28s rrna Search Results


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  • 99
    Millipore bovine 28s rrna
    Bovine 28s Rrna, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Agilent technologies 28s ribosomal rna
    Agonal factors and ribonucleic acid <t>(RNA)</t> degradation decrease Average Correlation Index (ACI). The ACI was calculated for the anterior cingulate cortex for 40 subjects and plotted on the y axis in the eight dot plots (A–H) . The x axis shows the corresponding (A) agonal factor score (AFS); (B) brain tissue pH; (C) percentage of 18S and <t>28S</t> ribosomal RNA to total RNA [%(18S+28S)]; (D) ratio of signal intensities for probe sets designed in 3′ and 5′ end region of glyceraldehyde phosphate dehydrogenase (GAPDH) gene (3′/5′ ratio of GAPDH; (E) the percentage of the total number of probe sets detected as present on the array (% Present Call); (F) coefficient of the RNA degradation slope from 5′ to 3′ ends of the averaged probe intensities calculated with the R -statistical program AffyRNAdeg (Degradation slope); (G) postmortem interval (PMI); and (H) freezer interval (FI). Average Correlation Index was significantly correlated with AFS and all of the RNA integrity indicators analyzed, whereas ACI showed no correlation with PMI and FI.
    28s Ribosomal Rna, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 88/100, based on 81 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Agilent technologies 28s 18s rrna ratio
    Ribosomal RNA processing is delayed in BCS patient cells. (A) In fibroblasts, nascent <t>rRNA</t> was metabolically labeled using (methyl- 3 H) methionine for 30 min, and RNA was isolated at twenty minute intervals to follow the processing rate of the large 45S rRNA precursor to the mature <t>18S</t> (arrowhead) and <t>28S</t> species (arrow). Equal counts were separated on a 0.7% agarose gel, and transferred to a positively-charged nylon membrane. The membrane was exposed to a phosphor storage screen, which was scanned using a phosphorimager. The precursor 45S rRNA, the 28S and 18S mature species, and the intermediates are indicated on the left side of the diagram. A representative image of three independent experiments is shown. (B, C) The intensity of each band in (A) was quantified using Image Lab software. The graphs indicate the mean density in arbitrary units of the 18S (B) and 28S rRNA (C) bands at each time point, and the error bars represent standard deviation. For this experiment, the average of three control cell lines and two BCS cell lines are shown. The 18S rRNA level was significantly reduced in BCS cells at the 20 minute time point (*p = 0.0287). (D) A representative image of an rRNA processing experiment in lymphoblasts. The experiment was performed essentially as in (A), except the rRNA was labeled with 32 P i for a twenty minute pulse. Following gel electrophoresis to separate the RNA species, the gel was dried and directly exposed to a phosphor storage screen. (E) and (F) show the intensity of each band. The 18S rRNA was significantly reduced in BCS cells at the 40 minute time point (*p = 0.0292).
    28s 18s Rrna Ratio, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 88/100, based on 85 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Agilent technologies 28s 18s ribosomal rna ratio
    Quantitative RT-PCR analysis of TNF-α regulated genes in U937 cells after sequential treatment with PGSE and TNF-α . U937 cells (1 × 10 6 ) were pretreated with or without 3 mg/ml PGSE for 24 hours and followed by 20 units/ml of TNF-α for 2 hours. DNase-treated <t>RNA</t> samples were reverse transcribed and the levels of mRNA induction of (A) CXCL-10, (B) IL-8 and (C) TNFAIP3 as well as the reference gene <t>18S</t> rRNA were determined by gene-specific TaqMan assays as described in Methods. The levels of induction were relative to the untreated cells. Values represent the average ± SD of three independent experiments and statistically analyzed by two tailed, paired t-test. *: p
    28s 18s Ribosomal Rna Ratio, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 88/100, based on 26 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Thermo Fisher 28s rrna probe
    RNA synthesis and growth of SAD L16, SAD M(R58G), and SAD M(R58E) viruses. (A) BSR cells were infected with the viruses at an MOI of 2, and after 1 and 2 days of infection, the RNA was isolated from the cells and analyzed in Northern hybridization with N or L probes. The amount of cellular <t>28S</t> <t>rRNA</t> at 1 day postinfection (dpi) was detected in a second hybridization step with 28S cDNA probe. n.i., not infected. (B) Transcription rates of SAD L16, SAD M(R58G), and SAD M(R58E) viruses. The transcription rates were calculated for each virus after quantitation of the N (1 and 2 dpi) and L (1 dpi) hybridization signals by phosphorimaging and normalization of the transcript levels with the genome RNA levels. The transcription rates are given as percentages (SAD L16 transcription rate, 100%). (C) Levels of virus RNAs in infected cells. The amounts of virus-specific RNAs in infected cells, detected with the N probe, were normalized to that of 28S rRNA. In SAD M(R58G)-infected cells, the level of virus-expressed RNAs increased to 239% (SAD L16 level, 100%), whereas the virus RNA levels in SAD M(R58E)-infected cells decreased to 27%. (D) One-step growth curves. The infectious virus titers were determined in supernatants of identical infection experiments at days 1, 2, and 3 postinfection.
    28s Rrna Probe, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 85/100, based on 12 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Thermo Fisher 28s ribosomal rna
    Induction of vascular endothelial growth factor (VEGF) mRNA expression by TGF-β and IL-1. Synovial fibroblasts were grown to confluence in T-75 flasks, incubated for another 48 h in serum-free medium and then fresh medium was added with or without TGF-β, IL-1 or both combined (10 ng/ml). After 6 h at 37°C total <t>RNA</t> was isolated. (a) Northern hybridization. Total RNA (10 μg/lane) and RNA size standards were electrophoresed and blotted onto nylon membranes and hybridized with a VEGF-specific 32 P-labelled cDNA. The VEGF transcript is seen at 4.2 kb. The blot was stripped and rehybridized with a <t>28S</t> RNA-specific labelled probe (28S). (b) Densitometric analysis. The bands on exposed films were imaged and digitized and the number of pixels associated with each band quantified. Induction of VEGF mRNA transcripts in cells treated with TGF-β and IL-1 combined for 6 h was increased approx. 25-fold above control cells.
    28s Ribosomal Rna, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 91/100, based on 31 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher 18 28s rrna
    Induction of vascular endothelial growth factor (VEGF) mRNA expression by TGF-β and IL-1. Synovial fibroblasts were grown to confluence in T-75 flasks, incubated for another 48 h in serum-free medium and then fresh medium was added with or without TGF-β, IL-1 or both combined (10 ng/ml). After 6 h at 37°C total <t>RNA</t> was isolated. (a) Northern hybridization. Total RNA (10 μg/lane) and RNA size standards were electrophoresed and blotted onto nylon membranes and hybridized with a VEGF-specific 32 P-labelled cDNA. The VEGF transcript is seen at 4.2 kb. The blot was stripped and rehybridized with a <t>28S</t> RNA-specific labelled probe (28S). (b) Densitometric analysis. The bands on exposed films were imaged and digitized and the number of pixels associated with each band quantified. Induction of VEGF mRNA transcripts in cells treated with TGF-β and IL-1 combined for 6 h was increased approx. 25-fold above control cells.
