Structured Review

Roche plasma hiv 1 rna levels
Profile of the study cohort. ART = Antiretroviral therapy; AZT = Zidovudine; D4T = Stavudine; EFV = efavirenz; NVP = Nevirapine; OBRs = optimized background regimens; TDF = tenofovir; 3TC = lamivudine. a Random sampling was performed using random numbers. b <t>HIV-1-infected</t> patients who simultaneously discontinued all drugs in NVP-based regimens when they experienced allergic reactions to NVP-based regimens. c HIV-1-infected patients who discontinued NVP first but continued use of the other NRTIs for a few days when they experienced allergic reactions to NVP-based regimens. d This criterion included patients who changed EFV to other NNRTIs or PIs. It did not include patients who only changed or modify NRTIs. e Virological failure was defined as either (1) having two consecutive results of plasma HIV-1 <t>RNA</t> > 400 copies/ml after 6 month of a EFV-based regimen or (2) having plasma HIV-1 RNA > 1,000 copies/ml plus having any genotypic resistance mutation to efavirenz-based regimen. f It included either (1) patients who did not have clinical visits for more than 3 months from appointment date or (2) patients who stopped efavirenz-based regimen for more than 3 months. g The cohort ended on 31 March 2010.
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1) Product Images from "Virological failure of staggered and simultaneous treatment interruption in HIV patients who began Efavirenz-based regimens after allergic reactions to nevirapine"

Article Title: Virological failure of staggered and simultaneous treatment interruption in HIV patients who began Efavirenz-based regimens after allergic reactions to nevirapine

Journal: AIDS Research and Therapy

doi: 10.1186/1742-6405-10-4

Profile of the study cohort. ART = Antiretroviral therapy; AZT = Zidovudine; D4T = Stavudine; EFV = efavirenz; NVP = Nevirapine; OBRs = optimized background regimens; TDF = tenofovir; 3TC = lamivudine. a Random sampling was performed using random numbers. b HIV-1-infected patients who simultaneously discontinued all drugs in NVP-based regimens when they experienced allergic reactions to NVP-based regimens. c HIV-1-infected patients who discontinued NVP first but continued use of the other NRTIs for a few days when they experienced allergic reactions to NVP-based regimens. d This criterion included patients who changed EFV to other NNRTIs or PIs. It did not include patients who only changed or modify NRTIs. e Virological failure was defined as either (1) having two consecutive results of plasma HIV-1 RNA > 400 copies/ml after 6 month of a EFV-based regimen or (2) having plasma HIV-1 RNA > 1,000 copies/ml plus having any genotypic resistance mutation to efavirenz-based regimen. f It included either (1) patients who did not have clinical visits for more than 3 months from appointment date or (2) patients who stopped efavirenz-based regimen for more than 3 months. g The cohort ended on 31 March 2010.
Figure Legend Snippet: Profile of the study cohort. ART = Antiretroviral therapy; AZT = Zidovudine; D4T = Stavudine; EFV = efavirenz; NVP = Nevirapine; OBRs = optimized background regimens; TDF = tenofovir; 3TC = lamivudine. a Random sampling was performed using random numbers. b HIV-1-infected patients who simultaneously discontinued all drugs in NVP-based regimens when they experienced allergic reactions to NVP-based regimens. c HIV-1-infected patients who discontinued NVP first but continued use of the other NRTIs for a few days when they experienced allergic reactions to NVP-based regimens. d This criterion included patients who changed EFV to other NNRTIs or PIs. It did not include patients who only changed or modify NRTIs. e Virological failure was defined as either (1) having two consecutive results of plasma HIV-1 RNA > 400 copies/ml after 6 month of a EFV-based regimen or (2) having plasma HIV-1 RNA > 1,000 copies/ml plus having any genotypic resistance mutation to efavirenz-based regimen. f It included either (1) patients who did not have clinical visits for more than 3 months from appointment date or (2) patients who stopped efavirenz-based regimen for more than 3 months. g The cohort ended on 31 March 2010.

Techniques Used: Sampling, Infection, Mutagenesis

2) Product Images from "Virological failure of staggered and simultaneous treatment interruption in HIV patients who began Efavirenz-based regimens after allergic reactions to nevirapine"

Article Title: Virological failure of staggered and simultaneous treatment interruption in HIV patients who began Efavirenz-based regimens after allergic reactions to nevirapine

Journal: AIDS Research and Therapy

doi: 10.1186/1742-6405-10-4

Profile of the study cohort. ART = Antiretroviral therapy; AZT = Zidovudine; D4T = Stavudine; EFV = efavirenz; NVP = Nevirapine; OBRs = optimized background regimens; TDF = tenofovir; 3TC = lamivudine. a Random sampling was performed using random numbers. b HIV-1-infected patients who simultaneously discontinued all drugs in NVP-based regimens when they experienced allergic reactions to NVP-based regimens. c HIV-1-infected patients who discontinued NVP first but continued use of the other NRTIs for a few days when they experienced allergic reactions to NVP-based regimens. d This criterion included patients who changed EFV to other NNRTIs or PIs. It did not include patients who only changed or modify NRTIs. e Virological failure was defined as either (1) having two consecutive results of plasma HIV-1 RNA > 400 copies/ml after 6 month of a EFV-based regimen or (2) having plasma HIV-1 RNA > 1,000 copies/ml plus having any genotypic resistance mutation to efavirenz-based regimen. f It included either (1) patients who did not have clinical visits for more than 3 months from appointment date or (2) patients who stopped efavirenz-based regimen for more than 3 months. g The cohort ended on 31 March 2010.
Figure Legend Snippet: Profile of the study cohort. ART = Antiretroviral therapy; AZT = Zidovudine; D4T = Stavudine; EFV = efavirenz; NVP = Nevirapine; OBRs = optimized background regimens; TDF = tenofovir; 3TC = lamivudine. a Random sampling was performed using random numbers. b HIV-1-infected patients who simultaneously discontinued all drugs in NVP-based regimens when they experienced allergic reactions to NVP-based regimens. c HIV-1-infected patients who discontinued NVP first but continued use of the other NRTIs for a few days when they experienced allergic reactions to NVP-based regimens. d This criterion included patients who changed EFV to other NNRTIs or PIs. It did not include patients who only changed or modify NRTIs. e Virological failure was defined as either (1) having two consecutive results of plasma HIV-1 RNA > 400 copies/ml after 6 month of a EFV-based regimen or (2) having plasma HIV-1 RNA > 1,000 copies/ml plus having any genotypic resistance mutation to efavirenz-based regimen. f It included either (1) patients who did not have clinical visits for more than 3 months from appointment date or (2) patients who stopped efavirenz-based regimen for more than 3 months. g The cohort ended on 31 March 2010.

Techniques Used: Sampling, Infection, Mutagenesis

3) Product Images from "Agonist-activated Ca2+ influx occurs at stable plasma membrane and endoplasmic reticulum junctions"

Article Title: Agonist-activated Ca2+ influx occurs at stable plasma membrane and endoplasmic reticulum junctions

Journal: Journal of Cell Science

doi: 10.1242/jcs.068387

Characteristics of HEK junctate and HEKT3 junctate overexpressing clones. Total RNA was extracted from HEK T3 and HEK T3-junctate clone D6 cells ( A ) or from control HEK293 and HEK293 junctate–YFP clone 21 cells ( B ) and converted into cDNA and
Figure Legend Snippet: Characteristics of HEK junctate and HEKT3 junctate overexpressing clones. Total RNA was extracted from HEK T3 and HEK T3-junctate clone D6 cells ( A ) or from control HEK293 and HEK293 junctate–YFP clone 21 cells ( B ) and converted into cDNA and

Techniques Used: Clone Assay

4) Product Images from "CP and CP-PGN protect mice against MRSA infection by inducing M1 macrophages"

Article Title: CP and CP-PGN protect mice against MRSA infection by inducing M1 macrophages

Journal: Scientific Reports

doi: 10.1038/s41598-017-17001-0

CP and CP-PGN skewed macrophages to a M1-like type. RAW264.7 cells ( a,d ) and peritoneal macrophages ( b,e ) were co-cultured with CP or CP-PGN ex vivo to detect the iNOS and arginase-1 by qRT-PCR and CD86 by Flow cytometric analysis. Macrophages pretreated with CP or CP-PGN for 24 h in vivo ( c,f ) were collected from the abdominal cavity of mice to analyzed iNOS, arginase-1 and CD86. Data shown are representative of three independent experiments. The significant differences compared with the control group were analyzed by Student’s t test, * P
Figure Legend Snippet: CP and CP-PGN skewed macrophages to a M1-like type. RAW264.7 cells ( a,d ) and peritoneal macrophages ( b,e ) were co-cultured with CP or CP-PGN ex vivo to detect the iNOS and arginase-1 by qRT-PCR and CD86 by Flow cytometric analysis. Macrophages pretreated with CP or CP-PGN for 24 h in vivo ( c,f ) were collected from the abdominal cavity of mice to analyzed iNOS, arginase-1 and CD86. Data shown are representative of three independent experiments. The significant differences compared with the control group were analyzed by Student’s t test, * P

Techniques Used: Cell Culture, Ex Vivo, Quantitative RT-PCR, Flow Cytometry, In Vivo, Mouse Assay

