quantitative real time pcr assay total rna  (Thermo Fisher)


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    Name:
    TRIzol Reagent
    Description:
    TRIzol Reagent is a complete ready to use reagent for the isolation of high quality total RNA or the simultaneous isolation of RNA DNA and protein from a variety of biological samples This monophasic solution of phenol and guanidine isothiocyanate is designed to isolate separate fractions of RNA DNA and proteins from cell and tissue samples of human animal plant yeast or bacterial origin within one hour Key features of TRIzol Reagent include • Permits the isolation of RNA DNA and protein from the same sample• Offers superior lysis capability even with difficult sample types• Optimized formulations and protocols for tissues cells serum virus and bacteriaReliably purify RNA from multiple sample volumes and sourcesTRIzol Reagent performs well with small quantities of tissue 50 100 mg and cells 5 × 106 as well as with large quantities of tissue 1 g and cells 107 and comes with prototcols for purification from samples of human animal plant or bacterial origin TRIzol Reagent maintains the integrity of the RNA due to highly effective inhibition of RNase activity while disrupting cells and dissolving cell components during sample homogenization The simplicity of the TRIzol Reagent method allows simultaneous processing of a large number of samples The entire procedure can be completed in 1 hour Total RNA isolated by TRIzol Reagent is free of protein and DNA contamination Formulated for isolation of multiple molecular targetsTRIzol Reagent allows you to perform sequential precipitation of RNA DNA and proteins from a single sample After homogenizing the sample with TRIzol Reagent chloroform is added and the homogenate is allowed to separate into a clear upper aqueous layer containing RNA and interphase and red lower organic layers containing the DNA and proteins RNA is precipitated from the aqueous layer with isopropanol DNA is precipitated from the interphase organic layer with ethanol Protein is precipitated from the phenol ethanol supernatant by isopropanol precipitation The precipitated RNA DNA or protein is washed to remove impurities and then resuspended for use in downstream applications
    Catalog Number:
    15596018
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    None
    Applications:
    DNA & RNA Purification & Analysis|RNAi, Epigenetics & Non-Coding RNA Research|RNA Extraction|Total RNA Isolation|Total RNA from Animal Cells & Tissues|Total RNA from Bacterial Samples|Total RNA from Plant Cells|Total RNA from Yeast|miRNA Isolation|miRNA & Non-Coding RNA Analysis
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    Structured Review

    Thermo Fisher quantitative real time pcr assay total rna
    Stable knockdown of Capn4 reduced ZEB1 expression and inhibited esophageal squamous cell carcinoma (ESCC) invasion and metastasis in vitro and in vivo. ( a ) Quantitative real‐time <t>PCR</t> (qRT‐PCR) analysis of Capn4 and ZEB1 expression in human esophageal epithelial and ESCC cell lines. ( b , c ) qRT‐PCR analyses were used to detect Capn4 and ZEB1 messenger <t>RNA</t> (mRNA) expression in ECA109 and TE7 cells stably transfected with the short hairpin negative control (shNC) vector or shCapn4 plasmid (** P
    TRIzol Reagent is a complete ready to use reagent for the isolation of high quality total RNA or the simultaneous isolation of RNA DNA and protein from a variety of biological samples This monophasic solution of phenol and guanidine isothiocyanate is designed to isolate separate fractions of RNA DNA and proteins from cell and tissue samples of human animal plant yeast or bacterial origin within one hour Key features of TRIzol Reagent include • Permits the isolation of RNA DNA and protein from the same sample• Offers superior lysis capability even with difficult sample types• Optimized formulations and protocols for tissues cells serum virus and bacteriaReliably purify RNA from multiple sample volumes and sourcesTRIzol Reagent performs well with small quantities of tissue 50 100 mg and cells 5 × 106 as well as with large quantities of tissue 1 g and cells 107 and comes with prototcols for purification from samples of human animal plant or bacterial origin TRIzol Reagent maintains the integrity of the RNA due to highly effective inhibition of RNase activity while disrupting cells and dissolving cell components during sample homogenization The simplicity of the TRIzol Reagent method allows simultaneous processing of a large number of samples The entire procedure can be completed in 1 hour Total RNA isolated by TRIzol Reagent is free of protein and DNA contamination Formulated for isolation of multiple molecular targetsTRIzol Reagent allows you to perform sequential precipitation of RNA DNA and proteins from a single sample After homogenizing the sample with TRIzol Reagent chloroform is added and the homogenate is allowed to separate into a clear upper aqueous layer containing RNA and interphase and red lower organic layers containing the DNA and proteins RNA is precipitated from the aqueous layer with isopropanol DNA is precipitated from the interphase organic layer with ethanol Protein is precipitated from the phenol ethanol supernatant by isopropanol precipitation The precipitated RNA DNA or protein is washed to remove impurities and then resuspended for use in downstream applications
    https://www.bioz.com/result/quantitative real time pcr assay total rna/product/Thermo Fisher
    Average 99 stars, based on 17 article reviews
    Price from $9.99 to $1999.99
    quantitative real time pcr assay total rna - by Bioz Stars, 2020-08
    99/100 stars