    18 28s Rrna, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 88/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Agilent technologies 28s 18s ribosomal rna rrna ratio
    Induction of vascular endothelial growth factor (VEGF) mRNA expression by TGF-β and IL-1. Synovial fibroblasts were grown to confluence in T-75 flasks, incubated for another 48 h in serum-free medium and then fresh medium was added with or without TGF-β, IL-1 or both combined (10 ng/ml). After 6 h at 37°C total <t>RNA</t> was isolated. (a) Northern hybridization. Total RNA (10 μg/lane) and RNA size standards were electrophoresed and blotted onto nylon membranes and hybridized with a VEGF-specific 32 P-labelled cDNA. The VEGF transcript is seen at 4.2 kb. The blot was stripped and rehybridized with a <t>28S</t> RNA-specific labelled probe (28S). (b) Densitometric analysis. The bands on exposed films were imaged and digitized and the number of pixels associated with each band quantified. Induction of VEGF mRNA transcripts in cells treated with TGF-β and IL-1 combined for 6 h was increased approx. 25-fold above control cells.
    28s 18s Ribosomal Rna Rrna Ratio, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 86/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Agilent technologies 28s 18s ribosomal rna peaks
    Induction of vascular endothelial growth factor (VEGF) mRNA expression by TGF-β and IL-1. Synovial fibroblasts were grown to confluence in T-75 flasks, incubated for another 48 h in serum-free medium and then fresh medium was added with or without TGF-β, IL-1 or both combined (10 ng/ml). After 6 h at 37°C total <t>RNA</t> was isolated. (a) Northern hybridization. Total RNA (10 μg/lane) and RNA size standards were electrophoresed and blotted onto nylon membranes and hybridized with a VEGF-specific 32 P-labelled cDNA. The VEGF transcript is seen at 4.2 kb. The blot was stripped and rehybridized with a <t>28S</t> RNA-specific labelled probe (28S). (b) Densitometric analysis. The bands on exposed films were imaged and digitized and the number of pixels associated with each band quantified. Induction of VEGF mRNA transcripts in cells treated with TGF-β and IL-1 combined for 6 h was increased approx. 25-fold above control cells.
    28s 18s Ribosomal Rna Peaks, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 85/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Stratagene 28s 18s rrna
    Induction of vascular endothelial growth factor (VEGF) mRNA expression by TGF-β and IL-1. Synovial fibroblasts were grown to confluence in T-75 flasks, incubated for another 48 h in serum-free medium and then fresh medium was added with or without TGF-β, IL-1 or both combined (10 ng/ml). After 6 h at 37°C total <t>RNA</t> was isolated. (a) Northern hybridization. Total RNA (10 μg/lane) and RNA size standards were electrophoresed and blotted onto nylon membranes and hybridized with a VEGF-specific 32 P-labelled cDNA. The VEGF transcript is seen at 4.2 kb. The blot was stripped and rehybridized with a <t>28S</t> RNA-specific labelled probe (28S). (b) Densitometric analysis. The bands on exposed films were imaged and digitized and the number of pixels associated with each band quantified. Induction of VEGF mRNA transcripts in cells treated with TGF-β and IL-1 combined for 6 h was increased approx. 25-fold above control cells.
    28s 18s Rrna, supplied by Stratagene, used in various techniques. Bioz Stars score: 88/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Agilent technologies rrna 28s
    Induction of vascular endothelial growth factor (VEGF) mRNA expression by TGF-β and IL-1. Synovial fibroblasts were grown to confluence in T-75 flasks, incubated for another 48 h in serum-free medium and then fresh medium was added with or without TGF-β, IL-1 or both combined (10 ng/ml). After 6 h at 37°C total <t>RNA</t> was isolated. (a) Northern hybridization. Total RNA (10 μg/lane) and RNA size standards were electrophoresed and blotted onto nylon membranes and hybridized with a VEGF-specific 32 P-labelled cDNA. The VEGF transcript is seen at 4.2 kb. The blot was stripped and rehybridized with a <t>28S</t> RNA-specific labelled probe (28S). (b) Densitometric analysis. The bands on exposed films were imaged and digitized and the number of pixels associated with each band quantified. Induction of VEGF mRNA transcripts in cells treated with TGF-β and IL-1 combined for 6 h was increased approx. 25-fold above control cells.
    Rrna 28s, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 88/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Millipore calf liver 18s 28s ribosomal rna
    Induction of vascular endothelial growth factor (VEGF) mRNA expression by TGF-β and IL-1. Synovial fibroblasts were grown to confluence in T-75 flasks, incubated for another 48 h in serum-free medium and then fresh medium was added with or without TGF-β, IL-1 or both combined (10 ng/ml). After 6 h at 37°C total <t>RNA</t> was isolated. (a) Northern hybridization. Total RNA (10 μg/lane) and RNA size standards were electrophoresed and blotted onto nylon membranes and hybridized with a VEGF-specific 32 P-labelled cDNA. The VEGF transcript is seen at 4.2 kb. The blot was stripped and rehybridized with a <t>28S</t> RNA-specific labelled probe (28S). (b) Densitometric analysis. The bands on exposed films were imaged and digitized and the number of pixels associated with each band quantified. Induction of VEGF mRNA transcripts in cells treated with TGF-β and IL-1 combined for 6 h was increased approx. 25-fold above control cells.
    Calf Liver 18s 28s Ribosomal Rna, supplied by Millipore, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Becton Dickinson human 28s rrna sequence
    Autoradiograph of a Northern blot of total RNA from wild-type mEFs and mEFs containing the knock-in alleles c- jun (S63A S73A) or c- jun (T91A T93A) incubated under aerobic conditions (air [5% CO 2 ]) or exposed to hypoxia (Hx) (pO 2 ≤ 0.01%) for the indicated times. The blot was probed sequentially for c- jun and <t>28S</t> <t>rRNA</t> transcripts.
    Human 28s Rrna Sequence, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 85/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher human 28s rrna gene
    Autoradiograph of a Northern blot of total RNA from wild-type mEFs and mEFs containing the knock-in alleles c- jun (S63A S73A) or c- jun (T91A T93A) incubated under aerobic conditions (air [5% CO 2 ]) or exposed to hypoxia (Hx) (pO 2 ≤ 0.01%) for the indicated times. The blot was probed sequentially for c- jun and <t>28S</t> <t>rRNA</t> transcripts.
    Human 28s Rrna Gene, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 85/100, based on 18 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Agonal factors and ribonucleic acid (RNA) degradation decrease Average Correlation Index (ACI). The ACI was calculated for the anterior cingulate cortex for 40 subjects and plotted on the y axis in the eight dot plots (A–H) . The x axis shows the corresponding (A) agonal factor score (AFS); (B) brain tissue pH; (C) percentage of 18S and 28S ribosomal RNA to total RNA [%(18S+28S)]; (D) ratio of signal intensities for probe sets designed in 3′ and 5′ end region of glyceraldehyde phosphate dehydrogenase (GAPDH) gene (3′/5′ ratio of GAPDH; (E) the percentage of the total number of probe sets detected as present on the array (% Present Call); (F) coefficient of the RNA degradation slope from 5′ to 3′ ends of the averaged probe intensities calculated with the R -statistical program AffyRNAdeg (Degradation slope); (G) postmortem interval (PMI); and (H) freezer interval (FI). Average Correlation Index was significantly correlated with AFS and all of the RNA integrity indicators analyzed, whereas ACI showed no correlation with PMI and FI.

