared from cells lysates were analyzed by quantitative real time PCR using the miRCURY LNA™ microRNA PCR System (Exiqon) for each of the miRNAs as indicated. A . Control treatment without inhibitors: HeLa cells were serum-starved for 24 hours and treated with EGF for 1 and 6 hours. B . Inhibitor treatment. HeLa cells were serum-starved for 24 hours and treated with EGF for 6 hours in the presence or absence of protein kinase inhibitors: AG1470 (EGFR inhibitor), U0126 (MEK inhibitor) and Wortmannin (PI3K inhibitor). In addition, HeLa cells were transfected with a constitutively active form of Ras (RasV12). Effective pathway inhibition was verified by western blotting in parallel samples from the same experiment (see Additional file 8 ).

    Techniques Used: Quantitative RT-PCR, Real-time Polymerase Chain Reaction, Polymerase Chain Reaction, Transfection, Inhibition, Western Blot

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    Quantitation Assay:

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    Hybridization:

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    Transfection:

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    Activation Assay:

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    Serial Dilution:

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    Northern Blot:

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    TaqMan microRNA Assay:

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    Digital PCR:

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    Polymerase Chain Reaction:

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    Article Snippet: .. All reactions were performed using the miRCURY LNA™ microRNA PCR system (Exiqon, Denmark) according to the manufacturer's instructions [ ]. .. The cycling conditions were 95°C for 10 min, followed by 42 cycles at 95°C for 20 s and 60°C for 1 min. All cycles were completed on an Mx3005P cycler (Stratagene, USA).

    Article Title: Visualization of mucosal field in HPV positive and negative oropharyngeal squamous cell carcinomas: combined genomic and radiology based 3D model
    Article Snippet: .. Droplet digital PCR analysis Regarding quantitative PCR analysis, we procured the Qiagen miRCury LNA miRNA PCR assays (Cat. no. Qiagen, Hilden, Denmark), in accordance with the following miRNA targets: hsa-miR-21-5p, hsa-miR-143, hsa-miR-155 and hsa-miR-221-5p. .. Master mixes were prepared to include 1 µl of miRCury LNA miRNA assay containing the target-miRNA specific forward and reverse primer pair, 12 µl of QX200 ddPCR EvaGreen Supermix (cat. no. 1864034; Bio-Rad) and 9 µl PCR grade water and 2 µl (~100 pg) of cDNA sample.

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    Article Snippet: .. For quantification of microRNA expression, qRT-PCR was performed using the miRCURY LNA™ PCR system (Exiqon, Vedbaek, Denmark) in accordance with the manufacturer′s instructions. .. Total RNA was diluted to a concentration of 10 ng in 4.5 μl using nuclease-free water and added to reagents from the miRCURY LNA™ First-strand cDNA Synthesis Kit.

    Cellular Antioxidant Activity Assay:

    Article Title: Decreased miR122 in hepatocellular carcinoma leads to chemoresistance with increased arginine
    Article Snippet: Hybridization was performed overnight at 42°C in ULTRAhyb-Oligo Buffer (Ambion) containing a biotinylated probe specific for miR122 (caa aca cca ttg tca cac tcc a), which had previously been heated to 95°C for 2 min. Membranes were washed at 42°C in 2 × SSC containing 0.1% SDS and the bound probe was visualized using a BrightStar BioDetect Kit (Ambion). .. To measure the amount of miR122 in the cells, a miRCURY LNA microRNA qPCR system (Exiqon, Vedbaek, Denmark) was used according to the manufacturer's instructions.

    Fluorescence:

    Article Title: miR-7 Modulates hESC Differentiation into Insulin-Producing Beta-like Cells and Contributes to Cell Maturation
    Article Snippet: Prior to qRT-PCR reactions, cDNA was diluted 1 in 80 for Exiqon assays. miRCURY LNA Universal RT microRNA PCR assays were performed using SYBR Green MasterMix Universal RT (Exiqon), according to the manufacturer’s instructions. .. ViiA 7 Software version 1.2 (Life Technologies, Carlsbad, CA, USA) was used to calculate the quantification cycle (Ct) value, which is defined as the number of cycles at which the fluorescence signal is significantly above the threshold; expression of each mRNA and miRNA was defined from the threshold cycle (Ct), and relative expression levels were calculated using the 2−ΔΔCt method after normalization with reference to expression of internal control.