    Images

    1) Product Images from "Capn4 promotes esophageal squamous cell carcinoma metastasis by regulating ZEB1 through the Wnt/β‐catenin signaling pathway"

    Article Title: Capn4 promotes esophageal squamous cell carcinoma metastasis by regulating ZEB1 through the Wnt/β‐catenin signaling pathway

    Journal: Thoracic Cancer

    doi: 10.1111/1759-7714.12893

    Stable knockdown of Capn4 reduced ZEB1 expression and inhibited esophageal squamous cell carcinoma (ESCC) invasion and metastasis in vitro and in vivo. ( a ) Quantitative real‐time PCR (qRT‐PCR) analysis of Capn4 and ZEB1 expression in human esophageal epithelial and ESCC cell lines. ( b , c ) qRT‐PCR analyses were used to detect Capn4 and ZEB1 messenger RNA (mRNA) expression in ECA109 and TE7 cells stably transfected with the short hairpin negative control (shNC) vector or shCapn4 plasmid (** P
    Figure Legend Snippet: Stable knockdown of Capn4 reduced ZEB1 expression and inhibited esophageal squamous cell carcinoma (ESCC) invasion and metastasis in vitro and in vivo. ( a ) Quantitative real‐time PCR (qRT‐PCR) analysis of Capn4 and ZEB1 expression in human esophageal epithelial and ESCC cell lines. ( b , c ) qRT‐PCR analyses were used to detect Capn4 and ZEB1 messenger RNA (mRNA) expression in ECA109 and TE7 cells stably transfected with the short hairpin negative control (shNC) vector or shCapn4 plasmid (** P

    Techniques Used: Expressing, In Vitro, In Vivo, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Stable Transfection, Transfection, Negative Control, Plasmid Preparation

    Capn4 overexpression is correlated with ZEB1 expression in human esophageal squamous cell carcinoma (ESCC). ( a , b ) Quantitative real‐time PCR analysis of Capn4 and ZEB1 messenger RNA (mRNA) expression in 86 ESCC and paired non‐tumor tissues (** P
    Figure Legend Snippet: Capn4 overexpression is correlated with ZEB1 expression in human esophageal squamous cell carcinoma (ESCC). ( a , b ) Quantitative real‐time PCR analysis of Capn4 and ZEB1 messenger RNA (mRNA) expression in 86 ESCC and paired non‐tumor tissues (** P

    Techniques Used: Over Expression, Expressing, Real-time Polymerase Chain Reaction

    2) Product Images from "Targeting Long Noncoding RNA HMMR-AS1 Suppresses and Radiosensitizes Glioblastoma"

    Article Title: Targeting Long Noncoding RNA HMMR-AS1 Suppresses and Radiosensitizes Glioblastoma

    Journal: Neoplasia (New York, N.Y.)

    doi: 10.1016/j.neo.2018.02.010

    HMMR-AS1 stabilizes HMMR mRNA. (A) Relative levels of HMMR-AS1 and HMMR mRNA in U87 and HA cells transfected with lentivirus containing HMMR-AS1 full-length sequence compare with control. (B) HMMR protein expression tested by Western blot in cells that HMMR-AS1 was downregulated or upregulated. (C) HMMR protein expression tested by immunofluorescence in control cells or cells with HMMR-AS1 or HMMR overexpression. Scale bar = 10 μm. (D) U87 cells differently transfected were treated with 50 μM α-amanitin for 6, 12, or 18 hours. Cells were then harvested for qRT-PCR to test the loss of HMMR mRNA. 18s RNA, a product of RNA polymerase I, was not affected by α-amanitin. Cells transfected with HMMR shRNA were used as positive control. β-Actin was used as a loading control for Western blot. The blot bands were quantified by ImageJ and represented by relative values compare with loading control (1.00). # P > .05, * P
    Figure Legend Snippet: HMMR-AS1 stabilizes HMMR mRNA. (A) Relative levels of HMMR-AS1 and HMMR mRNA in U87 and HA cells transfected with lentivirus containing HMMR-AS1 full-length sequence compare with control. (B) HMMR protein expression tested by Western blot in cells that HMMR-AS1 was downregulated or upregulated. (C) HMMR protein expression tested by immunofluorescence in control cells or cells with HMMR-AS1 or HMMR overexpression. Scale bar = 10 μm. (D) U87 cells differently transfected were treated with 50 μM α-amanitin for 6, 12, or 18 hours. Cells were then harvested for qRT-PCR to test the loss of HMMR mRNA. 18s RNA, a product of RNA polymerase I, was not affected by α-amanitin. Cells transfected with HMMR shRNA were used as positive control. β-Actin was used as a loading control for Western blot. The blot bands were quantified by ImageJ and represented by relative values compare with loading control (1.00). # P > .05, * P