    Journal: Biological psychiatry

    Article Title: Effect of Agonal and Postmortem Factors on Gene Expression Profile: Quality Control in Microarray Analyses of Postmortem Human Brain

    doi: 10.1016/j.biopsych.2003.10.013

    Figure Lengend Snippet: Agonal factors and ribonucleic acid (RNA) degradation decrease Average Correlation Index (ACI). The ACI was calculated for the anterior cingulate cortex for 40 subjects and plotted on the y axis in the eight dot plots (A–H) . The x axis shows the corresponding (A) agonal factor score (AFS); (B) brain tissue pH; (C) percentage of 18S and 28S ribosomal RNA to total RNA [%(18S+28S)]; (D) ratio of signal intensities for probe sets designed in 3′ and 5′ end region of glyceraldehyde phosphate dehydrogenase (GAPDH) gene (3′/5′ ratio of GAPDH; (E) the percentage of the total number of probe sets detected as present on the array (% Present Call); (F) coefficient of the RNA degradation slope from 5′ to 3′ ends of the averaged probe intensities calculated with the R -statistical program AffyRNAdeg (Degradation slope); (G) postmortem interval (PMI); and (H) freezer interval (FI). Average Correlation Index was significantly correlated with AFS and all of the RNA integrity indicators analyzed, whereas ACI showed no correlation with PMI and FI.

    Article Snippet: For the quantification of 18S and 28S ribosomal RNA, 1 μg of each RNA sample was applied to a 2100 Bioanalyzer (Agilent, Palo Alto, California).