Silencing of TLR2 down-regulated CP/CP-PGN-induced activation and differentiation of RAW264.7 cells. The expression levels of TLRs were analyzed by qRT-PCR to select out which TLR changed most obviously ( a ). RAW264.7 cells transfected 24 h with TLR2 siRNA/control siRNA, then the mRNA of TLR2 were assayed by qRT-PCR ( b ). CP and CP-PGN pretreated RAW264.7 or RAW264.7 transfected with TLR2 siRNA co-stimulated with MRSA to analysis the phagocytosis rate ( c ). RAW264.7 or RAW264.7 transfected with siTLR2 were co-cultured with CP or CP-PGN for 24 h to detect the production of NO ( d ), for 3 h to detect cytokines ( e,f,g ), iNOS ( h ) and arginase-1 ( i ) by qRT-PCR. CD86 were determined by flow cytometric analysis ( j ) and cell migration assay was shown ( k ). Each graph represents the average of three replications. The significant differences compared with the untreated group were analyzed by Student’s t test, * P
Figure Legend Snippet: Silencing of TLR2 down-regulated CP/CP-PGN-induced activation and differentiation of RAW264.7 cells. The expression levels of TLRs were analyzed by qRT-PCR to select out which TLR changed most obviously ( a ). RAW264.7 cells transfected 24 h with TLR2 siRNA/control siRNA, then the mRNA of TLR2 were assayed by qRT-PCR ( b ). CP and CP-PGN pretreated RAW264.7 or RAW264.7 transfected with TLR2 siRNA co-stimulated with MRSA to analysis the phagocytosis rate ( c ). RAW264.7 or RAW264.7 transfected with siTLR2 were co-cultured with CP or CP-PGN for 24 h to detect the production of NO ( d ), for 3 h to detect cytokines ( e,f,g ), iNOS ( h ) and arginase-1 ( i ) by qRT-PCR. CD86 were determined by flow cytometric analysis ( j ) and cell migration assay was shown ( k ). Each graph represents the average of three replications. The significant differences compared with the untreated group were analyzed by Student’s t test, * P

Techniques Used: Activation Assay, Expressing, Quantitative RT-PCR, Transfection, Cell Culture, Flow Cytometry, Cell Migration Assay

5) Product Images from "Virological failure of staggered and simultaneous treatment interruption in HIV patients who began Efavirenz-based regimens after allergic reactions to nevirapine"

Article Title: Virological failure of staggered and simultaneous treatment interruption in HIV patients who began Efavirenz-based regimens after allergic reactions to nevirapine

Journal: AIDS Research and Therapy

doi: 10.1186/1742-6405-10-4

Profile of the study cohort. ART = Antiretroviral therapy; AZT = Zidovudine; D4T = Stavudine; EFV = efavirenz; NVP = Nevirapine; OBRs = optimized background regimens; TDF = tenofovir; 3TC = lamivudine. a Random sampling was performed using random numbers. b HIV-1-infected patients who simultaneously discontinued all drugs in NVP-based regimens when they experienced allergic reactions to NVP-based regimens. c HIV-1-infected patients who discontinued NVP first but continued use of the other NRTIs for a few days when they experienced allergic reactions to NVP-based regimens. d This criterion included patients who changed EFV to other NNRTIs or PIs. It did not include patients who only changed or modify NRTIs. e Virological failure was defined as either (1) having two consecutive results of plasma HIV-1 RNA > 400 copies/ml after 6 month of a EFV-based regimen or (2) having plasma HIV-1 RNA > 1,000 copies/ml plus having any genotypic resistance mutation to efavirenz-based regimen. f It included either (1) patients who did not have clinical visits for more than 3 months from appointment date or (2) patients who stopped efavirenz-based regimen for more than 3 months. g The cohort ended on 31 March 2010.
Figure Legend Snippet: Profile of the study cohort. ART = Antiretroviral therapy; AZT = Zidovudine; D4T = Stavudine; EFV = efavirenz; NVP = Nevirapine; OBRs = optimized background regimens; TDF = tenofovir; 3TC = lamivudine. a Random sampling was performed using random numbers. b HIV-1-infected patients who simultaneously discontinued all drugs in NVP-based regimens when they experienced allergic reactions to NVP-based regimens. c HIV-1-infected patients who discontinued NVP first but continued use of the other NRTIs for a few days when they experienced allergic reactions to NVP-based regimens. d This criterion included patients who changed EFV to other NNRTIs or PIs. It did not include patients who only changed or modify NRTIs. e Virological failure was defined as either (1) having two consecutive results of plasma HIV-1 RNA > 400 copies/ml after 6 month of a EFV-based regimen or (2) having plasma HIV-1 RNA > 1,000 copies/ml plus having any genotypic resistance mutation to efavirenz-based regimen. f It included either (1) patients who did not have clinical visits for more than 3 months from appointment date or (2) patients who stopped efavirenz-based regimen for more than 3 months. g The cohort ended on 31 March 2010.

Techniques Used: Sampling, Infection, Mutagenesis

6) Product Images from "Comparison of Two Measures of Human Immunodeficiency Virus (HIV) Type 1 Load in HIV Risk Groups"

Article Title: Comparison of Two Measures of Human Immunodeficiency Virus (HIV) Type 1 Load in HIV Risk Groups

Journal: Journal of Clinical Microbiology

doi:

Estimated regression lines of log-based infectious viral load, measured as log IUPM, by log-based HIV RNA load, measured as log number of RNA copies per milliliter, for each population, excluding those whose estimated seroconversion date was within 6 months of the time that the viral load was measured.
Figure Legend Snippet: Estimated regression lines of log-based infectious viral load, measured as log IUPM, by log-based HIV RNA load, measured as log number of RNA copies per milliliter, for each population, excluding those whose estimated seroconversion date was within 6 months of the time that the viral load was measured.

Techniques Used:

Estimated regression lines of log-based HIV load by CD4 + lymphocyte cell count for each population, excluding those whose estimated seroconversion date was within 6 months of the time that the viral load was measured. The cell-associated infectious HIV-1 load (A) was measured as log IUPM, and the plasma HIV RNA concentration (B) was measured as the log number of RNA copies per milliliter.
Figure Legend Snippet: Estimated regression lines of log-based HIV load by CD4 + lymphocyte cell count for each population, excluding those whose estimated seroconversion date was within 6 months of the time that the viral load was measured. The cell-associated infectious HIV-1 load (A) was measured as log IUPM, and the plasma HIV RNA concentration (B) was measured as the log number of RNA copies per milliliter.

Techniques Used: Cell Counting, Concentration Assay

Related Articles

Centrifugation:

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Article Snippet: Extraction of DNA and RNA from Cells DNA and RNA were extracted from transfected and non-transfected hepatoma cells in a Magnapure robot (Roche Applied Science, Germany) using the Total NA protocol. .. Harvested cells were washed in 1 mL PBS and after centrifugation at 5000 rpm for 3 min the pellet was re-suspended in 800 µL RLT lysis buffer (Qiagen Sciences, MD, USA) before extraction.

Amplification:

Article Title: Depletion of the cdk Inhibitor p16INK4a Differentially Affects Proliferation of Established Cervical Carcinoma Cells
Article Snippet: For cDNA synthesis, 1 μg of total RNA was reverse transcribed with the Transcriptor first-strand cDNA synthesis kit (Roche Applied Science) and diluted 1:4. .. Amplification of bicistronic HPV-16, -18, and -45 E6/E7 mRNAs from HeLa, Caski, and MS-751 cells was carried out with the following primers: HPV-16E6E7 forward, 5′-GTTACTGCGACGTGAGGTATATG-3′; HPV-16E6E7 reverse, 5′-CATTTATCACATACAGCTATGGATTC-3′; HPV-18E6E7 forward, 5′-CAGAGGTATTTGAATTTGCATTT-3′; HPV-18E6E7 reverse, 5′-AATCTATACATTTATGGCATGCAG-3′; HPV-45E7 forward, 5′-TTGCATTTGGAACCTCAGAA-3′; and HPV-45E7 reverse, 5′-TTTCCTCCTCTGACTCGCTT-3′.

Article Title: Peroxisome proliferator-activated receptor (PPAR)? expression in T cells mediates gender differences in development of T cell-mediated autoimmunity
Article Snippet: .. 2 μg RNA was reverse-transcribed, and PPARα, PPARδ, PPARγ, and β-actin cDNAs were amplified according to methods described previously ( ) using a Lightcycler (Roche). ..

Article Title: MERS-coronavirus replication induces severe in vitro cytopathology and is strongly inhibited by cyclosporin A or interferon-? treatment
Article Snippet: RNA from 200 µl culture medium of CoV-infected cells was isolated with a MagnaPure LC Total Nucleic Acid Isolation kit (Roche) and eluted in 100 µl. .. RT-PCR conditions for quantifying MERS-CoV and SARS-CoV RNA and amplification parameters have been described previously ( ; ).

Article Title: Arabidopsis ETO1 specifically interacts with and negatively regulates type 2 1-aminocyclopropane-1-carboxylate synthases
Article Snippet: RNA (1 μg) was used in each RT-PCR following the manufacturer's protocol (Titan One Tube RT-PCR System, Roche Diagnostics). .. PCR amplification was performed for a first round of 10 cycles as follows: denaturation at 94°C for 30 sec, annealing at 55°C for 30 sec, and extension at 68°C for 45 sec. Then the second round of cycles was performed as follows: denaturation at 94°C for 30 sec, annealing at 55°C for 30 sec, and extension at 68°C for 45 sec + 5/cycle sec.

Article Title: Genome-wide analyses reveal the IRE1a-XBP1 pathway promotes T helper cell differentiation by resolving secretory stress and accelerating proliferation
Article Snippet: Reverse transcription quantitative PCR (RT-qPCR) Total RNA was isolated from two million cells by SV total RNA isolation kit (Promega). cDNA was prepared by annealing 500 ng RNA with oligo dT as per the manufacturer’s instructions (Transcriptor High Fidelity cDNA Synthesis kit, Roche). .. For XBP1, mRNA was amplified by PCR and products were separated by electrophoresis through a 2.5% agarose gel and visualized by ethidium bromide staining.

Mass Spectrometry:

Article Title: Depletion of the cdk Inhibitor p16INK4a Differentially Affects Proliferation of Established Cervical Carcinoma Cells
Article Snippet: For cDNA synthesis, 1 μg of total RNA was reverse transcribed with the Transcriptor first-strand cDNA synthesis kit (Roche Applied Science) and diluted 1:4. .. Amplification of bicistronic HPV-16, -18, and -45 E6/E7 mRNAs from HeLa, Caski, and MS-751 cells was carried out with the following primers: HPV-16E6E7 forward, 5′-GTTACTGCGACGTGAGGTATATG-3′; HPV-16E6E7 reverse, 5′-CATTTATCACATACAGCTATGGATTC-3′; HPV-18E6E7 forward, 5′-CAGAGGTATTTGAATTTGCATTT-3′; HPV-18E6E7 reverse, 5′-AATCTATACATTTATGGCATGCAG-3′; HPV-45E7 forward, 5′-TTGCATTTGGAACCTCAGAA-3′; and HPV-45E7 reverse, 5′-TTTCCTCCTCTGACTCGCTT-3′.