    Techniques Used: Transfection, Sequencing, Expressing, Western Blot, Immunofluorescence, Over Expression, Quantitative RT-PCR, shRNA, Positive Control

    3) Product Images from "OsACOS12, an orthologue of Arabidopsis acyl-CoA synthetase5, plays an important role in pollen exine formation and anther development in rice"

    Article Title: OsACOS12, an orthologue of Arabidopsis acyl-CoA synthetase5, plays an important role in pollen exine formation and anther development in rice

    Journal: BMC Plant Biology

    doi: 10.1186/s12870-016-0943-9

    OsACOS12 is specifically expressed in the anther. a RT-PCR analysis of RNA isolated from various tissues using OsACOS12 and OsACTIN primer sets. Le, lemma; Pa, palea; L2.5, Glumes length 2.5 mm; L3.0, Glumes length 3.0 mm; L3.5, Glumes length 3.5 mm; L4.0, Glumes length 4.0 mm; L5.6, Glumes length 5.6 mm. b Quantitative real-time PCR analysis of OsACOS12 . The OsACTIN gene served as the reference. Data are shown as the mean ± SD ( n = 3). c - j In situ hybridization of OsACOS12 in WT anthers. The anthers at the MMC stage ( c ), early meiosis stage ( d ), tetrad stage ( e and f ), microspore release stage ( g ), and microspore vacuolate stage ( h ) hybridized with an OsACOS12 antisense probe. The anthers at the tetrad stage ( i - j ) hybridized with an OsACOS12 sense probe. Msp, microspore; T, tapetum; Tds, tetrads. MMC, microspore mother cell; MC, meiotic cell; Dy, dyad cell. Bars = 50 μm. k - p Fluorescence confocal images of the OsACOS12-GFP fusion proteins at different stages. The green channel shows the GFP expression (530 nm), and the red channel shows the chlorophyll autofluorescence ( > 560 nm). The bright-field images of ( p ) show that these fusion proteins are not localized to the microspores. Bars = 10 μm; 100 μm
    Figure Legend Snippet: OsACOS12 is specifically expressed in the anther. a RT-PCR analysis of RNA isolated from various tissues using OsACOS12 and OsACTIN primer sets. Le, lemma; Pa, palea; L2.5, Glumes length 2.5 mm; L3.0, Glumes length 3.0 mm; L3.5, Glumes length 3.5 mm; L4.0, Glumes length 4.0 mm; L5.6, Glumes length 5.6 mm. b Quantitative real-time PCR analysis of OsACOS12 . The OsACTIN gene served as the reference. Data are shown as the mean ± SD ( n = 3). c - j In situ hybridization of OsACOS12 in WT anthers. The anthers at the MMC stage ( c ), early meiosis stage ( d ), tetrad stage ( e and f ), microspore release stage ( g ), and microspore vacuolate stage ( h ) hybridized with an OsACOS12 antisense probe. The anthers at the tetrad stage ( i - j ) hybridized with an OsACOS12 sense probe. Msp, microspore; T, tapetum; Tds, tetrads. MMC, microspore mother cell; MC, meiotic cell; Dy, dyad cell. Bars = 50 μm. k - p Fluorescence confocal images of the OsACOS12-GFP fusion proteins at different stages. The green channel shows the GFP expression (530 nm), and the red channel shows the chlorophyll autofluorescence ( > 560 nm). The bright-field images of ( p ) show that these fusion proteins are not localized to the microspores. Bars = 10 μm; 100 μm

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, Isolation, Real-time Polymerase Chain Reaction, In Situ Hybridization, Fluorescence, Expressing

    Related Articles

    Isolation:

    Article Title: A highly effective and versatile technology for the isolation of RNAs from grapevines and other woody perennials for use in virus diagnostics
    Article Snippet: .. Commercial RNA isolation kits Five commercial kits were selected for this study, which included TRIzol Reagent (Life Technologies), RNeasy Plant mini kit (Qiagen), Spectrum™ Plant Total RNA kit (Sigma), AccuPrep viral RNA extraction kit (Bioneer) and Plant/fungi total RNA kit (Norgen BioTek). .. Methods of sample homogenization and RNA isolation

    Article Title: Modulation of microRNA Expression in Subjects with Metabolic Syndrome and Decrease of Cholesterol Efflux from Macrophages via microRNA-33-Mediated Attenuation of ATP-Binding Cassette Transporter A1 Expression by Statins
    Article Snippet: .. Total RNA was isolated from human plasma using Trizol reagent (Invitrogen, Carlsbad, CA, USA). microRNA was prepared using the mirVANA microRNA isolation kit (Ambion, Austin, TX, USA) according to the manufacturer’s instructions. .. RNA concentrations were determined using a Nanodrop 1000 spectrophotometer (Nanodrop Technologies).