    Techniques:

    Ribosomal RNA processing is delayed in BCS patient cells. (A) In fibroblasts, nascent rRNA was metabolically labeled using (methyl- 3 H) methionine for 30 min, and RNA was isolated at twenty minute intervals to follow the processing rate of the large 45S rRNA precursor to the mature 18S (arrowhead) and 28S species (arrow). Equal counts were separated on a 0.7% agarose gel, and transferred to a positively-charged nylon membrane. The membrane was exposed to a phosphor storage screen, which was scanned using a phosphorimager. The precursor 45S rRNA, the 28S and 18S mature species, and the intermediates are indicated on the left side of the diagram. A representative image of three independent experiments is shown. (B, C) The intensity of each band in (A) was quantified using Image Lab software. The graphs indicate the mean density in arbitrary units of the 18S (B) and 28S rRNA (C) bands at each time point, and the error bars represent standard deviation. For this experiment, the average of three control cell lines and two BCS cell lines are shown. The 18S rRNA level was significantly reduced in BCS cells at the 20 minute time point (*p = 0.0287). (D) A representative image of an rRNA processing experiment in lymphoblasts. The experiment was performed essentially as in (A), except the rRNA was labeled with 32 P i for a twenty minute pulse. Following gel electrophoresis to separate the RNA species, the gel was dried and directly exposed to a phosphor storage screen. (E) and (F) show the intensity of each band. The 18S rRNA was significantly reduced in BCS cells at the 40 minute time point (*p = 0.0292).

    Journal: BBA Clinical

    Article Title: Mutation of EMG1 causing Bowen–Conradi syndrome results in reduced cell proliferation rates concomitant with G2/M arrest and 18S rRNA processing delay

    doi: 10.1016/j.bbacli.2014.05.002

    Figure Lengend Snippet: Ribosomal RNA processing is delayed in BCS patient cells. (A) In fibroblasts, nascent rRNA was metabolically labeled using (methyl- 3 H) methionine for 30 min, and RNA was isolated at twenty minute intervals to follow the processing rate of the large 45S rRNA precursor to the mature 18S (arrowhead) and 28S species (arrow). Equal counts were separated on a 0.7% agarose gel, and transferred to a positively-charged nylon membrane. The membrane was exposed to a phosphor storage screen, which was scanned using a phosphorimager. The precursor 45S rRNA, the 28S and 18S mature species, and the intermediates are indicated on the left side of the diagram. A representative image of three independent experiments is shown. (B, C) The intensity of each band in (A) was quantified using Image Lab software. The graphs indicate the mean density in arbitrary units of the 18S (B) and 28S rRNA (C) bands at each time point, and the error bars represent standard deviation. For this experiment, the average of three control cell lines and two BCS cell lines are shown. The 18S rRNA level was significantly reduced in BCS cells at the 20 minute time point (*p = 0.0287). (D) A representative image of an rRNA processing experiment in lymphoblasts. The experiment was performed essentially as in (A), except the rRNA was labeled with 32 P i for a twenty minute pulse. Following gel electrophoresis to separate the RNA species, the gel was dried and directly exposed to a phosphor storage screen. (E) and (F) show the intensity of each band. The 18S rRNA was significantly reduced in BCS cells at the 40 minute time point (*p = 0.0292).

    Article Snippet: 2.9 Determination of 28S/18S rRNA ratio For each cell line, three independent samples of RNA were analyzed by capillary electrophoresis using the RNA 6000 Nano LabChip Kit in the Agilent 2100 Bioanalyzer according to the manufacturer's instructions.

    Techniques: Metabolic Labelling, Labeling, Isolation, Agarose Gel Electrophoresis, Software, Standard Deviation, Nucleic Acid Electrophoresis

    Ribosomal RNA levels at steady-state are normal in BCS patient cells. (A, B) Total RNA was isolated from unaffected control and BCS-affected lymphoblasts (A) and fibroblasts (B), and separated by capillary chromatography using the 6000 RNA Nano kit in an Agilent Bioanalyzer. The resulting electropherograms show the 18S and 28S peaks, as well as a smaller peak which encompasses the 5S, 5.8S, and tRNA. (C, D) 28S/18S ratios for lymphoblasts (C) and fibroblasts (D). The area of each peak was calculated by the Agilent software, and the ratios of 28S to 18S rRNA were calculated. The mean of three individual experiments, performed in triplicate, and SEM are shown. No significant difference was found between the control and BCS patient cells.

    Journal: BBA Clinical

    Article Title: Mutation of EMG1 causing Bowen–Conradi syndrome results in reduced cell proliferation rates concomitant with G2/M arrest and 18S rRNA processing delay

    doi: 10.1016/j.bbacli.2014.05.002

    Figure Lengend Snippet: Ribosomal RNA levels at steady-state are normal in BCS patient cells. (A, B) Total RNA was isolated from unaffected control and BCS-affected lymphoblasts (A) and fibroblasts (B), and separated by capillary chromatography using the 6000 RNA Nano kit in an Agilent Bioanalyzer. The resulting electropherograms show the 18S and 28S peaks, as well as a smaller peak which encompasses the 5S, 5.8S, and tRNA. (C, D) 28S/18S ratios for lymphoblasts (C) and fibroblasts (D). The area of each peak was calculated by the Agilent software, and the ratios of 28S to 18S rRNA were calculated. The mean of three individual experiments, performed in triplicate, and SEM are shown. No significant difference was found between the control and BCS patient cells.

    Article Snippet: 2.9 Determination of 28S/18S rRNA ratio For each cell line, three independent samples of RNA were analyzed by capillary electrophoresis using the RNA 6000 Nano LabChip Kit in the Agilent 2100 Bioanalyzer according to the manufacturer's instructions.