Synthesized:

Article Title: Chronic hypoxia‐induced slug promotes invasive behavior of prostate cancer cells by activating expression of ephrin‐B1, et al. Chronic hypoxia‐induced slug promotes invasive behavior of prostate cancer cells by activating expression of ephrin‐B1
Article Snippet: .. 2.5 Real‐time quantitative RT‐PCR First‐strand cDNA was synthesized from the total RNA using ThermoScript RT‐PCR System (Roche, Indianapolis, IN, USA). .. PCR was performed on a LightCycler system (Roche) using LightCycler FastStart DNA Master SYBR Green I reaction mix (Roche) and QuantiTect Primer Assays (QIAGEN, Hilden, Germany).

Article Title: Ddx42p--a human DEAD box protein with RNA chaperone activities
Article Snippet: A ddx42-specific antisense RNA probe (complementary to nt 1797–3074 of the ddx42 cDNA) was synthesized by NheI-digest of pGEMddx42 (see below) and in vitro transcription with T7 RNA polymerase in the presence of Dig-RNA labeling mix (Roche). .. Northern blotting of 100 ng poly(A)+ RNA was carried out using the Dig-labeled ddx42-specific antisense RNA probe and detected with CSPD (Roche).

Quantitative RT-PCR:

Article Title: Prospective Evaluation of Whole Genome MicroRNA Expression Profiling in Childhood Acute Lymphoblastic Leukemia
Article Snippet: Quantitative Real Time Polymerase Chain Reaction and Validation Significantly dysregulated miRNAs were validated by Quantitative PCR (LightCycler 480, Roche Applied Science, Mannheim, Germany) after comparing the BM microarray miRNA expressions of patients with the controls. .. A master mix was designed for each primer set in accordance with the recommendations of the real time RT-PCR setup for “individual assays,” suggested in the kit.

Article Title: Chronic hypoxia‐induced slug promotes invasive behavior of prostate cancer cells by activating expression of ephrin‐B1, et al. Chronic hypoxia‐induced slug promotes invasive behavior of prostate cancer cells by activating expression of ephrin‐B1
Article Snippet: .. 2.5 Real‐time quantitative RT‐PCR First‐strand cDNA was synthesized from the total RNA using ThermoScript RT‐PCR System (Roche, Indianapolis, IN, USA). .. PCR was performed on a LightCycler system (Roche) using LightCycler FastStart DNA Master SYBR Green I reaction mix (Roche) and QuantiTect Primer Assays (QIAGEN, Hilden, Germany).

Article Title: Depletion of the cdk Inhibitor p16INK4a Differentially Affects Proliferation of Established Cervical Carcinoma Cells
Article Snippet: Paragraph title: Quantitative real-time PCR (qRT-PCR) analysis of mRNAs. ... For cDNA synthesis, 1 μg of total RNA was reverse transcribed with the Transcriptor first-strand cDNA synthesis kit (Roche Applied Science) and diluted 1:4.

Article Title: Effects of GADL1 overexpression on cell migration and the associated morphological changes
Article Snippet: Paragraph title: Extraction of mRNA from cells and real-time quantitative PCR (RT-qPCR) ... The extracted RNA was reverse transcribed into cDNA using a reverse transcription kit (Roche, Switzerland).

Article Title: Peroxisome proliferator-activated receptor (PPAR)? expression in T cells mediates gender differences in development of T cell-mediated autoimmunity
Article Snippet: Paragraph title: Real-time RT-PCR. ... 2 μg RNA was reverse-transcribed, and PPARα, PPARδ, PPARγ, and β-actin cDNAs were amplified according to methods described previously ( ) using a Lightcycler (Roche).

Article Title: HIV-1 RNA Levels and Antiretroviral Drug Resistance in Blood and Non-Blood Compartments from HIV-1-Infected Men and Women enrolled in AIDS Clinical Trials Group Study A5077
Article Snippet: Laboratory Testing HIV RNA was measured in blood plasma using RT-PCR (Monitor version 1.5 (Roche Molecular) at a central laboratory. .. HIV RNA was extracted into NucliSens extraction buffer from saliva, seminal plasma or endocervical wicks and quantified using published NucliSens (bioMerieux; saliva, seminal plasma) or real-time RT-PCR assay methods , , , .

Article Title: Homologous Recombination Occurs in Entamoeba and Is Enhanced during Growth Stress and Stage Conversion
Article Snippet: .. RNA purification and Real Time RT-PCR Entamoeba cells were harvested and suspended immediately in TriZol reagent (Invitrogen) and RNA was isolated and treated with DNase I (Roche) according to manufacturer's protocol. .. RNA was reverse transcribed into cDNA using Superscript III reverse transcriptase (Invitrogen) and random hexamers used as a primer.

Article Title: Genome-wide analyses reveal the IRE1a-XBP1 pathway promotes T helper cell differentiation by resolving secretory stress and accelerating proliferation
Article Snippet: .. Reverse transcription quantitative PCR (RT-qPCR) Total RNA was isolated from two million cells by SV total RNA isolation kit (Promega). cDNA was prepared by annealing 500 ng RNA with oligo dT as per the manufacturer’s instructions (Transcriptor High Fidelity cDNA Synthesis kit, Roche). ..

Real-time Polymerase Chain Reaction:

Article Title: Prospective Evaluation of Whole Genome MicroRNA Expression Profiling in Childhood Acute Lymphoblastic Leukemia
Article Snippet: .. Quantitative Real Time Polymerase Chain Reaction and Validation Significantly dysregulated miRNAs were validated by Quantitative PCR (LightCycler 480, Roche Applied Science, Mannheim, Germany) after comparing the BM microarray miRNA expressions of patients with the controls. .. RNA was reverse-transcribed to cDNA by using a cDNA synthesis kit (Exiqon).

Article Title: Hepatitis B Viral DNA Decline at Loss of HBeAg Is Mainly Explained by Reduced cccDNA Load - Down-Regulated Transcription of PgRNA Has Limited Impact
Article Snippet: Extraction of DNA and RNA from Cells DNA and RNA were extracted from transfected and non-transfected hepatoma cells in a Magnapure robot (Roche Applied Science, Germany) using the Total NA protocol. .. Half the output from cells were subjected to quantification of DNA by real-time PCR, whereas the other half was treated with DNase (Turbo DNA-free kit, Ambion Inc, TX, USA) before analysis with reverse transcriptase (RT) real-time PCR.

Article Title: Depletion of the cdk Inhibitor p16INK4a Differentially Affects Proliferation of Established Cervical Carcinoma Cells
Article Snippet: Paragraph title: Quantitative real-time PCR (qRT-PCR) analysis of mRNAs. ... For cDNA synthesis, 1 μg of total RNA was reverse transcribed with the Transcriptor first-strand cDNA synthesis kit (Roche Applied Science) and diluted 1:4.

Article Title: Effects of GADL1 overexpression on cell migration and the associated morphological changes
Article Snippet: Paragraph title: Extraction of mRNA from cells and real-time quantitative PCR (RT-qPCR) ... The extracted RNA was reverse transcribed into cDNA using a reverse transcription kit (Roche, Switzerland).

Article Title: Homologous Recombination Occurs in Entamoeba and Is Enhanced during Growth Stress and Stage Conversion
Article Snippet: RNA purification and Real Time RT-PCR Entamoeba cells were harvested and suspended immediately in TriZol reagent (Invitrogen) and RNA was isolated and treated with DNase I (Roche) according to manufacturer's protocol. .. This was carried out using an ABI real-time PCR system 7300 (Applied Biosystems) with Power SYBR Green PCR Master Mix.

Article Title: Genome-wide analyses reveal the IRE1a-XBP1 pathway promotes T helper cell differentiation by resolving secretory stress and accelerating proliferation
Article Snippet: .. Reverse transcription quantitative PCR (RT-qPCR) Total RNA was isolated from two million cells by SV total RNA isolation kit (Promega). cDNA was prepared by annealing 500 ng RNA with oligo dT as per the manufacturer’s instructions (Transcriptor High Fidelity cDNA Synthesis kit, Roche). ..

Microarray:

Article Title: Prospective Evaluation of Whole Genome MicroRNA Expression Profiling in Childhood Acute Lymphoblastic Leukemia
Article Snippet: .. Quantitative Real Time Polymerase Chain Reaction and Validation Significantly dysregulated miRNAs were validated by Quantitative PCR (LightCycler 480, Roche Applied Science, Mannheim, Germany) after comparing the BM microarray miRNA expressions of patients with the controls. .. RNA was reverse-transcribed to cDNA by using a cDNA synthesis kit (Exiqon).

RNA Detection:

Article Title: HIV-1 RNA Levels and Antiretroviral Drug Resistance in Blood and Non-Blood Compartments from HIV-1-Infected Men and Women enrolled in AIDS Clinical Trials Group Study A5077
Article Snippet: Laboratory Testing HIV RNA was measured in blood plasma using RT-PCR (Monitor version 1.5 (Roche Molecular) at a central laboratory. .. The lower limits of viral RNA detection were 50 copies/mL in blood plasma, 400 copies/mL in saliva, 400 copies/mL in semen plasma, 1,500 copies/mL in cervical Sno-strip fluid, and 75 copies/mL in CVL fluid.

Expressing:

Article Title: Homologous Recombination Occurs in Entamoeba and Is Enhanced during Growth Stress and Stage Conversion
Article Snippet: RNA purification and Real Time RT-PCR Entamoeba cells were harvested and suspended immediately in TriZol reagent (Invitrogen) and RNA was isolated and treated with DNase I (Roche) according to manufacturer's protocol. .. Real-time RT-PCR was used to quantify HR and meiotic specific mRNA expression of E. invadens and E. histolytica during encystation and serum starvation respectively.