    Isolation:

    Article Title: Stemness, Pluripotentiality, and Wnt Antagonism: sFRP4, a Wnt antagonist Mediates Pluripotency and Stemness in Glioblastoma
    Article Snippet: .. MicroRNA 885 Analysis Total RNA was isolated from U87 treated cells by using the Trizol method. miRCURY LNATM Universal RT microRNA PCR kit (Exiqon, Woburn, MA, USA), hsa-miR-885-5p (Accession ID: MIMAT0004947; Sequence-UCCAUUACACUACCCUGCCUCU) and hsa-miR-885-3p (Accession ID: MIMAT0004948; Sequence-AGGCAGCGGGGUGUAGUGGAUA) were used in the miRNA885 analysis. miRCURY LNA miRNA Detection probe (hsa-miR-885-3p; Exiqon) was used to localize mature miRNA activity in treated U87 cells by using in situ hybridization as previously described [ ]. .. RNA Sequencing Total RNA from U87 cells subjected to either sFRP4 OE or sFRP4 silencing and untreated cells was isolated and sequenced with Illumina HiSeq 2500 System (Illumina Inc.) for the whole transcriptome analysis.

    Article Title: miR-7 Modulates hESC Differentiation into Insulin-Producing Beta-like Cells and Contributes to Cell Maturation
    Article Snippet: RNA Extraction and qRT-PCR Total RNA from cells was isolated using TRIzol (Invitrogen, Carlsbad, CA, USA) according to the manufacturer’s instructions. .. Prior to qRT-PCR reactions, cDNA was diluted 1 in 80 for Exiqon assays. miRCURY LNA Universal RT microRNA PCR assays were performed using SYBR Green MasterMix Universal RT (Exiqon), according to the manufacturer’s instructions.

    Article Title: Microarray and deep sequencing cross-platform analysis of the mirRNome and isomiR variation in response to epidermal growth factor
    Article Snippet: .. miRNA RT-qPCR Quantitative real time PCR was performed using the miRCURY LNA™ microRNA PCR System (Exiqon) on total RNA extracted from HeLa cells treated at different times with EGF (with or without protein kinase inhibitors) with miRVana’s isolation kit (Ambion) following the manufacturer’s instructions. .. PCR amplification and detection were performed with the ROCHE LightCycler 480 detector, using 2x SYBR GREEN Master Mix.

    Article Title: Effects of repeated sprints training on fracture risk-associated miRNA
    Article Snippet: Circulating miRNA analyses MicroRNA-enriched total RNA was extracted from plasma with a miRCURY™ RNA Isolation Kit (Exiqon A/S, Denmark). .. Real-time PCR was performed using specific miRCURY LNA™ microRNA PCR primers (Exiqon; ).

    Size-exclusion Chromatography:

    Article Title: Microarray and deep sequencing cross-platform analysis of the mirRNome and isomiR variation in response to epidermal growth factor
    Article Snippet: miRNA RT-qPCR Quantitative real time PCR was performed using the miRCURY LNA™ microRNA PCR System (Exiqon) on total RNA extracted from HeLa cells treated at different times with EGF (with or without protein kinase inhibitors) with miRVana’s isolation kit (Ambion) following the manufacturer’s instructions. .. The reaction profile was: Polymerase Activation/Denaturation cycle (95°C for 10 min) followed by 40 amplification cycles (95°C-10 sec, 60°C-20 sec). miRNA levels were calculated using the LightCycler 480 software.