    Article Title: UTX Affects Neural Stem Cell Proliferation and Differentiation through PTEN Signaling
    Article Snippet: .. RNA Isolation and qRT-PCR Total RNA was extracted with TRIzol (Invitrogen) and then inverse transcribed into cDNA using the FastQuant RT Kit (Tiangen Biotech). .. Real-time qPCR was performed according to the Super Real PreMix Plus (SYBR Green I) (Tiangen Biotech) in a 20-μL system on an ABI PRISM 7500 sequence detector system (Applied Biosystems).

    RNA Extraction:

    Article Title: High-quality RNA extraction from the sea urchin Paracentrotus lividus embryos
    Article Snippet: .. RNA extraction Five different methods of RNA extraction were compared. i) TRIzol® RNA extraction method Total RNA was extracted using TRIzol (Invitrogen, Life Technologies, Carlsbad, CA, USA) according to the manufacturer’s instructions, homogenizing with TissueLyser (Qiagen, Austin, TX, US) with 3 mm sterile aluminium beads at 20.1 Hertz (Hz) for 5 minutes. ..

    Article Title: A highly effective and versatile technology for the isolation of RNAs from grapevines and other woody perennials for use in virus diagnostics
    Article Snippet: .. Commercial RNA isolation kits Five commercial kits were selected for this study, which included TRIzol Reagent (Life Technologies), RNeasy Plant mini kit (Qiagen), Spectrum™ Plant Total RNA kit (Sigma), AccuPrep viral RNA extraction kit (Bioneer) and Plant/fungi total RNA kit (Norgen BioTek). .. Methods of sample homogenization and RNA isolation

    Article Title: Dysregulation of Lipid Metabolism in Mkp-1 Deficient Mice during Gram-Negative Sepsis
    Article Snippet: .. Liver RNA Extraction Total RNA was extracted from frozen liver tissues using Trizol reagent (Thermo Fisher Scientific, Waltham, MA, USA), solubilized in UltraPure RNase/DNase-free water (Thermo Fisher Scientific), and quantified by using NanoDrop ND-1000 spectrophotometer (Marshall Scientific, Hampton, NH, USA). .. RNA-Seq For RNA-seq analyses, 1 µg total RNA was used as starting material.

    Quantitative RT-PCR:

    Article Title: UTX Affects Neural Stem Cell Proliferation and Differentiation through PTEN Signaling
    Article Snippet: .. RNA Isolation and qRT-PCR Total RNA was extracted with TRIzol (Invitrogen) and then inverse transcribed into cDNA using the FastQuant RT Kit (Tiangen Biotech). .. Real-time qPCR was performed according to the Super Real PreMix Plus (SYBR Green I) (Tiangen Biotech) in a 20-μL system on an ABI PRISM 7500 sequence detector system (Applied Biosystems).

    Spectrophotometry:

    Article Title: Dysregulation of Lipid Metabolism in Mkp-1 Deficient Mice during Gram-Negative Sepsis
    Article Snippet: .. Liver RNA Extraction Total RNA was extracted from frozen liver tissues using Trizol reagent (Thermo Fisher Scientific, Waltham, MA, USA), solubilized in UltraPure RNase/DNase-free water (Thermo Fisher Scientific), and quantified by using NanoDrop ND-1000 spectrophotometer (Marshall Scientific, Hampton, NH, USA). .. RNA-Seq For RNA-seq analyses, 1 µg total RNA was used as starting material.

    Purification:

    Article Title: gp63 Homologues in Trypanosoma cruzi: Surface Antigens with Metalloprotease Activity and a Possible Role in Host Cell Infection
    Article Snippet: .. RNA was purified using TRIzol reagent (Life Technologies, Inc.) by following the manufacturer's instructions. ..