Transfection:

Article Title: Hepatitis B Viral DNA Decline at Loss of HBeAg Is Mainly Explained by Reduced cccDNA Load - Down-Regulated Transcription of PgRNA Has Limited Impact
Article Snippet: .. Extraction of DNA and RNA from Cells DNA and RNA were extracted from transfected and non-transfected hepatoma cells in a Magnapure robot (Roche Applied Science, Germany) using the Total NA protocol. .. Harvested cells were washed in 1 mL PBS and after centrifugation at 5000 rpm for 3 min the pellet was re-suspended in 800 µL RLT lysis buffer (Qiagen Sciences, MD, USA) before extraction.

SYBR Green Assay:

Article Title: Chronic hypoxia‐induced slug promotes invasive behavior of prostate cancer cells by activating expression of ephrin‐B1, et al. Chronic hypoxia‐induced slug promotes invasive behavior of prostate cancer cells by activating expression of ephrin‐B1
Article Snippet: 2.5 Real‐time quantitative RT‐PCR First‐strand cDNA was synthesized from the total RNA using ThermoScript RT‐PCR System (Roche, Indianapolis, IN, USA). .. PCR was performed on a LightCycler system (Roche) using LightCycler FastStart DNA Master SYBR Green I reaction mix (Roche) and QuantiTect Primer Assays (QIAGEN, Hilden, Germany).

Article Title: Effects of GADL1 overexpression on cell migration and the associated morphological changes
Article Snippet: The extracted RNA was reverse transcribed into cDNA using a reverse transcription kit (Roche, Switzerland). .. Expressions of GADL1 (PPH22451A), FN1 (PPH00143B), ITGA2 (PPH00625F), ITGAV (PPH00628C), and CCL2 (PPH00192F) were examined with SYBR Green (Qiagen, Germany) using gene-specific primers (all designed by Qiagen) in triplicates.

Article Title: Homologous Recombination Occurs in Entamoeba and Is Enhanced during Growth Stress and Stage Conversion
Article Snippet: RNA purification and Real Time RT-PCR Entamoeba cells were harvested and suspended immediately in TriZol reagent (Invitrogen) and RNA was isolated and treated with DNase I (Roche) according to manufacturer's protocol. .. This was carried out using an ABI real-time PCR system 7300 (Applied Biosystems) with Power SYBR Green PCR Master Mix.

Article Title: Genome-wide analyses reveal the IRE1a-XBP1 pathway promotes T helper cell differentiation by resolving secretory stress and accelerating proliferation
Article Snippet: Reverse transcription quantitative PCR (RT-qPCR) Total RNA was isolated from two million cells by SV total RNA isolation kit (Promega). cDNA was prepared by annealing 500 ng RNA with oligo dT as per the manufacturer’s instructions (Transcriptor High Fidelity cDNA Synthesis kit, Roche). .. Two microliters of cDNA was used in 12 μl qPCR reactions with appropriate primers and SYBR Green PCR Master Mix (Applied Biosystems).

Northern Blot:

Article Title: Ddx42p--a human DEAD box protein with RNA chaperone activities
Article Snippet: .. Northern blotting of 100 ng poly(A)+ RNA was carried out using the Dig-labeled ddx42-specific antisense RNA probe and detected with CSPD (Roche). .. A Dig-labeled β-actin RNA probe (Roche) was used to detect the ubiquitously abundant β-actin mRNA as a loading control.

Article Title: Arabidopsis ETO1 specifically interacts with and negatively regulates type 2 1-aminocyclopropane-1-carboxylate synthases
Article Snippet: RNA (1 μg) was used in each RT-PCR following the manufacturer's protocol (Titan One Tube RT-PCR System, Roche Diagnostics). .. For Northern blot analysis, total RNA (10 μg) was used for each lanes and blotted onto positively charged nylon membranes (Roche Diagnostics).

Polymerase Chain Reaction:

Article Title: Prospective Evaluation of Whole Genome MicroRNA Expression Profiling in Childhood Acute Lymphoblastic Leukemia
Article Snippet: Quantitative Real Time Polymerase Chain Reaction and Validation Significantly dysregulated miRNAs were validated by Quantitative PCR (LightCycler 480, Roche Applied Science, Mannheim, Germany) after comparing the BM microarray miRNA expressions of patients with the controls. .. For miRNA quantification, the miRCURY LNA Universal RT microRNA PCR system (Exiqon) was used in combination with the predesigned primers (Exiqon).

Article Title: Chronic hypoxia‐induced slug promotes invasive behavior of prostate cancer cells by activating expression of ephrin‐B1, et al. Chronic hypoxia‐induced slug promotes invasive behavior of prostate cancer cells by activating expression of ephrin‐B1
Article Snippet: 2.5 Real‐time quantitative RT‐PCR First‐strand cDNA was synthesized from the total RNA using ThermoScript RT‐PCR System (Roche, Indianapolis, IN, USA). .. PCR was performed on a LightCycler system (Roche) using LightCycler FastStart DNA Master SYBR Green I reaction mix (Roche) and QuantiTect Primer Assays (QIAGEN, Hilden, Germany).

Article Title: Arabidopsis ETO1 specifically interacts with and negatively regulates type 2 1-aminocyclopropane-1-carboxylate synthases
Article Snippet: RNA (1 μg) was used in each RT-PCR following the manufacturer's protocol (Titan One Tube RT-PCR System, Roche Diagnostics). .. PCR amplification was performed for a first round of 10 cycles as follows: denaturation at 94°C for 30 sec, annealing at 55°C for 30 sec, and extension at 68°C for 45 sec. Then the second round of cycles was performed as follows: denaturation at 94°C for 30 sec, annealing at 55°C for 30 sec, and extension at 68°C for 45 sec + 5/cycle sec.

Article Title: Homologous Recombination Occurs in Entamoeba and Is Enhanced during Growth Stress and Stage Conversion
Article Snippet: RNA purification and Real Time RT-PCR Entamoeba cells were harvested and suspended immediately in TriZol reagent (Invitrogen) and RNA was isolated and treated with DNase I (Roche) according to manufacturer's protocol. .. This was carried out using an ABI real-time PCR system 7300 (Applied Biosystems) with Power SYBR Green PCR Master Mix.

Article Title: Genome-wide analyses reveal the IRE1a-XBP1 pathway promotes T helper cell differentiation by resolving secretory stress and accelerating proliferation
Article Snippet: Reverse transcription quantitative PCR (RT-qPCR) Total RNA was isolated from two million cells by SV total RNA isolation kit (Promega). cDNA was prepared by annealing 500 ng RNA with oligo dT as per the manufacturer’s instructions (Transcriptor High Fidelity cDNA Synthesis kit, Roche). .. Two microliters of cDNA was used in 12 μl qPCR reactions with appropriate primers and SYBR Green PCR Master Mix (Applied Biosystems).

Isolation:

Article Title: Depletion of the cdk Inhibitor p16INK4a Differentially Affects Proliferation of Established Cervical Carcinoma Cells
Article Snippet: RNA was isolated using the RNeasy Minikit (Qiagen) and quantified as described above. .. For cDNA synthesis, 1 μg of total RNA was reverse transcribed with the Transcriptor first-strand cDNA synthesis kit (Roche Applied Science) and diluted 1:4.

Article Title: Effects of GADL1 overexpression on cell migration and the associated morphological changes
Article Snippet: Extraction of mRNA from cells and real-time quantitative PCR (RT-qPCR) Total RNA was extracted SH-SY5Y or GADL1 -overexpressing cells pooling from sextuplicate wells using the NucleoSpin RNA/protein isolation kit (MACHEREY-NAGEL, Germany). .. The extracted RNA was reverse transcribed into cDNA using a reverse transcription kit (Roche, Switzerland).

Article Title: Peroxisome proliferator-activated receptor (PPAR)? expression in T cells mediates gender differences in development of T cell-mediated autoimmunity
Article Snippet: Total RNA was isolated from these T cells using the Absolutely RNA RT-PCR Miniprep kit (Statagene). .. 2 μg RNA was reverse-transcribed, and PPARα, PPARδ, PPARγ, and β-actin cDNAs were amplified according to methods described previously ( ) using a Lightcycler (Roche).

Article Title: MERS-coronavirus replication induces severe in vitro cytopathology and is strongly inhibited by cyclosporin A or interferon-? treatment
Article Snippet: .. RNA from 200 µl culture medium of CoV-infected cells was isolated with a MagnaPure LC Total Nucleic Acid Isolation kit (Roche) and eluted in 100 µl. .. RT-PCR conditions for quantifying MERS-CoV and SARS-CoV RNA and amplification parameters have been described previously ( ; ).

Article Title: Ddx42p--a human DEAD box protein with RNA chaperone activities
Article Snippet: Northern blotting Poly(A)+ RNA was isolated from different mammalian cell lines (∼107 cells each) using the Oligotex Direct mRNA kit (Qiagen) according to supplier's recommendations. .. Northern blotting of 100 ng poly(A)+ RNA was carried out using the Dig-labeled ddx42-specific antisense RNA probe and detected with CSPD (Roche).

Article Title: Arabidopsis ETO1 specifically interacts with and negatively regulates type 2 1-aminocyclopropane-1-carboxylate synthases
Article Snippet: Nucleic acid analysis Genomic DNA was isolated using Isoplant II (Nippon Gene). .. RNA (1 μg) was used in each RT-PCR following the manufacturer's protocol (Titan One Tube RT-PCR System, Roche Diagnostics).

Article Title: Homologous Recombination Occurs in Entamoeba and Is Enhanced during Growth Stress and Stage Conversion
Article Snippet: .. RNA purification and Real Time RT-PCR Entamoeba cells were harvested and suspended immediately in TriZol reagent (Invitrogen) and RNA was isolated and treated with DNase I (Roche) according to manufacturer's protocol. .. RNA was reverse transcribed into cDNA using Superscript III reverse transcriptase (Invitrogen) and random hexamers used as a primer.