    Techniques: Isolation, Chromatography, Software

    Involvement of human r-proteins in pre-rRNA processing. ( a ) The 28S/18S ratio calculated from Agilent bioanalyzer electropherograms. Data are shown for the two different siRNAs used (siRNA #1 and #2). ( b ) Major pre-rRNA intermediates and probes used in this work. Three of the four rRNAs are produced by RNA Pol I as a long 47S primary transcript. The 18S, 5.8S and 28S rRNAs are separated by noncoding external (ETS) and internal (ITS) transcribed spacers. Probes a, b and c are the oligonucleotides LD1844, LD1827 and LD1828, respectively (Methods section). ( c ) Pre-rRNA processing inhibitions after depletion of SSU r-proteins. On the northern blots ( www.RibosomalProteins.com , Supplementary Figs 5,6 and 11 ), all RNA species were quantified with a Phosphorimager, normalized with respect to the non-targeting control (SCR), and their abundances represented on a heatmap using the colour code indicated. The heatmap profiles were clustered with ‘R' and the corresponding proteins grouped in classes of r-proteins affecting the same or similar processing steps. The different siRNAs used are indicated (#). Asterisks (*) refer to r-proteins assigned to two groups according to the siRNA used. ( d ) As in c for LSU r-proteins.

    Journal: Nature Communications

    Article Title: Involvement of human ribosomal proteins in nucleolar structure and p53-dependent nucleolar stress

    doi: 10.1038/ncomms11390

    Figure Lengend Snippet: Involvement of human r-proteins in pre-rRNA processing. ( a ) The 28S/18S ratio calculated from Agilent bioanalyzer electropherograms. Data are shown for the two different siRNAs used (siRNA #1 and #2). ( b ) Major pre-rRNA intermediates and probes used in this work. Three of the four rRNAs are produced by RNA Pol I as a long 47S primary transcript. The 18S, 5.8S and 28S rRNAs are separated by noncoding external (ETS) and internal (ITS) transcribed spacers. Probes a, b and c are the oligonucleotides LD1844, LD1827 and LD1828, respectively (Methods section). ( c ) Pre-rRNA processing inhibitions after depletion of SSU r-proteins. On the northern blots ( www.RibosomalProteins.com , Supplementary Figs 5,6 and 11 ), all RNA species were quantified with a Phosphorimager, normalized with respect to the non-targeting control (SCR), and their abundances represented on a heatmap using the colour code indicated. The heatmap profiles were clustered with ‘R' and the corresponding proteins grouped in classes of r-proteins affecting the same or similar processing steps. The different siRNAs used are indicated (#). Asterisks (*) refer to r-proteins assigned to two groups according to the siRNA used. ( d ) As in c for LSU r-proteins.

    Article Snippet: The 28S/18S rRNA ratios were calculated from Agilent bioanalyzer electropherograms according to the manufacturer's instructions.

    Techniques: Produced, Northern Blot

    Systematic screening of human r-proteins reveals that uL5 (RPL11) and uL18 (RPL5) are the strongest contributors to nucleolar structure maintenance. ( a ) Experimental strategy: all 80 r-proteins were depleted one by one in human cells by use of specific siRNAs. The nucleolar structure (fluorescence microscopy), the accumulation of mature 18S and 28S rRNAs (electropherograms), pre-rRNA processing (high-resolution northern blotting), and steady-state accumulation of p53 (fluorescent western blotting) were monitored. ( b ) PCA showing a classification of r-proteins according to their requirement for nucleolar structure maintenance. Each r-protein was depleted in three knockdown experiments, each performed with a different siRNA. The image-processing algorithm that we designed for this analysis involves selecting five discriminant shape and textural features, computing five d k values, and reducing the five dimensions to two by PCA. In the resulting plot, each coloured dot represents one population of cells treated with one siRNA. Dot colour is indicative of the targeted protein: green for SSU r-proteins and magenta for LSU r-proteins. The mean of three populations of cells treated with a non-targeting control siRNA (SCR) is shown in red. Blue symbols represent the six calibration controls (FBL, GFP, nucleolin, nucleophosmin, MOCK and TIF1A, see Supplementary Fig. 1 ). Insets show images of the nuclei of cells depleted of representative proteins with the DNA stained in blue and the nucleoli appearing in green (FBL). For a few representative examples, a specific symbol is used (for example, a diamond for uL5). RPL, r-proteins of the LSU; RPS, r-proteins of the SSU. ( c ) r-proteins and calibration controls classified according to the severity of nucleolar disruption caused by their absence. The iNo was defined as the sum of the d k values of the five most discriminant shape and textural features identified in this work (Methods section). Higher iNo correspond to more severe disruption. Colour-coding as in b . The coloured dots are the means of three individual experiments (shown in grey). Note: the r-proteins are named according to a recently revised nomenclature 24 where the ‘e' prefix stands for eukaryote-specific and ‘u' for universal (present in bacteria, archaea and eukaryotes).