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    Thermo Fisher quantitative rt pcr analysis total rna
    Stable forced SUMO1 expression enhanced the colony formation, proliferation, migration, cell cycle progression, and invasion of A549 cells in vitro. ( a ) Detection of messenger <t>RNA</t> (mRNA) expression of SUMO1 in different lung cancer cell lines by quantitative real time <t>(qRT)‐PCR.</t> ( b ) Similar results were obtained through Western blot analysis. ( c ) qRT‐PCR analysis revealed that SUMO1 mRNA expression levels were increased in SUMO1 overexpressed A549 cells compared to control cells. ( d ) Similar results were obtained through Western blot analysis (passages 15 and 30). Upregulation of SUMO1 enhanced the ( e , f ) colony‐formation ability, ( g ) proliferation, ( h , i ) migration, and ( k , l ) invasion of A549 cells. ( j ) Forced expression of SUMO1 increased the number of A549 cells in the S phase of the cell cycle. * P
    Quantitative Rt Pcr Analysis Total Rna, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 169 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/quantitative rt pcr analysis total rna/product/Thermo Fisher
    Average 94 stars, based on 169 article reviews
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    Thermo Fisher quantitative real time polymerase chain reaction qrt pcr total rna
    Up-regulation of miR-96 and down-regulation of FOXO1 in tumor tissues and HepG2 cells. a miR-96 relative expression in tumor tissues and non-tumor tissues by <t>qRT-PCR;</t> b FOXO1 mRNA relative expression in tumor tissues and non-tumor tissues by qRT-PCR; c FOXO1 protein relative expression in tumor tissues and non-tumor tissues by Western blot; d miR-96 relative expression in L02 and HepG2 cells by qRT-PCR; e FOXO1 mRNA relative expression in L02 and HepG2 cells by qRT-PCR; f FOXO1 protein relative expression in L02 and HepG2 cells by Western blot. ** P
    Quantitative Real Time Polymerase Chain Reaction Qrt Pcr Total Rna, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 424 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/quantitative real time polymerase chain reaction qrt pcr total rna/product/Thermo Fisher
    Average 94 stars, based on 424 article reviews
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    95
    Thermo Fisher rna
    MD001 stimulates fatty acid oxidation in vitro . HepG2 ( A ), differentiated 3T3-L1 adipocyte ( B ), and differentiated C2C12 myotubes ( C ) were treated with vehicle or MD001 (10 μM) for 24 h and total <t>RNA</t> was isolated for <t>cDNA</t> synthesis. Relative gene expressions were analysed by qRT-PCR. HepG2 ( D ), differentiated 3T3-L1 adipocytes ( E ), and differentiated C2C12 myotubes ( F ) were treated with vehicle, MD001 (10 μM), or WY14643 (WY) (10 μM) for 24 h and the fatty acid oxidation rate was analysed as described in the Methods section. To confirm whether the fatty acid oxidation rate enhanced by MD001 is mediated by PPARα, HepG2 ( G ), differentiated 3T3-L1 ( H ), and differentiated C2C12 myotubes ( I ) were transfected with control or PPARα siRNA (20 nM) for 48 h, followed by treatment with vehicle, MD001 (10 μM), or WY14643 (WY) (10 μM) for 24 h. WY14643 was treated as positive control. *, **, vs. vehicle; † and ‡ , vs. vehicle/control siRNA; § , vs. WY/control siRNA; ¶ , vs. MD001/control siRNA. Data represent the mean ± SD of three independent experiments. * P
    Rna, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 95/100, based on 3653 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Thermo Fisher real time quantitative pcr qpcr total rna
    Pcf11 Expression Is Regulated by IPA (A) Schematic of mouse Pcf11 gene. Multiple PASs are indicated. EPA, internal exonic polyadenylation; IPA, intronic polyadenylation; TPA, 3′ terminal exon polyadenylation. 3′READS data of brain, heart, and testis are shown. (B) Features of intron 1 compared with other introns in the mouse genome. Each feature has ten bins, and the bin containing intron 1 of Pcf11 is highlighted in red. Distance to TSS is based on all IPA sites in PolyA_DB. (C) <t>RT-qPCR</t> analysis of IPA and TPA isoform expression in siPcf11 versus siCtrl cells. Error bars are SD on the basis of three replicates. (D) Top: schematic showing two sgRNAs used to remove a region flanking the IPA site. Bottom: validation of IPA site knockout (IPA Pcf11 -KO) by genomic DNA <t>PCR</t> using primers flanking the KO region. WT, wild-type 4T1 cells. (E) RT-qPCR analysis of Pcf11 isoform expression in WT and IPA Pcf11 -KO cells. Error bars are SD on the basis of three replicates. (F) Western blot analysis of PCF11 in WT and IPA Pcf11 -KO cells. (G) 3′UTR APA changes in IPA Pcf11 -KO cells. (H) IPA changes in IPA Pcf11 -KO cells. IPA isoforms (all combined) were compared with TPA isoforms (all combined) in analysis. (I) IPA distribution maps of activated IPA events in IPA Pcf11 -KO cells. (J) Left: schematic of PASs around the transcription start site (TSS); right: metagene analysis of PAS usage around the TSS in WT and IPA Pcf1 1 -KO cells. (K) Gene expression changes in IPA Pcf11 -KO cells for different groups of genes on the basis of IPA site frequency in PolyA_DB. ***p
    Real Time Quantitative Pcr Qpcr Total Rna, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 92/100, based on 101 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Stable forced SUMO1 expression enhanced the colony formation, proliferation, migration, cell cycle progression, and invasion of A549 cells in vitro. ( a ) Detection of messenger RNA (mRNA) expression of SUMO1 in different lung cancer cell lines by quantitative real time (qRT)‐PCR. ( b ) Similar results were obtained through Western blot analysis. ( c ) qRT‐PCR analysis revealed that SUMO1 mRNA expression levels were increased in SUMO1 overexpressed A549 cells compared to control cells. ( d ) Similar results were obtained through Western blot analysis (passages 15 and 30). Upregulation of SUMO1 enhanced the ( e , f ) colony‐formation ability, ( g ) proliferation, ( h , i ) migration, and ( k , l ) invasion of A549 cells. ( j ) Forced expression of SUMO1 increased the number of A549 cells in the S phase of the cell cycle. * P