Article Title: Genome-wide analyses reveal the IRE1a-XBP1 pathway promotes T helper cell differentiation by resolving secretory stress and accelerating proliferation
Article Snippet: .. Reverse transcription quantitative PCR (RT-qPCR) Total RNA was isolated from two million cells by SV total RNA isolation kit (Promega). cDNA was prepared by annealing 500 ng RNA with oligo dT as per the manufacturer’s instructions (Transcriptor High Fidelity cDNA Synthesis kit, Roche). ..

Size-exclusion Chromatography:

Article Title: Arabidopsis ETO1 specifically interacts with and negatively regulates type 2 1-aminocyclopropane-1-carboxylate synthases
Article Snippet: RNA (1 μg) was used in each RT-PCR following the manufacturer's protocol (Titan One Tube RT-PCR System, Roche Diagnostics). .. PCR amplification was performed for a first round of 10 cycles as follows: denaturation at 94°C for 30 sec, annealing at 55°C for 30 sec, and extension at 68°C for 45 sec. Then the second round of cycles was performed as follows: denaturation at 94°C for 30 sec, annealing at 55°C for 30 sec, and extension at 68°C for 45 sec + 5/cycle sec.

Labeling:

Article Title: Ddx42p--a human DEAD box protein with RNA chaperone activities
Article Snippet: A ddx42-specific antisense RNA probe (complementary to nt 1797–3074 of the ddx42 cDNA) was synthesized by NheI-digest of pGEMddx42 (see below) and in vitro transcription with T7 RNA polymerase in the presence of Dig-RNA labeling mix (Roche). .. Northern blotting of 100 ng poly(A)+ RNA was carried out using the Dig-labeled ddx42-specific antisense RNA probe and detected with CSPD (Roche).

Purification:

Article Title: Peroxisome proliferator-activated receptor (PPAR)? expression in T cells mediates gender differences in development of T cell-mediated autoimmunity
Article Snippet: CD4+ and CD3+ T cells were purified by negative selection (columns from R & D Systems). .. 2 μg RNA was reverse-transcribed, and PPARα, PPARδ, PPARγ, and β-actin cDNAs were amplified according to methods described previously ( ) using a Lightcycler (Roche).

Article Title: Homologous Recombination Occurs in Entamoeba and Is Enhanced during Growth Stress and Stage Conversion
Article Snippet: .. RNA purification and Real Time RT-PCR Entamoeba cells were harvested and suspended immediately in TriZol reagent (Invitrogen) and RNA was isolated and treated with DNase I (Roche) according to manufacturer's protocol. .. RNA was reverse transcribed into cDNA using Superscript III reverse transcriptase (Invitrogen) and random hexamers used as a primer.

Reverse Transcription Polymerase Chain Reaction:

Article Title: Chronic hypoxia‐induced slug promotes invasive behavior of prostate cancer cells by activating expression of ephrin‐B1, et al. Chronic hypoxia‐induced slug promotes invasive behavior of prostate cancer cells by activating expression of ephrin‐B1
Article Snippet: .. 2.5 Real‐time quantitative RT‐PCR First‐strand cDNA was synthesized from the total RNA using ThermoScript RT‐PCR System (Roche, Indianapolis, IN, USA). .. PCR was performed on a LightCycler system (Roche) using LightCycler FastStart DNA Master SYBR Green I reaction mix (Roche) and QuantiTect Primer Assays (QIAGEN, Hilden, Germany).

Article Title: Peroxisome proliferator-activated receptor (PPAR)? expression in T cells mediates gender differences in development of T cell-mediated autoimmunity
Article Snippet: Total RNA was isolated from these T cells using the Absolutely RNA RT-PCR Miniprep kit (Statagene). .. 2 μg RNA was reverse-transcribed, and PPARα, PPARδ, PPARγ, and β-actin cDNAs were amplified according to methods described previously ( ) using a Lightcycler (Roche).

Article Title: MERS-coronavirus replication induces severe in vitro cytopathology and is strongly inhibited by cyclosporin A or interferon-? treatment
Article Snippet: Paragraph title: Real-time reverse transcription-PCR (RT-PCR). ... RNA from 200 µl culture medium of CoV-infected cells was isolated with a MagnaPure LC Total Nucleic Acid Isolation kit (Roche) and eluted in 100 µl.

Article Title: HIV-1 RNA Levels and Antiretroviral Drug Resistance in Blood and Non-Blood Compartments from HIV-1-Infected Men and Women enrolled in AIDS Clinical Trials Group Study A5077
Article Snippet: .. Laboratory Testing HIV RNA was measured in blood plasma using RT-PCR (Monitor version 1.5 (Roche Molecular) at a central laboratory. .. HIV RNA was extracted into NucliSens extraction buffer from saliva, seminal plasma or endocervical wicks and quantified using published NucliSens (bioMerieux; saliva, seminal plasma) or real-time RT-PCR assay methods , , , .

Article Title: Arabidopsis ETO1 specifically interacts with and negatively regulates type 2 1-aminocyclopropane-1-carboxylate synthases
Article Snippet: .. RNA (1 μg) was used in each RT-PCR following the manufacturer's protocol (Titan One Tube RT-PCR System, Roche Diagnostics). ..

Lysis:

Article Title: Hepatitis B Viral DNA Decline at Loss of HBeAg Is Mainly Explained by Reduced cccDNA Load - Down-Regulated Transcription of PgRNA Has Limited Impact
Article Snippet: Extraction of DNA and RNA from Cells DNA and RNA were extracted from transfected and non-transfected hepatoma cells in a Magnapure robot (Roche Applied Science, Germany) using the Total NA protocol. .. Harvested cells were washed in 1 mL PBS and after centrifugation at 5000 rpm for 3 min the pellet was re-suspended in 800 µL RLT lysis buffer (Qiagen Sciences, MD, USA) before extraction.

Electrophoresis:

Article Title: Genome-wide analyses reveal the IRE1a-XBP1 pathway promotes T helper cell differentiation by resolving secretory stress and accelerating proliferation
Article Snippet: Reverse transcription quantitative PCR (RT-qPCR) Total RNA was isolated from two million cells by SV total RNA isolation kit (Promega). cDNA was prepared by annealing 500 ng RNA with oligo dT as per the manufacturer’s instructions (Transcriptor High Fidelity cDNA Synthesis kit, Roche). .. For XBP1, mRNA was amplified by PCR and products were separated by electrophoresis through a 2.5% agarose gel and visualized by ethidium bromide staining.

Selection:

Article Title: Peroxisome proliferator-activated receptor (PPAR)? expression in T cells mediates gender differences in development of T cell-mediated autoimmunity
Article Snippet: CD4+ and CD3+ T cells were purified by negative selection (columns from R & D Systems). .. 2 μg RNA was reverse-transcribed, and PPARα, PPARδ, PPARγ, and β-actin cDNAs were amplified according to methods described previously ( ) using a Lightcycler (Roche).

Agarose Gel Electrophoresis:

Article Title: Genome-wide analyses reveal the IRE1a-XBP1 pathway promotes T helper cell differentiation by resolving secretory stress and accelerating proliferation
Article Snippet: Reverse transcription quantitative PCR (RT-qPCR) Total RNA was isolated from two million cells by SV total RNA isolation kit (Promega). cDNA was prepared by annealing 500 ng RNA with oligo dT as per the manufacturer’s instructions (Transcriptor High Fidelity cDNA Synthesis kit, Roche). .. For XBP1, mRNA was amplified by PCR and products were separated by electrophoresis through a 2.5% agarose gel and visualized by ethidium bromide staining.

In Vitro:

Article Title: Ddx42p--a human DEAD box protein with RNA chaperone activities
Article Snippet: A ddx42-specific antisense RNA probe (complementary to nt 1797–3074 of the ddx42 cDNA) was synthesized by NheI-digest of pGEMddx42 (see below) and in vitro transcription with T7 RNA polymerase in the presence of Dig-RNA labeling mix (Roche). .. Northern blotting of 100 ng poly(A)+ RNA was carried out using the Dig-labeled ddx42-specific antisense RNA probe and detected with CSPD (Roche).

Staining:

Article Title: Genome-wide analyses reveal the IRE1a-XBP1 pathway promotes T helper cell differentiation by resolving secretory stress and accelerating proliferation
Article Snippet: Reverse transcription quantitative PCR (RT-qPCR) Total RNA was isolated from two million cells by SV total RNA isolation kit (Promega). cDNA was prepared by annealing 500 ng RNA with oligo dT as per the manufacturer’s instructions (Transcriptor High Fidelity cDNA Synthesis kit, Roche). .. For XBP1, mRNA was amplified by PCR and products were separated by electrophoresis through a 2.5% agarose gel and visualized by ethidium bromide staining.