    Titration:

    Article Title: Evaluation of extraction kits and RT-qPCR systems adapted to high-throughput platform for circulating miRNAs
    Article Snippet: Reverse transcription and quantitative real-time PCR (RT-qPCR) High-throughput RT-qPCR was carried out on three different systems: TaqMan miRNA PCR system (Applied Biosystems, California, USA), miRCURY LNA microRNA PCR system (Exiqon) and miScript miRNA PCR system (Qiagen). .. Samples from serial dilution of pooled synthetic miRNAs (Integrated DNA Technologies) and pooled cell line RNAs were used to construct standard curves and titration curves, respectively.

    Sequencing:

    Article Title: Distinct MicroRNAs Expression Profile in Primary Biliary Cirrhosis and Evaluation of miR 505-3p and miR197-3p as Novel Biomarkers
    Article Snippet: The relative expression levels from 9 differentially expressed miRNAs were analyzed with the TaqMan MicroRNA assay (Applied Biosystems) or miRCURY LNA microRNA PCR system (Exiqon). .. The relative expression levels from 9 differentially expressed miRNAs were analyzed with the TaqMan MicroRNA assay (Applied Biosystems) or miRCURY LNA microRNA PCR system (Exiqon).

    Article Title: Stemness, Pluripotentiality, and Wnt Antagonism: sFRP4, a Wnt antagonist Mediates Pluripotency and Stemness in Glioblastoma
    Article Snippet: .. MicroRNA 885 Analysis Total RNA was isolated from U87 treated cells by using the Trizol method. miRCURY LNATM Universal RT microRNA PCR kit (Exiqon, Woburn, MA, USA), hsa-miR-885-5p (Accession ID: MIMAT0004947; Sequence-UCCAUUACACUACCCUGCCUCU) and hsa-miR-885-3p (Accession ID: MIMAT0004948; Sequence-AGGCAGCGGGGUGUAGUGGAUA) were used in the miRNA885 analysis. miRCURY LNA miRNA Detection probe (hsa-miR-885-3p; Exiqon) was used to localize mature miRNA activity in treated U87 cells by using in situ hybridization as previously described [ ]. .. RNA Sequencing Total RNA from U87 cells subjected to either sFRP4 OE or sFRP4 silencing and untreated cells was isolated and sequenced with Illumina HiSeq 2500 System (Illumina Inc.) for the whole transcriptome analysis.

    RNA Extraction:

    Article Title: miR-7 Modulates hESC Differentiation into Insulin-Producing Beta-like Cells and Contributes to Cell Maturation
    Article Snippet: Paragraph title: RNA Extraction and qRT-PCR ... Prior to qRT-PCR reactions, cDNA was diluted 1 in 80 for Exiqon assays. miRCURY LNA Universal RT microRNA PCR assays were performed using SYBR Green MasterMix Universal RT (Exiqon), according to the manufacturer’s instructions.

    Construct:

    Article Title: Evaluation of extraction kits and RT-qPCR systems adapted to high-throughput platform for circulating miRNAs
    Article Snippet: Reverse transcription and quantitative real-time PCR (RT-qPCR) High-throughput RT-qPCR was carried out on three different systems: TaqMan miRNA PCR system (Applied Biosystems, California, USA), miRCURY LNA microRNA PCR system (Exiqon) and miScript miRNA PCR system (Qiagen). .. Samples from serial dilution of pooled synthetic miRNAs (Integrated DNA Technologies) and pooled cell line RNAs were used to construct standard curves and titration curves, respectively.

    In Situ Hybridization:

    Article Title: Stemness, Pluripotentiality, and Wnt Antagonism: sFRP4, a Wnt antagonist Mediates Pluripotency and Stemness in Glioblastoma
    Article Snippet: .. MicroRNA 885 Analysis Total RNA was isolated from U87 treated cells by using the Trizol method. miRCURY LNATM Universal RT microRNA PCR kit (Exiqon, Woburn, MA, USA), hsa-miR-885-5p (Accession ID: MIMAT0004947; Sequence-UCCAUUACACUACCCUGCCUCU) and hsa-miR-885-3p (Accession ID: MIMAT0004948; Sequence-AGGCAGCGGGGUGUAGUGGAUA) were used in the miRNA885 analysis. miRCURY LNA miRNA Detection probe (hsa-miR-885-3p; Exiqon) was used to localize mature miRNA activity in treated U87 cells by using in situ hybridization as previously described [ ]. .. RNA Sequencing Total RNA from U87 cells subjected to either sFRP4 OE or sFRP4 silencing and untreated cells was isolated and sequenced with Illumina HiSeq 2500 System (Illumina Inc.) for the whole transcriptome analysis.