    Journal: Thoracic Cancer

    Article Title: SUMO1 promotes the proliferation and invasion of non‐small cell lung cancer cells by regulating NF‐κB

    doi: 10.1111/1759-7714.12895

    Figure Lengend Snippet: Stable forced SUMO1 expression enhanced the colony formation, proliferation, migration, cell cycle progression, and invasion of A549 cells in vitro. ( a ) Detection of messenger RNA (mRNA) expression of SUMO1 in different lung cancer cell lines by quantitative real time (qRT)‐PCR. ( b ) Similar results were obtained through Western blot analysis. ( c ) qRT‐PCR analysis revealed that SUMO1 mRNA expression levels were increased in SUMO1 overexpressed A549 cells compared to control cells. ( d ) Similar results were obtained through Western blot analysis (passages 15 and 30). Upregulation of SUMO1 enhanced the ( e , f ) colony‐formation ability, ( g ) proliferation, ( h , i ) migration, and ( k , l ) invasion of A549 cells. ( j ) Forced expression of SUMO1 increased the number of A549 cells in the S phase of the cell cycle. * P

    Article Snippet: Quantitative RT‐PCR analysis Total RNA was isolated using the TRIzol reagent according to the manufacturer's protocol (Thermo Fisher Scientific).

    Techniques: Expressing, Migration, In Vitro, Quantitative RT-PCR, Western Blot

    Downregulation of SUMO1 suppresses the proliferation, cell cycle progression, colony formation, migration, and invasion of Calu‐1 cells in vitro. ( a ) Quantitative real time‐PCR analysis revealed that the messenger RNA (mRNA) expression levels of SUMO1 in short hairpin RNA (shRNA)‐SUMO1‐1 Calu‐1 cells were significantly downregulated. ( b ) Similar results were obtained through Western blot analysis. ( c ) Downregulation of SUMO1 expression significantly inhibited the ( c ) proliferation, ( e , f ) colony‐formation ability, ( g , h ) migration, and ( i , j ) invasion of Calu‐1 cells. ( d ) Downregulation of SUMO1 reduced the number of Calu‐1 cells in the S phase of the cell cycle.

    Journal: Thoracic Cancer

    Article Title: SUMO1 promotes the proliferation and invasion of non‐small cell lung cancer cells by regulating NF‐κB

    doi: 10.1111/1759-7714.12895

    Figure Lengend Snippet: Downregulation of SUMO1 suppresses the proliferation, cell cycle progression, colony formation, migration, and invasion of Calu‐1 cells in vitro. ( a ) Quantitative real time‐PCR analysis revealed that the messenger RNA (mRNA) expression levels of SUMO1 in short hairpin RNA (shRNA)‐SUMO1‐1 Calu‐1 cells were significantly downregulated. ( b ) Similar results were obtained through Western blot analysis. ( c ) Downregulation of SUMO1 expression significantly inhibited the ( c ) proliferation, ( e , f ) colony‐formation ability, ( g , h ) migration, and ( i , j ) invasion of Calu‐1 cells. ( d ) Downregulation of SUMO1 reduced the number of Calu‐1 cells in the S phase of the cell cycle.

    Article Snippet: Quantitative RT‐PCR analysis Total RNA was isolated using the TRIzol reagent according to the manufacturer's protocol (Thermo Fisher Scientific).