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  • 93
    Roche automatic immunohistochemistry system
    Data analysis workflow. Fifty nine tumor samples from 56 NF1-glioma patients with 43 matched normal were profiled with WES, DNA Methylation profiles (31 tumors) and RNA sequencing (29 tumors). WES was used to call NF1 germline mutations using HaplotypeCaller and Somatic-germline log odds filter. Somatic SNVs were called from WES data by integrating the results of five algorithms (Freebayes, MuTect, Strelka, VarDict and VarScan). Recurrent CNVs were detected by GATK and GISTIC2. SNVs and CNVs were validated by Sanger sequencing (93% validation rate) and genomic qPCR (96% validation rate), respectively. Neoantigen prediction was obtained using netMHCpan and HLA genotype was determined by Polysolver, Optitype, Phlat and Seq2hla and validated by affinity binding kinetics. COSMIC cancer mutation signatures were identified by deconstructSig and compared to those occurring in sporadic glioma. DNA Methylation arrays were used to classify NF1 glioma in the methylation subtypes of sporadic glioma form the TCGA pan-glioma dataset (KNN). RNAseq was used to define gene expression clusters and immune subtypes of low-grade NF1-glioma and results were confirmed by RT-qPCR and <t>immunohistochemistry.</t> Integrative analysis of gene expression and DNA Methylation identified epigenetic signatures characterizing immune subtypes of low-grade glioma. A pan-glioma gene regulatory network was used to identify MRs of the ATRX -mutant phenotype in LGm6 sporadic and NF1-glioma (RGBM). Finally, the impact of ATRX mutation on survival was assessed using TCGA pan-glioma and NF1-glioma data.
    Automatic Immunohistochemistry System, supplied by Roche, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/automatic immunohistochemistry system/product/Roche
    Average 93 stars, based on 1 article reviews
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    automatic immunohistochemistry system - by Bioz Stars, 2020-04
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    93
    Roche atrx immunohistochemistry
    Data analysis workflow. Fifty nine tumor samples from 56 NF1-glioma patients with 43 matched normal were profiled with WES, DNA Methylation profiles (31 tumors) and RNA sequencing (29 tumors). WES was used to call NF1 germline mutations using HaplotypeCaller and Somatic-germline log odds filter. Somatic SNVs were called from WES data by integrating the results of five algorithms (Freebayes, MuTect, Strelka, VarDict and VarScan). Recurrent CNVs were detected by GATK and GISTIC2. SNVs and CNVs were validated by Sanger sequencing (93% validation rate) and genomic qPCR (96% validation rate), respectively. Neoantigen prediction was obtained using netMHCpan and HLA genotype was determined by Polysolver, Optitype, Phlat and Seq2hla and validated by affinity binding kinetics. COSMIC cancer mutation signatures were identified by deconstructSig and compared to those occurring in sporadic glioma. DNA Methylation arrays were used to classify NF1 glioma in the methylation subtypes of sporadic glioma form the TCGA pan-glioma dataset (KNN). RNAseq was used to define gene expression clusters and immune subtypes of low-grade NF1-glioma and results were confirmed by RT-qPCR and <t>immunohistochemistry.</t> Integrative analysis of gene expression and DNA Methylation identified epigenetic signatures characterizing immune subtypes of low-grade glioma. A pan-glioma gene regulatory network was used to identify MRs of the <t>ATRX</t> -mutant phenotype in LGm6 sporadic and NF1-glioma (RGBM). Finally, the impact of ATRX mutation on survival was assessed using TCGA pan-glioma and NF1-glioma data.
    Atrx Immunohistochemistry, supplied by Roche, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Roche rna fluorescent in situ hybridization rna fish lncrna loc100129973 subcellular localization
    <t>LncRNA</t> <t>LOC100129973</t> might function as a miRNA sponge. ( A,B ) Potential sites targeted by miR-4707-5p and miR-4767 in lncRNA LOC100129973 . ( C ) <t>RNA</t> Fluorescent in situ hybridization of the LncRNA LOC100129973 in HUVECs. The antisense probe was used as a negative control. Scale bar: 10 μm. ( D ) RNA-binding protein immunoprecipitation experiments were performed using Ago2 antibody in HUVECs. IgG was used as a negative control. Quantitative reverse transcription–polymerase chain reaction (qRT-PCR) was performed to detect pulled-down lncRNA LOC100129973 . The lncRNA LOC100129973 RNA level of input was set as 100%.
    Rna Fluorescent In Situ Hybridization Rna Fish Lncrna Loc100129973 Subcellular Localization, supplied by Roche, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 91 stars, based on 1 article reviews
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    85
    Roche in vitro reverse transcriptase polymerase chain reaction rt pcr assay
    <t>LncRNA</t> <t>LOC100129973</t> might function as a miRNA sponge. ( A,B ) Potential sites targeted by miR-4707-5p and miR-4767 in lncRNA LOC100129973 . ( C ) <t>RNA</t> Fluorescent in situ hybridization of the LncRNA LOC100129973 in HUVECs. The antisense probe was used as a negative control. Scale bar: 10 μm. ( D ) RNA-binding protein immunoprecipitation experiments were performed using Ago2 antibody in HUVECs. IgG was used as a negative control. Quantitative reverse transcription–polymerase chain reaction (qRT-PCR) was performed to detect pulled-down lncRNA LOC100129973 . The lncRNA LOC100129973 RNA level of input was set as 100%.
    In Vitro Reverse Transcriptase Polymerase Chain Reaction Rt Pcr Assay, supplied by Roche, used in various techniques. Bioz Stars score: 85/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/in vitro reverse transcriptase polymerase chain reaction rt pcr assay/product/Roche
    Average 85 stars, based on 3 article reviews
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    in vitro reverse transcriptase polymerase chain reaction rt pcr assay - by Bioz Stars, 2020-04
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    Data analysis workflow. Fifty nine tumor samples from 56 NF1-glioma patients with 43 matched normal were profiled with WES, DNA Methylation profiles (31 tumors) and RNA sequencing (29 tumors). WES was used to call NF1 germline mutations using HaplotypeCaller and Somatic-germline log odds filter. Somatic SNVs were called from WES data by integrating the results of five algorithms (Freebayes, MuTect, Strelka, VarDict and VarScan). Recurrent CNVs were detected by GATK and GISTIC2. SNVs and CNVs were validated by Sanger sequencing (93% validation rate) and genomic qPCR (96% validation rate), respectively. Neoantigen prediction was obtained using netMHCpan and HLA genotype was determined by Polysolver, Optitype, Phlat and Seq2hla and validated by affinity binding kinetics. COSMIC cancer mutation signatures were identified by deconstructSig and compared to those occurring in sporadic glioma. DNA Methylation arrays were used to classify NF1 glioma in the methylation subtypes of sporadic glioma form the TCGA pan-glioma dataset (KNN). RNAseq was used to define gene expression clusters and immune subtypes of low-grade NF1-glioma and results were confirmed by RT-qPCR and immunohistochemistry. Integrative analysis of gene expression and DNA Methylation identified epigenetic signatures characterizing immune subtypes of low-grade glioma. A pan-glioma gene regulatory network was used to identify MRs of the ATRX -mutant phenotype in LGm6 sporadic and NF1-glioma (RGBM). Finally, the impact of ATRX mutation on survival was assessed using TCGA pan-glioma and NF1-glioma data.

    Journal: Nature medicine

    Article Title: The molecular landscape of glioma in patients with Neurofibromatosis 1

    doi: 10.1038/s41591-018-0263-8

    Figure Lengend Snippet: Data analysis workflow. Fifty nine tumor samples from 56 NF1-glioma patients with 43 matched normal were profiled with WES, DNA Methylation profiles (31 tumors) and RNA sequencing (29 tumors). WES was used to call NF1 germline mutations using HaplotypeCaller and Somatic-germline log odds filter. Somatic SNVs were called from WES data by integrating the results of five algorithms (Freebayes, MuTect, Strelka, VarDict and VarScan). Recurrent CNVs were detected by GATK and GISTIC2. SNVs and CNVs were validated by Sanger sequencing (93% validation rate) and genomic qPCR (96% validation rate), respectively. Neoantigen prediction was obtained using netMHCpan and HLA genotype was determined by Polysolver, Optitype, Phlat and Seq2hla and validated by affinity binding kinetics. COSMIC cancer mutation signatures were identified by deconstructSig and compared to those occurring in sporadic glioma. DNA Methylation arrays were used to classify NF1 glioma in the methylation subtypes of sporadic glioma form the TCGA pan-glioma dataset (KNN). RNAseq was used to define gene expression clusters and immune subtypes of low-grade NF1-glioma and results were confirmed by RT-qPCR and immunohistochemistry. Integrative analysis of gene expression and DNA Methylation identified epigenetic signatures characterizing immune subtypes of low-grade glioma. A pan-glioma gene regulatory network was used to identify MRs of the ATRX -mutant phenotype in LGm6 sporadic and NF1-glioma (RGBM). Finally, the impact of ATRX mutation on survival was assessed using TCGA pan-glioma and NF1-glioma data.

    Article Snippet: For ATRX immunohistochemistry, deparaffinization and immunolabeling of sections were performed by a fully automatic immunohistochemistry system, Ventana BenchMark XT (Roche), using a streptavidin-peroxidase complex with diaminobenzidine as chromogen and hematoxylin counterstaining of nuclei.

    Techniques: DNA Methylation Assay, RNA Sequencing Assay, Sequencing, Real-time Polymerase Chain Reaction, Binding Assay, Mutagenesis, Methylation, Expressing, Quantitative RT-PCR, Immunohistochemistry

    Analysis of ATRX somatic mutations in NF1-glioma patients. a , ATRX mutations were identified by WES. The spectrum of ATRX somatic variants (SNVs and indels) is represented with each mutation shown only once per patient. We identified and validated by Sanger sequencing ATRX pathogenic mutations in 10 patients (1 low-grade glioma, 3%; 9 high-grade gliomas, 37.5%). b , The relative frequency of age distribution is represented for all patients (dashed black line, n = 55), ATRX wild-type (green line, n = 46), and ATRX mutant gliomas (red line, n = 9; for one patient carrying ATRX mutation age was unknown and was not included in the analysis). c , Microphotographs of ATRX immunohistochemistry in gliomas from NF1 patients. Representative images are from n = 7 low-grade gliomas (left) and n = 16 high-grade gliomas (right). Results were validated on more than ten independent samples to ensure the staining pattern on human tissue was reproducible. High-grade glioma samples were negative for ATRX expression whereas low-grade gliomas retained ATRX protein expression. d , Contingency table shows loss of ATRX protein expression in 8 of 16 high-grade and in none of 7 low-grade NF1-gliomas ( P = 0.05, two-sided Fisher’s exact test). e , C-circle (CC) assay was performed to measure ALT activity in NF1-glioma samples. The scatter plot reports the normalized CC content for each glioma according to ATRX mutational status: ATRX wild-type (blue, n = 11) and ATRX mutant gliomas (red, n = 10). For each group the median with interquartile range is indicated. All ATRX mutant gliomas but only one ATRX wild-type glioma showed increased ALT activity (normalized CC content greater than 1; P = 2.3×10 −5 , two-sided MWW test).