    Software:

    Article Title: miR-7 Modulates hESC Differentiation into Insulin-Producing Beta-like Cells and Contributes to Cell Maturation
    Article Snippet: Analysis (qRT-PCR) was performed in triplicate using a ViiA 7 Real-Time PCR System (Applied Biosystems, Foster City, CA, USA), ViiA 7 Software (Applied Biosystems), and SYBR Green SuperMix Low ROX (BIOLINE, Luckenwalde, Germany), using the standard instrument protocol. mRNA levels were normalized to the expression of RPLP (TATAA-Biocenter, Gothenburg, Sweden). .. Prior to qRT-PCR reactions, cDNA was diluted 1 in 80 for Exiqon assays. miRCURY LNA Universal RT microRNA PCR assays were performed using SYBR Green MasterMix Universal RT (Exiqon), according to the manufacturer’s instructions.

    Article Title: Microarray and deep sequencing cross-platform analysis of the mirRNome and isomiR variation in response to epidermal growth factor
    Article Snippet: miRNA RT-qPCR Quantitative real time PCR was performed using the miRCURY LNA™ microRNA PCR System (Exiqon) on total RNA extracted from HeLa cells treated at different times with EGF (with or without protein kinase inhibitors) with miRVana’s isolation kit (Ambion) following the manufacturer’s instructions. .. The reaction profile was: Polymerase Activation/Denaturation cycle (95°C for 10 min) followed by 40 amplification cycles (95°C-10 sec, 60°C-20 sec). miRNA levels were calculated using the LightCycler 480 software.

    SYBR Green Assay:

    Article Title: miR-7 Modulates hESC Differentiation into Insulin-Producing Beta-like Cells and Contributes to Cell Maturation
    Article Snippet: .. Prior to qRT-PCR reactions, cDNA was diluted 1 in 80 for Exiqon assays. miRCURY LNA Universal RT microRNA PCR assays were performed using SYBR Green MasterMix Universal RT (Exiqon), according to the manufacturer’s instructions. .. Reactions were performed, according to the manufacturer’s instructions, using a ViiA 7 Real-Time PCR System (Applied Biosystems). gives details of the qRT-PCR primers used.

    Article Title: Microarray and deep sequencing cross-platform analysis of the mirRNome and isomiR variation in response to epidermal growth factor
    Article Snippet: miRNA RT-qPCR Quantitative real time PCR was performed using the miRCURY LNA™ microRNA PCR System (Exiqon) on total RNA extracted from HeLa cells treated at different times with EGF (with or without protein kinase inhibitors) with miRVana’s isolation kit (Ambion) following the manufacturer’s instructions. .. PCR amplification and detection were performed with the ROCHE LightCycler 480 detector, using 2x SYBR GREEN Master Mix.

    Article Title: Serum miR-146a and miR-150 as Potential New Biomarkers for Hip Fracture-Induced Acute Lung Injury
    Article Snippet: All reactions were performed using the miRCURY LNA™ microRNA PCR system (Exiqon, Denmark) according to the manufacturer's instructions [ ]. .. The miRNA expression levels were measured using SYBR green qRT-PCR, normalised to those of U6 snRNA, and calculated by the comparative CT method.

    Article Title: MicroRNA-215 Regulates Fibroblast Function: Insights from a Human Fibrotic Disease
    Article Snippet: For quantification of microRNA expression, qRT-PCR was performed using the miRCURY LNA™ PCR system (Exiqon, Vedbaek, Denmark) in accordance with the manufacturer′s instructions. .. Resultant complementary cDNA products were diluted 2-fold and 4 μl was added to reagents in the miRCURY LNA™ SYBR® Green Master Mix.