    Journal: Nature Communications

    Article Title: Involvement of human ribosomal proteins in nucleolar structure and p53-dependent nucleolar stress

    doi: 10.1038/ncomms11390

    Figure Lengend Snippet: Systematic screening of human r-proteins reveals that uL5 (RPL11) and uL18 (RPL5) are the strongest contributors to nucleolar structure maintenance. ( a ) Experimental strategy: all 80 r-proteins were depleted one by one in human cells by use of specific siRNAs. The nucleolar structure (fluorescence microscopy), the accumulation of mature 18S and 28S rRNAs (electropherograms), pre-rRNA processing (high-resolution northern blotting), and steady-state accumulation of p53 (fluorescent western blotting) were monitored. ( b ) PCA showing a classification of r-proteins according to their requirement for nucleolar structure maintenance. Each r-protein was depleted in three knockdown experiments, each performed with a different siRNA. The image-processing algorithm that we designed for this analysis involves selecting five discriminant shape and textural features, computing five d k values, and reducing the five dimensions to two by PCA. In the resulting plot, each coloured dot represents one population of cells treated with one siRNA. Dot colour is indicative of the targeted protein: green for SSU r-proteins and magenta for LSU r-proteins. The mean of three populations of cells treated with a non-targeting control siRNA (SCR) is shown in red. Blue symbols represent the six calibration controls (FBL, GFP, nucleolin, nucleophosmin, MOCK and TIF1A, see Supplementary Fig. 1 ). Insets show images of the nuclei of cells depleted of representative proteins with the DNA stained in blue and the nucleoli appearing in green (FBL). For a few representative examples, a specific symbol is used (for example, a diamond for uL5). RPL, r-proteins of the LSU; RPS, r-proteins of the SSU. ( c ) r-proteins and calibration controls classified according to the severity of nucleolar disruption caused by their absence. The iNo was defined as the sum of the d k values of the five most discriminant shape and textural features identified in this work (Methods section). Higher iNo correspond to more severe disruption. Colour-coding as in b . The coloured dots are the means of three individual experiments (shown in grey). Note: the r-proteins are named according to a recently revised nomenclature 24 where the ‘e' prefix stands for eukaryote-specific and ‘u' for universal (present in bacteria, archaea and eukaryotes).

    Article Snippet: The 28S/18S rRNA ratios were calculated from Agilent bioanalyzer electropherograms according to the manufacturer's instructions.

    Techniques: Fluorescence, Microscopy, Northern Blot, Western Blot, Staining

    Quantitative RT-PCR analysis of TNF-α regulated genes in U937 cells after sequential treatment with PGSE and TNF-α . U937 cells (1 × 10 6 ) were pretreated with or without 3 mg/ml PGSE for 24 hours and followed by 20 units/ml of TNF-α for 2 hours. DNase-treated RNA samples were reverse transcribed and the levels of mRNA induction of (A) CXCL-10, (B) IL-8 and (C) TNFAIP3 as well as the reference gene 18S rRNA were determined by gene-specific TaqMan assays as described in Methods. The levels of induction were relative to the untreated cells. Values represent the average ± SD of three independent experiments and statistically analyzed by two tailed, paired t-test. *: p

    Journal: Journal of Translational Medicine

    Article Title: Bioactivity-guided identification and cell signaling technology to delineate the immunomodulatory effects of Panax ginseng on human promonocytic U937 cells

    doi: 10.1186/1479-5876-7-34

    Figure Lengend Snippet: Quantitative RT-PCR analysis of TNF-α regulated genes in U937 cells after sequential treatment with PGSE and TNF-α . U937 cells (1 × 10 6 ) were pretreated with or without 3 mg/ml PGSE for 24 hours and followed by 20 units/ml of TNF-α for 2 hours. DNase-treated RNA samples were reverse transcribed and the levels of mRNA induction of (A) CXCL-10, (B) IL-8 and (C) TNFAIP3 as well as the reference gene 18S rRNA were determined by gene-specific TaqMan assays as described in Methods. The levels of induction were relative to the untreated cells. Values represent the average ± SD of three independent experiments and statistically analyzed by two tailed, paired t-test. *: p

    Article Snippet: The RNA integrity was determined by the ratio of 28S/18S ribosomal RNA using Agilent 2100 Bioanalzyer.

    Techniques: Quantitative RT-PCR, Two Tailed Test

    RNA synthesis and growth of SAD L16, SAD M(R58G), and SAD M(R58E) viruses. (A) BSR cells were infected with the viruses at an MOI of 2, and after 1 and 2 days of infection, the RNA was isolated from the cells and analyzed in Northern hybridization with N or L probes. The amount of cellular 28S rRNA at 1 day postinfection (dpi) was detected in a second hybridization step with 28S cDNA probe. n.i., not infected. (B) Transcription rates of SAD L16, SAD M(R58G), and SAD M(R58E) viruses. The transcription rates were calculated for each virus after quantitation of the N (1 and 2 dpi) and L (1 dpi) hybridization signals by phosphorimaging and normalization of the transcript levels with the genome RNA levels. The transcription rates are given as percentages (SAD L16 transcription rate, 100%). (C) Levels of virus RNAs in infected cells. The amounts of virus-specific RNAs in infected cells, detected with the N probe, were normalized to that of 28S rRNA. In SAD M(R58G)-infected cells, the level of virus-expressed RNAs increased to 239% (SAD L16 level, 100%), whereas the virus RNA levels in SAD M(R58E)-infected cells decreased to 27%. (D) One-step growth curves. The infectious virus titers were determined in supernatants of identical infection experiments at days 1, 2, and 3 postinfection.