    Journal: Nature medicine

    Article Title: The molecular landscape of glioma in patients with Neurofibromatosis 1

    doi: 10.1038/s41591-018-0263-8

    Figure Lengend Snippet: Analysis of ATRX somatic mutations in NF1-glioma patients. a , ATRX mutations were identified by WES. The spectrum of ATRX somatic variants (SNVs and indels) is represented with each mutation shown only once per patient. We identified and validated by Sanger sequencing ATRX pathogenic mutations in 10 patients (1 low-grade glioma, 3%; 9 high-grade gliomas, 37.5%). b , The relative frequency of age distribution is represented for all patients (dashed black line, n = 55), ATRX wild-type (green line, n = 46), and ATRX mutant gliomas (red line, n = 9; for one patient carrying ATRX mutation age was unknown and was not included in the analysis). c , Microphotographs of ATRX immunohistochemistry in gliomas from NF1 patients. Representative images are from n = 7 low-grade gliomas (left) and n = 16 high-grade gliomas (right). Results were validated on more than ten independent samples to ensure the staining pattern on human tissue was reproducible. High-grade glioma samples were negative for ATRX expression whereas low-grade gliomas retained ATRX protein expression. d , Contingency table shows loss of ATRX protein expression in 8 of 16 high-grade and in none of 7 low-grade NF1-gliomas ( P = 0.05, two-sided Fisher’s exact test). e , C-circle (CC) assay was performed to measure ALT activity in NF1-glioma samples. The scatter plot reports the normalized CC content for each glioma according to ATRX mutational status: ATRX wild-type (blue, n = 11) and ATRX mutant gliomas (red, n = 10). For each group the median with interquartile range is indicated. All ATRX mutant gliomas but only one ATRX wild-type glioma showed increased ALT activity (normalized CC content greater than 1; P = 2.3×10 −5 , two-sided MWW test).

    Article Snippet: For ATRX immunohistochemistry, deparaffinization and immunolabeling of sections were performed by a fully automatic immunohistochemistry system, Ventana BenchMark XT (Roche), using a streptavidin-peroxidase complex with diaminobenzidine as chromogen and hematoxylin counterstaining of nuclei.

    Techniques: Mutagenesis, Sequencing, Immunohistochemistry, Staining, Expressing, Activity Assay

    T cell infiltration and neoantigen analysis in low-grade NF1-glioma subclusters. a , Representative microphotographs of CD3 immunohistochemistry in low-grade/high-immune (left panels) and low-grade/low-immune (right panels) NF1-gliomas. Results were validated on more than ten independent samples to ensure that the staining pattern on human tissue was reproducible. b , The number of CD3-positive cells was scored in at least 5 pictures from low-grade/high-immune ( n = 6, red dots) and low-grade/low-immune ( n = 6, green dots) (* P = 0.003, two-sided t -test with Welch correction; scatter plots show mean with minimum to maximum range). c , Representative microphotographs of CD8 immunohistochemistry in low-grade/high-immune (left panels) and low-grade/low-immune (right panels) NFI-gliomas. Results were validated on more than ten independent samples to ensure that the staining pattern on human tissue was reproducible. d , The number of CD8-positive cells was scored in at least 5 pictures from low-grade/high-immune ( n = 6, red dots) and low-grade/low-immune ( n = 6, green dots) (* P = 0.017, two-sided t -test with Welch correction; scatter plots show mean with minimum to maximum range). e , Representative microphotographs of GZMB immunohistochemistry in low-grade/high-immune (left panels) and low-grade/low-immune (right panels) NFI-gliomas. Results were validated on more than ten independent samples to ensure that the staining pattern on human tissue was reproducible. f , The number of GZMB-positive cells was scored in at least 5 pictures from low-grade/high-immune ( n = 6, red dots) and low-grade/low-immune ( n = 6, green dots) (* P = 0.004, two-sided t -test with Welch correction; scatter plots show mean with minimum to maximum range). g , Quantification of neoantigens in low-grade NFI-glioma subclusters. The number of neoantigens per somatic mutation was significantly higher in the set of low-grade/high-immune NFI-gliomas ( P = 0.034, two-sided MWW test). h , In vitro binding affinity kinetics of neoantigens and corresponding wild-type peptides for their restricted HLA class I allele. Data are shown as counts per second with increasing peptide concentration (log 10 M). Data are mean of two independent experiments. MT, mutant peptide; WT, wild-type peptide.

    Journal: Nature medicine

    Article Title: The molecular landscape of glioma in patients with Neurofibromatosis 1

    doi: 10.1038/s41591-018-0263-8

    Figure Lengend Snippet: T cell infiltration and neoantigen analysis in low-grade NF1-glioma subclusters. a , Representative microphotographs of CD3 immunohistochemistry in low-grade/high-immune (left panels) and low-grade/low-immune (right panels) NF1-gliomas. Results were validated on more than ten independent samples to ensure that the staining pattern on human tissue was reproducible. b , The number of CD3-positive cells was scored in at least 5 pictures from low-grade/high-immune ( n = 6, red dots) and low-grade/low-immune ( n = 6, green dots) (* P = 0.003, two-sided t -test with Welch correction; scatter plots show mean with minimum to maximum range). c , Representative microphotographs of CD8 immunohistochemistry in low-grade/high-immune (left panels) and low-grade/low-immune (right panels) NFI-gliomas. Results were validated on more than ten independent samples to ensure that the staining pattern on human tissue was reproducible. d , The number of CD8-positive cells was scored in at least 5 pictures from low-grade/high-immune ( n = 6, red dots) and low-grade/low-immune ( n = 6, green dots) (* P = 0.017, two-sided t -test with Welch correction; scatter plots show mean with minimum to maximum range). e , Representative microphotographs of GZMB immunohistochemistry in low-grade/high-immune (left panels) and low-grade/low-immune (right panels) NFI-gliomas. Results were validated on more than ten independent samples to ensure that the staining pattern on human tissue was reproducible. f , The number of GZMB-positive cells was scored in at least 5 pictures from low-grade/high-immune ( n = 6, red dots) and low-grade/low-immune ( n = 6, green dots) (* P = 0.004, two-sided t -test with Welch correction; scatter plots show mean with minimum to maximum range). g , Quantification of neoantigens in low-grade NFI-glioma subclusters. The number of neoantigens per somatic mutation was significantly higher in the set of low-grade/high-immune NFI-gliomas ( P = 0.034, two-sided MWW test). h , In vitro binding affinity kinetics of neoantigens and corresponding wild-type peptides for their restricted HLA class I allele. Data are shown as counts per second with increasing peptide concentration (log 10 M). Data are mean of two independent experiments. MT, mutant peptide; WT, wild-type peptide.

    Article Snippet: For ATRX immunohistochemistry, deparaffinization and immunolabeling of sections were performed by a fully automatic immunohistochemistry system, Ventana BenchMark XT (Roche), using a streptavidin-peroxidase complex with diaminobenzidine as chromogen and hematoxylin counterstaining of nuclei.

    Techniques: Immunohistochemistry, Staining, Mutagenesis, In Vitro, Binding Assay, Concentration Assay

    Data analysis workflow. Fifty nine tumor samples from 56 NF1-glioma patients with 43 matched normal were profiled with WES, DNA Methylation profiles (31 tumors) and RNA sequencing (29 tumors). WES was used to call NF1 germline mutations using HaplotypeCaller and Somatic-germline log odds filter. Somatic SNVs were called from WES data by integrating the results of five algorithms (Freebayes, MuTect, Strelka, VarDict and VarScan). Recurrent CNVs were detected by GATK and GISTIC2. SNVs and CNVs were validated by Sanger sequencing (93% validation rate) and genomic qPCR (96% validation rate), respectively. Neoantigen prediction was obtained using netMHCpan and HLA genotype was determined by Polysolver, Optitype, Phlat and Seq2hla and validated by affinity binding kinetics. COSMIC cancer mutation signatures were identified by deconstructSig and compared to those occurring in sporadic glioma. DNA Methylation arrays were used to classify NF1 glioma in the methylation subtypes of sporadic glioma form the TCGA pan-glioma dataset (KNN). RNAseq was used to define gene expression clusters and immune subtypes of low-grade NF1-glioma and results were confirmed by RT-qPCR and immunohistochemistry. Integrative analysis of gene expression and DNA Methylation identified epigenetic signatures characterizing immune subtypes of low-grade glioma. A pan-glioma gene regulatory network was used to identify MRs of the ATRX -mutant phenotype in LGm6 sporadic and NF1-glioma (RGBM). Finally, the impact of ATRX mutation on survival was assessed using TCGA pan-glioma and NF1-glioma data.

    Journal: Nature medicine

    Article Title: The molecular landscape of glioma in patients with Neurofibromatosis 1

    doi: 10.1038/s41591-018-0263-8

    Figure Lengend Snippet: Data analysis workflow. Fifty nine tumor samples from 56 NF1-glioma patients with 43 matched normal were profiled with WES, DNA Methylation profiles (31 tumors) and RNA sequencing (29 tumors). WES was used to call NF1 germline mutations using HaplotypeCaller and Somatic-germline log odds filter. Somatic SNVs were called from WES data by integrating the results of five algorithms (Freebayes, MuTect, Strelka, VarDict and VarScan). Recurrent CNVs were detected by GATK and GISTIC2. SNVs and CNVs were validated by Sanger sequencing (93% validation rate) and genomic qPCR (96% validation rate), respectively. Neoantigen prediction was obtained using netMHCpan and HLA genotype was determined by Polysolver, Optitype, Phlat and Seq2hla and validated by affinity binding kinetics. COSMIC cancer mutation signatures were identified by deconstructSig and compared to those occurring in sporadic glioma. DNA Methylation arrays were used to classify NF1 glioma in the methylation subtypes of sporadic glioma form the TCGA pan-glioma dataset (KNN). RNAseq was used to define gene expression clusters and immune subtypes of low-grade NF1-glioma and results were confirmed by RT-qPCR and immunohistochemistry. Integrative analysis of gene expression and DNA Methylation identified epigenetic signatures characterizing immune subtypes of low-grade glioma. A pan-glioma gene regulatory network was used to identify MRs of the ATRX -mutant phenotype in LGm6 sporadic and NF1-glioma (RGBM). Finally, the impact of ATRX mutation on survival was assessed using TCGA pan-glioma and NF1-glioma data.