    Negative Control:

    Article Title: Evaluation of extraction kits and RT-qPCR systems adapted to high-throughput platform for circulating miRNAs
    Article Snippet: Reverse transcription and quantitative real-time PCR (RT-qPCR) High-throughput RT-qPCR was carried out on three different systems: TaqMan miRNA PCR system (Applied Biosystems, California, USA), miRCURY LNA microRNA PCR system (Exiqon) and miScript miRNA PCR system (Qiagen). .. Nuclease free water was used as negative control.

    Concentration Assay:

    Article Title: Visualization of mucosal field in HPV positive and negative oropharyngeal squamous cell carcinomas: combined genomic and radiology based 3D model
    Article Snippet: Droplet digital PCR analysis Regarding quantitative PCR analysis, we procured the Qiagen miRCury LNA miRNA PCR assays (Cat. no. Qiagen, Hilden, Denmark), in accordance with the following miRNA targets: hsa-miR-21-5p, hsa-miR-143, hsa-miR-155 and hsa-miR-221-5p. .. Application onto the plate was perform using approximately 100 pg/µl template concentration.

    Article Title: MicroRNA-215 Regulates Fibroblast Function: Insights from a Human Fibrotic Disease
    Article Snippet: For quantification of microRNA expression, qRT-PCR was performed using the miRCURY LNA™ PCR system (Exiqon, Vedbaek, Denmark) in accordance with the manufacturer′s instructions. .. Total RNA was diluted to a concentration of 10 ng in 4.5 μl using nuclease-free water and added to reagents from the miRCURY LNA™ First-strand cDNA Synthesis Kit.

    High Throughput Screening Assay:

    Article Title: Evaluation of extraction kits and RT-qPCR systems adapted to high-throughput platform for circulating miRNAs
    Article Snippet: .. Reverse transcription and quantitative real-time PCR (RT-qPCR) High-throughput RT-qPCR was carried out on three different systems: TaqMan miRNA PCR system (Applied Biosystems, California, USA), miRCURY LNA microRNA PCR system (Exiqon) and miScript miRNA PCR system (Qiagen). .. The RT, preamplification and qPCR reactions were carried out according to previous publication and as indicated in .

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    Qiagen miscript mirna pcr system
    <t>miRNA</t> recovery of different extraction kits were evaluated by the C q values for spike-in controls (cel-miR-39 and cel-miR-54) assayed in the (a) TaqMan and (b) <t>miScript</t> RT-qPCR system. Box plots with whiskers show 5–95 percentile of the C q values for spike-in controls and the corresponding CVs are shown. The extent of extraction bias across samples was reflected in the range and CV of C q values for spike-in controls. Higher median C q compared to other extraction kits indicate poorer performance in miRNA recovery. Q, miRNeasy Serum/Plasma kit; E, miRCURY™ RNA Isolation Kit - Biofluids; A, mirVana™ PARIS™ Kit; MN, NucleoSpin® miRNA Plasma; NB, Plasma/Serum Circulating RNA Purification Kit.
    Miscript Mirna Pcr System, supplied by Qiagen, used in various techniques. Bioz Stars score: 90/100, based on 13 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/miscript mirna pcr system/product/Qiagen
    Average 90 stars, based on 13 article reviews
    Price from $9.99 to $1999.99
    miscript mirna pcr system - by Bioz Stars, 2020-04
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    99
    Qiagen mircury lna sybr green pcr kit
    <t>miRNA</t> recovery of different extraction kits were evaluated by the C q values for spike-in controls (cel-miR-39 and cel-miR-54) assayed in the (a) TaqMan and (b) <t>miScript</t> RT-qPCR system. Box plots with whiskers show 5–95 percentile of the C q values for spike-in controls and the corresponding CVs are shown. The extent of extraction bias across samples was reflected in the range and CV of C q values for spike-in controls. Higher median C q compared to other extraction kits indicate poorer performance in miRNA recovery. Q, miRNeasy Serum/Plasma kit; E, miRCURY™ RNA Isolation Kit - Biofluids; A, mirVana™ PARIS™ Kit; MN, NucleoSpin® miRNA Plasma; NB, Plasma/Serum Circulating RNA Purification Kit.
    Mircury Lna Sybr Green Pcr Kit, supplied by Qiagen, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mircury lna sybr green pcr kit/product/Qiagen
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    mircury lna sybr green pcr kit - by Bioz Stars, 2020-04
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    miRNA recovery of different extraction kits were evaluated by the C q values for spike-in controls (cel-miR-39 and cel-miR-54) assayed in the (a) TaqMan and (b) miScript RT-qPCR system. Box plots with whiskers show 5–95 percentile of the C q values for spike-in controls and the corresponding CVs are shown. The extent of extraction bias across samples was reflected in the range and CV of C q values for spike-in controls. Higher median C q compared to other extraction kits indicate poorer performance in miRNA recovery. Q, miRNeasy Serum/Plasma kit; E, miRCURY™ RNA Isolation Kit - Biofluids; A, mirVana™ PARIS™ Kit; MN, NucleoSpin® miRNA Plasma; NB, Plasma/Serum Circulating RNA Purification Kit.