    Journal: Journal of Virology

    Article Title: Dissociation of Rabies Virus Matrix Protein Functions in Regulation of Viral RNA Synthesis and Virus Assembly

    doi: 10.1128/JVI.77.22.12074-12082.2003

    Figure Lengend Snippet: RNA synthesis and growth of SAD L16, SAD M(R58G), and SAD M(R58E) viruses. (A) BSR cells were infected with the viruses at an MOI of 2, and after 1 and 2 days of infection, the RNA was isolated from the cells and analyzed in Northern hybridization with N or L probes. The amount of cellular 28S rRNA at 1 day postinfection (dpi) was detected in a second hybridization step with 28S cDNA probe. n.i., not infected. (B) Transcription rates of SAD L16, SAD M(R58G), and SAD M(R58E) viruses. The transcription rates were calculated for each virus after quantitation of the N (1 and 2 dpi) and L (1 dpi) hybridization signals by phosphorimaging and normalization of the transcript levels with the genome RNA levels. The transcription rates are given as percentages (SAD L16 transcription rate, 100%). (C) Levels of virus RNAs in infected cells. The amounts of virus-specific RNAs in infected cells, detected with the N probe, were normalized to that of 28S rRNA. In SAD M(R58G)-infected cells, the level of virus-expressed RNAs increased to 239% (SAD L16 level, 100%), whereas the virus RNA levels in SAD M(R58E)-infected cells decreased to 27%. (D) One-step growth curves. The infectious virus titers were determined in supernatants of identical infection experiments at days 1, 2, and 3 postinfection.

    Article Snippet: To detect cellular RNA, a 28S rRNA probe (Ambion) was used.

    Techniques: Infection, Isolation, Northern Blot, Hybridization, Quantitation Assay

    Induction of vascular endothelial growth factor (VEGF) mRNA expression by TGF-β and IL-1. Synovial fibroblasts were grown to confluence in T-75 flasks, incubated for another 48 h in serum-free medium and then fresh medium was added with or without TGF-β, IL-1 or both combined (10 ng/ml). After 6 h at 37°C total RNA was isolated. (a) Northern hybridization. Total RNA (10 μg/lane) and RNA size standards were electrophoresed and blotted onto nylon membranes and hybridized with a VEGF-specific 32 P-labelled cDNA. The VEGF transcript is seen at 4.2 kb. The blot was stripped and rehybridized with a 28S RNA-specific labelled probe (28S). (b) Densitometric analysis. The bands on exposed films were imaged and digitized and the number of pixels associated with each band quantified. Induction of VEGF mRNA transcripts in cells treated with TGF-β and IL-1 combined for 6 h was increased approx. 25-fold above control cells.

    Journal: Clinical and Experimental Immunology

    Article Title: Hypoxia augments cytokine (transforming growth factor-beta (TGF-?) and IL-1)-induced vascular endothelial growth factor secretion by human synovial fibroblasts

    doi: 10.1046/j.1365-2249.1999.00775.x

    Figure Lengend Snippet: Induction of vascular endothelial growth factor (VEGF) mRNA expression by TGF-β and IL-1. Synovial fibroblasts were grown to confluence in T-75 flasks, incubated for another 48 h in serum-free medium and then fresh medium was added with or without TGF-β, IL-1 or both combined (10 ng/ml). After 6 h at 37°C total RNA was isolated. (a) Northern hybridization. Total RNA (10 μg/lane) and RNA size standards were electrophoresed and blotted onto nylon membranes and hybridized with a VEGF-specific 32 P-labelled cDNA. The VEGF transcript is seen at 4.2 kb. The blot was stripped and rehybridized with a 28S RNA-specific labelled probe (28S). (b) Densitometric analysis. The bands on exposed films were imaged and digitized and the number of pixels associated with each band quantified. Induction of VEGF mRNA transcripts in cells treated with TGF-β and IL-1 combined for 6 h was increased approx. 25-fold above control cells.

    Article Snippet: To quantify RNA loading, the blots were hybridized with a 32 P-labelled cDNA designed to hybridize with 28S ribosomal RNA (pTri-RNA-28S,; Ambion).

    Techniques: Expressing, Incubation, Isolation, Northern Blot, Hybridization

    Combined effect of hypoxic culture and cytokines on vascular endothelial growth factor (VEGF) mRNA levels. After 48 h of culture in serum-free medium, confluent fibroblast cultures were changed to fresh medium with or without addition of TGF-β (10 ng/ml) and IL-1 (10 ng/ml) added together. Replicate flasks were cultured under either normoxic (21% oxygen) or hypoxic (3% oxygen) conditions for 6 h or 24 h. Total RNA was isolated from cells in six flasks in each case, and Northern hybridization for VEGF performed with 5 μg total RNA. The blot was rehybridized for 28S RNA to assess equality of loading The absence or presence of cytokines, the incubation time period and normoxic or hypoxic culture are indicated in the figure.

    Journal: Clinical and Experimental Immunology

    Article Title: Hypoxia augments cytokine (transforming growth factor-beta (TGF-?) and IL-1)-induced vascular endothelial growth factor secretion by human synovial fibroblasts

    doi: 10.1046/j.1365-2249.1999.00775.x

    Figure Lengend Snippet: Combined effect of hypoxic culture and cytokines on vascular endothelial growth factor (VEGF) mRNA levels. After 48 h of culture in serum-free medium, confluent fibroblast cultures were changed to fresh medium with or without addition of TGF-β (10 ng/ml) and IL-1 (10 ng/ml) added together. Replicate flasks were cultured under either normoxic (21% oxygen) or hypoxic (3% oxygen) conditions for 6 h or 24 h. Total RNA was isolated from cells in six flasks in each case, and Northern hybridization for VEGF performed with 5 μg total RNA. The blot was rehybridized for 28S RNA to assess equality of loading The absence or presence of cytokines, the incubation time period and normoxic or hypoxic culture are indicated in the figure.