    Techniques: Migration, In Vitro, Real-time Polymerase Chain Reaction, Expressing, shRNA, Western Blot

    Up-regulation of miR-96 and down-regulation of FOXO1 in tumor tissues and HepG2 cells. a miR-96 relative expression in tumor tissues and non-tumor tissues by qRT-PCR; b FOXO1 mRNA relative expression in tumor tissues and non-tumor tissues by qRT-PCR; c FOXO1 protein relative expression in tumor tissues and non-tumor tissues by Western blot; d miR-96 relative expression in L02 and HepG2 cells by qRT-PCR; e FOXO1 mRNA relative expression in L02 and HepG2 cells by qRT-PCR; f FOXO1 protein relative expression in L02 and HepG2 cells by Western blot. ** P

    Journal: Cancer Cell International

    Article Title: miR-96 exerts carcinogenic effect by activating AKT/GSK-3β/β-catenin signaling pathway through targeting inhibition of FOXO1 in hepatocellular carcinoma

    doi: 10.1186/s12935-019-0756-7

    Figure Lengend Snippet: Up-regulation of miR-96 and down-regulation of FOXO1 in tumor tissues and HepG2 cells. a miR-96 relative expression in tumor tissues and non-tumor tissues by qRT-PCR; b FOXO1 mRNA relative expression in tumor tissues and non-tumor tissues by qRT-PCR; c FOXO1 protein relative expression in tumor tissues and non-tumor tissues by Western blot; d miR-96 relative expression in L02 and HepG2 cells by qRT-PCR; e FOXO1 mRNA relative expression in L02 and HepG2 cells by qRT-PCR; f FOXO1 protein relative expression in L02 and HepG2 cells by Western blot. ** P

    Article Snippet: Quantitative real time polymerase chain reaction (qRT-PCR) Total RNA in tumor tissues and cells was obtained by Trizol kit (Thermo Fisher Scientific, Waltham, MA, USA) according to the the instruction manual.

    Techniques: Expressing, Quantitative RT-PCR, Western Blot

    FOXO1 was directly suppressed by miR-96. a Prediction of binding sites for FOXO1 and miR-96 by Target Scan; b Luciferase reporter assay; c FOXO1 mRNA expression by qRT-PCR; d FOXO1 protein expression by Western blot. ## P

    Journal: Cancer Cell International

    Article Title: miR-96 exerts carcinogenic effect by activating AKT/GSK-3β/β-catenin signaling pathway through targeting inhibition of FOXO1 in hepatocellular carcinoma

    doi: 10.1186/s12935-019-0756-7

    Figure Lengend Snippet: FOXO1 was directly suppressed by miR-96. a Prediction of binding sites for FOXO1 and miR-96 by Target Scan; b Luciferase reporter assay; c FOXO1 mRNA expression by qRT-PCR; d FOXO1 protein expression by Western blot. ## P

    Article Snippet: Quantitative real time polymerase chain reaction (qRT-PCR) Total RNA in tumor tissues and cells was obtained by Trizol kit (Thermo Fisher Scientific, Waltham, MA, USA) according to the the instruction manual.

    Techniques: Binding Assay, Luciferase, Reporter Assay, Expressing, Quantitative RT-PCR, Western Blot

    MD001 stimulates fatty acid oxidation in vitro . HepG2 ( A ), differentiated 3T3-L1 adipocyte ( B ), and differentiated C2C12 myotubes ( C ) were treated with vehicle or MD001 (10 μM) for 24 h and total RNA was isolated for cDNA synthesis. Relative gene expressions were analysed by qRT-PCR. HepG2 ( D ), differentiated 3T3-L1 adipocytes ( E ), and differentiated C2C12 myotubes ( F ) were treated with vehicle, MD001 (10 μM), or WY14643 (WY) (10 μM) for 24 h and the fatty acid oxidation rate was analysed as described in the Methods section. To confirm whether the fatty acid oxidation rate enhanced by MD001 is mediated by PPARα, HepG2 ( G ), differentiated 3T3-L1 ( H ), and differentiated C2C12 myotubes ( I ) were transfected with control or PPARα siRNA (20 nM) for 48 h, followed by treatment with vehicle, MD001 (10 μM), or WY14643 (WY) (10 μM) for 24 h. WY14643 was treated as positive control. *, **, vs. vehicle; † and ‡ , vs. vehicle/control siRNA; § , vs. WY/control siRNA; ¶ , vs. MD001/control siRNA. Data represent the mean ± SD of three independent experiments. * P