    Journal: Scientific Reports

    Article Title: Evaluation of extraction kits and RT-qPCR systems adapted to high-throughput platform for circulating miRNAs

    doi: 10.1038/srep09430

    Figure Lengend Snippet: miRNA recovery of different extraction kits were evaluated by the C q values for spike-in controls (cel-miR-39 and cel-miR-54) assayed in the (a) TaqMan and (b) miScript RT-qPCR system. Box plots with whiskers show 5–95 percentile of the C q values for spike-in controls and the corresponding CVs are shown. The extent of extraction bias across samples was reflected in the range and CV of C q values for spike-in controls. Higher median C q compared to other extraction kits indicate poorer performance in miRNA recovery. Q, miRNeasy Serum/Plasma kit; E, miRCURY™ RNA Isolation Kit - Biofluids; A, mirVana™ PARIS™ Kit; MN, NucleoSpin® miRNA Plasma; NB, Plasma/Serum Circulating RNA Purification Kit.

    Article Snippet: Reverse transcription and quantitative real-time PCR (RT-qPCR) High-throughput RT-qPCR was carried out on three different systems: TaqMan miRNA PCR system (Applied Biosystems, California, USA), miRCURY LNA microRNA PCR system (Exiqon) and miScript miRNA PCR system (Qiagen).

    Techniques: Quantitative RT-PCR, Isolation, Purification

    Comparison of TaqMan and miScript RT-qPCR systems adapted to the high-throughput platform by performance parameters which include reproducibility, accuracy and sensitivity. Transformation of data to z-score was carried out to facilitate direct comparison. A1, accuracy when measuring fold change of abundant miRNAs; A2, accuracy when measuring fold change of less abundant miRNAs, R1, reproducibility of RT replicates; R2, reproducibility of RT replicates after normalisation; R3, reproducibility of qPCR replicates; S1, sensitivity to detect the presence of plasma miRNAs and S2, sensitivity to detect the presence of miRNAs in titration points.

    Journal: Scientific Reports

    Article Title: Evaluation of extraction kits and RT-qPCR systems adapted to high-throughput platform for circulating miRNAs

    doi: 10.1038/srep09430

    Figure Lengend Snippet: Comparison of TaqMan and miScript RT-qPCR systems adapted to the high-throughput platform by performance parameters which include reproducibility, accuracy and sensitivity. Transformation of data to z-score was carried out to facilitate direct comparison. A1, accuracy when measuring fold change of abundant miRNAs; A2, accuracy when measuring fold change of less abundant miRNAs, R1, reproducibility of RT replicates; R2, reproducibility of RT replicates after normalisation; R3, reproducibility of qPCR replicates; S1, sensitivity to detect the presence of plasma miRNAs and S2, sensitivity to detect the presence of miRNAs in titration points.

    Article Snippet: Reverse transcription and quantitative real-time PCR (RT-qPCR) High-throughput RT-qPCR was carried out on three different systems: TaqMan miRNA PCR system (Applied Biosystems, California, USA), miRCURY LNA microRNA PCR system (Exiqon) and miScript miRNA PCR system (Qiagen).