    Article Snippet: For ATRX immunohistochemistry, deparaffinization and immunolabeling of sections were performed by a fully automatic immunohistochemistry system, Ventana BenchMark XT (Roche), using a streptavidin-peroxidase complex with diaminobenzidine as chromogen and hematoxylin counterstaining of nuclei.

    Techniques: DNA Methylation Assay, RNA Sequencing Assay, Sequencing, Real-time Polymerase Chain Reaction, Binding Assay, Mutagenesis, Methylation, Expressing, Quantitative RT-PCR, Immunohistochemistry

    Analysis of ATRX somatic mutations in NF1-glioma patients. a , ATRX mutations were identified by WES. The spectrum of ATRX somatic variants (SNVs and indels) is represented with each mutation shown only once per patient. We identified and validated by Sanger sequencing ATRX pathogenic mutations in 10 patients (1 low-grade glioma, 3%; 9 high-grade gliomas, 37.5%). b , The relative frequency of age distribution is represented for all patients (dashed black line, n = 55), ATRX wild-type (green line, n = 46), and ATRX mutant gliomas (red line, n = 9; for one patient carrying ATRX mutation age was unknown and was not included in the analysis). c , Microphotographs of ATRX immunohistochemistry in gliomas from NF1 patients. Representative images are from n = 7 low-grade gliomas (left) and n = 16 high-grade gliomas (right). Results were validated on more than ten independent samples to ensure the staining pattern on human tissue was reproducible. High-grade glioma samples were negative for ATRX expression whereas low-grade gliomas retained ATRX protein expression. d , Contingency table shows loss of ATRX protein expression in 8 of 16 high-grade and in none of 7 low-grade NF1-gliomas ( P = 0.05, two-sided Fisher’s exact test). e , C-circle (CC) assay was performed to measure ALT activity in NF1-glioma samples. The scatter plot reports the normalized CC content for each glioma according to ATRX mutational status: ATRX wild-type (blue, n = 11) and ATRX mutant gliomas (red, n = 10). For each group the median with interquartile range is indicated. All ATRX mutant gliomas but only one ATRX wild-type glioma showed increased ALT activity (normalized CC content greater than 1; P = 2.3×10 −5 , two-sided MWW test).

    Journal: Nature medicine

    Article Title: The molecular landscape of glioma in patients with Neurofibromatosis 1

    doi: 10.1038/s41591-018-0263-8

    Figure Lengend Snippet: Analysis of ATRX somatic mutations in NF1-glioma patients. a , ATRX mutations were identified by WES. The spectrum of ATRX somatic variants (SNVs and indels) is represented with each mutation shown only once per patient. We identified and validated by Sanger sequencing ATRX pathogenic mutations in 10 patients (1 low-grade glioma, 3%; 9 high-grade gliomas, 37.5%). b , The relative frequency of age distribution is represented for all patients (dashed black line, n = 55), ATRX wild-type (green line, n = 46), and ATRX mutant gliomas (red line, n = 9; for one patient carrying ATRX mutation age was unknown and was not included in the analysis). c , Microphotographs of ATRX immunohistochemistry in gliomas from NF1 patients. Representative images are from n = 7 low-grade gliomas (left) and n = 16 high-grade gliomas (right). Results were validated on more than ten independent samples to ensure the staining pattern on human tissue was reproducible. High-grade glioma samples were negative for ATRX expression whereas low-grade gliomas retained ATRX protein expression. d , Contingency table shows loss of ATRX protein expression in 8 of 16 high-grade and in none of 7 low-grade NF1-gliomas ( P = 0.05, two-sided Fisher’s exact test). e , C-circle (CC) assay was performed to measure ALT activity in NF1-glioma samples. The scatter plot reports the normalized CC content for each glioma according to ATRX mutational status: ATRX wild-type (blue, n = 11) and ATRX mutant gliomas (red, n = 10). For each group the median with interquartile range is indicated. All ATRX mutant gliomas but only one ATRX wild-type glioma showed increased ALT activity (normalized CC content greater than 1; P = 2.3×10 −5 , two-sided MWW test).

    Article Snippet: For ATRX immunohistochemistry, deparaffinization and immunolabeling of sections were performed by a fully automatic immunohistochemistry system, Ventana BenchMark XT (Roche), using a streptavidin-peroxidase complex with diaminobenzidine as chromogen and hematoxylin counterstaining of nuclei.

    Techniques: Mutagenesis, Sequencing, Immunohistochemistry, Staining, Expressing, Activity Assay

    LncRNA LOC100129973 might function as a miRNA sponge. ( A,B ) Potential sites targeted by miR-4707-5p and miR-4767 in lncRNA LOC100129973 . ( C ) RNA Fluorescent in situ hybridization of the LncRNA LOC100129973 in HUVECs. The antisense probe was used as a negative control. Scale bar: 10 μm. ( D ) RNA-binding protein immunoprecipitation experiments were performed using Ago2 antibody in HUVECs. IgG was used as a negative control. Quantitative reverse transcription–polymerase chain reaction (qRT-PCR) was performed to detect pulled-down lncRNA LOC100129973 . The lncRNA LOC100129973 RNA level of input was set as 100%.

    Journal: Scientific Reports

    Article Title: Long Noncoding RNA LOC100129973 Suppresses Apoptosis by Targeting miR-4707-5p and miR-4767 in Vascular Endothelial Cells

    doi: 10.1038/srep21620

    Figure Lengend Snippet: LncRNA LOC100129973 might function as a miRNA sponge. ( A,B ) Potential sites targeted by miR-4707-5p and miR-4767 in lncRNA LOC100129973 . ( C ) RNA Fluorescent in situ hybridization of the LncRNA LOC100129973 in HUVECs. The antisense probe was used as a negative control. Scale bar: 10 μm. ( D ) RNA-binding protein immunoprecipitation experiments were performed using Ago2 antibody in HUVECs. IgG was used as a negative control. Quantitative reverse transcription–polymerase chain reaction (qRT-PCR) was performed to detect pulled-down lncRNA LOC100129973 . The lncRNA LOC100129973 RNA level of input was set as 100%.

    Article Snippet: RNA fluorescent in situ hybridization (RNA-FISH) LncRNA LOC100129973 subcellular localization in HUVECs was detected by use of a FISH kit (Roche Applied Science, Germany).

    Techniques: In Situ Hybridization, Negative Control, RNA Binding Assay, Immunoprecipitation, Reverse Transcription Polymerase Chain Reaction, Quantitative RT-PCR

    LncRNA L OC100129973 suppressed the serum and FGF-2 starvation-induced apoptosis in HUVECs. ( A ) Quantified real-time PCR analysis of lncRNA LOC100129973 overexpression and knock down efficiency. HUVECs were transfected with pcDNA3.1- LOC100129973 at 0.1, 0.2, 0.4 μg/mL or si LOC100129973 at 10, 20, 40 nM for 24 h and then starvation for another 24 h. ( B ) Hoechst 33258 staining of apoptotic HUVECs. Nor: M199 medium (with FGF-2 and serum); OE-Ctr: transfected with pcDNA3.1-empty vector; OE- LOC100129973 : transfected with pcDNA3.1- LOC100129973 at 0.2 μg/mL; siCtr: transfected with scramble RNA for negative control; si LOC100129973 : transfected with si LOC100129973 at 40 nM. After being transfected for 24 h, all the cells described above were deprived of serum and FGF-2 for another 12 or 24 h. Scale bar: 10 μm. Images are representative of at least 3 independent experiments. The percent of apoptosis was measured (%). ( C , D ) Western blot analysis of cleaved PARP protein level. HUVECs were transfected with pcDNA3.1- LOC100129973 at 0.1, 0.2, 0.4 μg/mL or si LOC100129973 at 10, 20, 40 nM for 24 h and starvation for another 24 h. The level of cleaved PARP was relative to that of β-actin. (Cropped, full-length blots are in Supplementary Fig. S4 ) Data are mean ± SEM. of three independent experiments. *P

    Journal: Scientific Reports

    Article Title: Long Noncoding RNA LOC100129973 Suppresses Apoptosis by Targeting miR-4707-5p and miR-4767 in Vascular Endothelial Cells

    doi: 10.1038/srep21620

    Figure Lengend Snippet: LncRNA L OC100129973 suppressed the serum and FGF-2 starvation-induced apoptosis in HUVECs. ( A ) Quantified real-time PCR analysis of lncRNA LOC100129973 overexpression and knock down efficiency. HUVECs were transfected with pcDNA3.1- LOC100129973 at 0.1, 0.2, 0.4 μg/mL or si LOC100129973 at 10, 20, 40 nM for 24 h and then starvation for another 24 h. ( B ) Hoechst 33258 staining of apoptotic HUVECs. Nor: M199 medium (with FGF-2 and serum); OE-Ctr: transfected with pcDNA3.1-empty vector; OE- LOC100129973 : transfected with pcDNA3.1- LOC100129973 at 0.2 μg/mL; siCtr: transfected with scramble RNA for negative control; si LOC100129973 : transfected with si LOC100129973 at 40 nM. After being transfected for 24 h, all the cells described above were deprived of serum and FGF-2 for another 12 or 24 h. Scale bar: 10 μm. Images are representative of at least 3 independent experiments. The percent of apoptosis was measured (%). ( C , D ) Western blot analysis of cleaved PARP protein level. HUVECs were transfected with pcDNA3.1- LOC100129973 at 0.1, 0.2, 0.4 μg/mL or si LOC100129973 at 10, 20, 40 nM for 24 h and starvation for another 24 h. The level of cleaved PARP was relative to that of β-actin. (Cropped, full-length blots are in Supplementary Fig. S4 ) Data are mean ± SEM. of three independent experiments. *P

    Article Snippet: RNA fluorescent in situ hybridization (RNA-FISH) LncRNA LOC100129973 subcellular localization in HUVECs was detected by use of a FISH kit (Roche Applied Science, Germany).

    Techniques: Real-time Polymerase Chain Reaction, Over Expression, Transfection, Staining, Plasmid Preparation, Negative Control, Western Blot