    Article Snippet: To quantify RNA loading, the blots were hybridized with a 32 P-labelled cDNA designed to hybridize with 28S ribosomal RNA (pTri-RNA-28S,; Ambion).

    Techniques: Cell Culture, Isolation, Northern Blot, Hybridization, Incubation

    Autoradiograph of a Northern blot of total RNA from wild-type mEFs and mEFs containing the knock-in alleles c- jun (S63A S73A) or c- jun (T91A T93A) incubated under aerobic conditions (air [5% CO 2 ]) or exposed to hypoxia (Hx) (pO 2 ≤ 0.01%) for the indicated times. The blot was probed sequentially for c- jun and 28S rRNA transcripts.

    Journal: Molecular and Cellular Biology

    Article Title: The Response of c-Jun/AP-1 to Chronic Hypoxia Is Hypoxia-Inducible Factor 1? Dependent

    doi: 10.1128/MCB.22.8.2515-2523.2002

    Figure Lengend Snippet: Autoradiograph of a Northern blot of total RNA from wild-type mEFs and mEFs containing the knock-in alleles c- jun (S63A S73A) or c- jun (T91A T93A) incubated under aerobic conditions (air [5% CO 2 ]) or exposed to hypoxia (Hx) (pO 2 ≤ 0.01%) for the indicated times. The blot was probed sequentially for c- jun and 28S rRNA transcripts.

    Article Snippet: To provide a normalization standard for RNA loading and transfer, blots were stripped and probed with a DNA oligomer corresponding to a human 28S rRNA sequence (BD Biosciences Clontech, Palo Alto, Calif.) end labeled with [γ-32 P]ATP (Amersham).

    Techniques: Autoradiography, Northern Blot, Knock-In, Incubation

    (A) Hypoxia Hx) induces the immediate-early accumulation of c- jun mRNA independently of the presence of HIF-1α. Autoradiograph of a Northern blot of total RNA from wild-type and HIF-1α null mEFs incubated under aerobic conditions (air [5% CO 2 ]) or exposed to hypoxia (pO 2 ≤ 1%) for the indicated times. The blot was probed sequentially for c- jun and 28S rRNA transcripts. This finding is representative of three independent experiments. (B) The early hypoxia-inducible accumulation of c- jun mRNA is dependent on serum (10% FBS). Histogram of relative amounts of c- jun mRNA detected by quantitative RT-PCR amplification of total RNA harvested at the indicated times. For details, see Materials and Methods.

    Journal: Molecular and Cellular Biology

    Article Title: The Response of c-Jun/AP-1 to Chronic Hypoxia Is Hypoxia-Inducible Factor 1? Dependent

    doi: 10.1128/MCB.22.8.2515-2523.2002

    Figure Lengend Snippet: (A) Hypoxia Hx) induces the immediate-early accumulation of c- jun mRNA independently of the presence of HIF-1α. Autoradiograph of a Northern blot of total RNA from wild-type and HIF-1α null mEFs incubated under aerobic conditions (air [5% CO 2 ]) or exposed to hypoxia (pO 2 ≤ 1%) for the indicated times. The blot was probed sequentially for c- jun and 28S rRNA transcripts. This finding is representative of three independent experiments. (B) The early hypoxia-inducible accumulation of c- jun mRNA is dependent on serum (10% FBS). Histogram of relative amounts of c- jun mRNA detected by quantitative RT-PCR amplification of total RNA harvested at the indicated times. For details, see Materials and Methods.

    Article Snippet: To provide a normalization standard for RNA loading and transfer, blots were stripped and probed with a DNA oligomer corresponding to a human 28S rRNA sequence (BD Biosciences Clontech, Palo Alto, Calif.) end labeled with [γ-32 P]ATP (Amersham).

    Techniques: Autoradiography, Northern Blot, Incubation, Quantitative RT-PCR, Amplification

    The delayed accumulation of c- jun mRNA induced by prolonged hypoxia (Hx) is dependent on the presence of HIF-1α. Autoradiograph of a Northern blot of total RNA from wild-type and HIF-1α null mEFs incubated under aerobic conditions (air [5% CO 2 ]) or exposed to hypoxia (pO 2 ≤ 0.01%) for the indicated times. The blot was probed sequentially for the mRNAs for c- jun and 28S rRNA. This finding is representative of three independent experiments.

    Journal: Molecular and Cellular Biology

    Article Title: The Response of c-Jun/AP-1 to Chronic Hypoxia Is Hypoxia-Inducible Factor 1? Dependent

    doi: 10.1128/MCB.22.8.2515-2523.2002

    Figure Lengend Snippet: The delayed accumulation of c- jun mRNA induced by prolonged hypoxia (Hx) is dependent on the presence of HIF-1α. Autoradiograph of a Northern blot of total RNA from wild-type and HIF-1α null mEFs incubated under aerobic conditions (air [5% CO 2 ]) or exposed to hypoxia (pO 2 ≤ 0.01%) for the indicated times. The blot was probed sequentially for the mRNAs for c- jun and 28S rRNA. This finding is representative of three independent experiments.

    Article Snippet: To provide a normalization standard for RNA loading and transfer, blots were stripped and probed with a DNA oligomer corresponding to a human 28S rRNA sequence (BD Biosciences Clontech, Palo Alto, Calif.) end labeled with [γ-32 P]ATP (Amersham).

    Techniques: Autoradiography, Northern Blot, Incubation