    Techniques: Quantitative RT-PCR, High Throughput Screening Assay, Transformation Assay, Real-time Polymerase Chain Reaction, Titration

    Evaluation of consistency between technical replicatesin RT-qPCR. Consistencies among (a) RT replicates in TaqMan system, (b) RT replicates in miScript system, (c) qPCR replicates in TaqMan system and (d) qPCR replicates in miScript system were evaluated by measuring the levels of 11–16 miRNAs in a same set of RNA samples (n = 5–14). Each data point in scatter plot represents the average C q value obtained from the duplicate or triplicate wells of qPCR. Overall, significant correlations (ICC = 0.712–0.996, p

    Journal: Scientific Reports

    Article Title: Evaluation of extraction kits and RT-qPCR systems adapted to high-throughput platform for circulating miRNAs

    doi: 10.1038/srep09430

    Figure Lengend Snippet: Evaluation of consistency between technical replicatesin RT-qPCR. Consistencies among (a) RT replicates in TaqMan system, (b) RT replicates in miScript system, (c) qPCR replicates in TaqMan system and (d) qPCR replicates in miScript system were evaluated by measuring the levels of 11–16 miRNAs in a same set of RNA samples (n = 5–14). Each data point in scatter plot represents the average C q value obtained from the duplicate or triplicate wells of qPCR. Overall, significant correlations (ICC = 0.712–0.996, p

    Article Snippet: Reverse transcription and quantitative real-time PCR (RT-qPCR) High-throughput RT-qPCR was carried out on three different systems: TaqMan miRNA PCR system (Applied Biosystems, California, USA), miRCURY LNA microRNA PCR system (Exiqon) and miScript miRNA PCR system (Qiagen).

    Techniques: Quantitative RT-PCR, Real-time Polymerase Chain Reaction, Immunocytochemistry

    Comparison of different RT-qPCR systems adapted to the BioMark high-throughput platform. (a) Average C q values for endogeneous plasma miRNAs were depicted in box plot with whiskers showing the 5–95 percentile. Irrespective of extraction kits being used, within the same set of samples, more undetermined C q values were obtained in the miScript system. (b) Correlation of qPCR results from TaqMan and miScript systems may vary among different miRNA assays. Each data point represents the normalised C q value transformed into z-score. Q, miRNeasy Serum/Plasma kit; E, miRCURY™ RNA Isolation Kit - Biofluids; A, mirVana™ PARIS™ Kit; MN, NucleoSpin® miRNA Plasma; NB, Plasma/Serum Circulating RNA Purification Kit.

    Journal: Scientific Reports

    Article Title: Evaluation of extraction kits and RT-qPCR systems adapted to high-throughput platform for circulating miRNAs

    doi: 10.1038/srep09430

    Figure Lengend Snippet: Comparison of different RT-qPCR systems adapted to the BioMark high-throughput platform. (a) Average C q values for endogeneous plasma miRNAs were depicted in box plot with whiskers showing the 5–95 percentile. Irrespective of extraction kits being used, within the same set of samples, more undetermined C q values were obtained in the miScript system. (b) Correlation of qPCR results from TaqMan and miScript systems may vary among different miRNA assays. Each data point represents the normalised C q value transformed into z-score. Q, miRNeasy Serum/Plasma kit; E, miRCURY™ RNA Isolation Kit - Biofluids; A, mirVana™ PARIS™ Kit; MN, NucleoSpin® miRNA Plasma; NB, Plasma/Serum Circulating RNA Purification Kit.

    Article Snippet: Reverse transcription and quantitative real-time PCR (RT-qPCR) High-throughput RT-qPCR was carried out on three different systems: TaqMan miRNA PCR system (Applied Biosystems, California, USA), miRCURY LNA microRNA PCR system (Exiqon) and miScript miRNA PCR system (Qiagen).

    Techniques: Quantitative RT-PCR, High Throughput Screening Assay, Real-time Polymerase Chain Reaction, Transformation Assay, Isolation, Purification