    Journal: Scientific Reports

    Article Title: MD001, a Novel Peroxisome Proliferator-activated Receptor α/γ Agonist, Improves Glucose and Lipid Metabolism

    doi: 10.1038/s41598-018-38281-0

    Figure Lengend Snippet: MD001 stimulates fatty acid oxidation in vitro . HepG2 ( A ), differentiated 3T3-L1 adipocyte ( B ), and differentiated C2C12 myotubes ( C ) were treated with vehicle or MD001 (10 μM) for 24 h and total RNA was isolated for cDNA synthesis. Relative gene expressions were analysed by qRT-PCR. HepG2 ( D ), differentiated 3T3-L1 adipocytes ( E ), and differentiated C2C12 myotubes ( F ) were treated with vehicle, MD001 (10 μM), or WY14643 (WY) (10 μM) for 24 h and the fatty acid oxidation rate was analysed as described in the Methods section. To confirm whether the fatty acid oxidation rate enhanced by MD001 is mediated by PPARα, HepG2 ( G ), differentiated 3T3-L1 ( H ), and differentiated C2C12 myotubes ( I ) were transfected with control or PPARα siRNA (20 nM) for 48 h, followed by treatment with vehicle, MD001 (10 μM), or WY14643 (WY) (10 μM) for 24 h. WY14643 was treated as positive control. *, **, vs. vehicle; † and ‡ , vs. vehicle/control siRNA; § , vs. WY/control siRNA; ¶ , vs. MD001/control siRNA. Data represent the mean ± SD of three independent experiments. * P

    Article Snippet: Complementary DNA (cDNA) was synthesised by reverse transcription using 0.5 μg of total RNA, and quantitative RT-PCR was performed using Power SYBR Green PCR Master Mix (ABI Thermo Fisher Scientific, Foster City, CA, USA) on a StepOne 48-well real-time PCR system (ABI/Thermo Fisher Scientific).

    Techniques: In Vitro, Isolation, Quantitative RT-PCR, Transfection, Positive Control

    MD001 induces the expression of target genes through PPARα and PPARγ activation. HEK293 cells were transiently co-transfected with human HA-PPARα ( A ) and HA-PPARγ ( B ) expression vectors along with the reporter plasmid (PPRE-pk-Luc) or control reporter plasmid (pk-Luc) with Renilla vector for 24 h. Cells were treated with MD001 (10 μM), rosiglitazone (rosi) (10 μM) or WY14643 (WY) (10 μM) for 24 h. Luciferase activity was normalised to Renilla luciferase activity as described in the Methods section. **, vs. vehicle. ( C ) HepG2 cells were treated with MD001 (10 μM) for 24 h, and total RNA was isolated and synthesised into cDNA. Relative expression was quantitated using qRT-PCR. **, vs. vehicle. ( D , E ) HepG2 cells were transfected with control siRNA, PPARα siRNA ( D ), or PPARγ siRNA ( E ) for 48 h and treated with vehicle, WY14643 (10 μM), rosiglitazone (10 μM) or MD001 (10 μM) for 24 h. Subsequently, cells were harvested for total RNA isolation. Relative expression of target genes was quantitated using qRT-PCR. # , vs. control siRNA; ‡ , vs. control siRNA/vehicle; § , vs. control siRNA/WY; * and **, vs. control siRNA/MD001. Data represent the mean ± SD of three independent experiments. * P

    Journal: Scientific Reports

    Article Title: MD001, a Novel Peroxisome Proliferator-activated Receptor α/γ Agonist, Improves Glucose and Lipid Metabolism

    doi: 10.1038/s41598-018-38281-0

    Figure Lengend Snippet: MD001 induces the expression of target genes through PPARα and PPARγ activation. HEK293 cells were transiently co-transfected with human HA-PPARα ( A ) and HA-PPARγ ( B ) expression vectors along with the reporter plasmid (PPRE-pk-Luc) or control reporter plasmid (pk-Luc) with Renilla vector for 24 h. Cells were treated with MD001 (10 μM), rosiglitazone (rosi) (10 μM) or WY14643 (WY) (10 μM) for 24 h. Luciferase activity was normalised to Renilla luciferase activity as described in the Methods section. **, vs. vehicle. ( C ) HepG2 cells were treated with MD001 (10 μM) for 24 h, and total RNA was isolated and synthesised into cDNA. Relative expression was quantitated using qRT-PCR. **, vs. vehicle. ( D , E ) HepG2 cells were transfected with control siRNA, PPARα siRNA ( D ), or PPARγ siRNA ( E ) for 48 h and treated with vehicle, WY14643 (10 μM), rosiglitazone (10 μM) or MD001 (10 μM) for 24 h. Subsequently, cells were harvested for total RNA isolation. Relative expression of target genes was quantitated using qRT-PCR. # , vs. control siRNA; ‡ , vs. control siRNA/vehicle; § , vs. control siRNA/WY; * and **, vs. control siRNA/MD001. Data represent the mean ± SD of three independent experiments. * P

    Article Snippet: Complementary DNA (cDNA) was synthesised by reverse transcription using 0.5 μg of total RNA, and quantitative RT-PCR was performed using Power SYBR Green PCR Master Mix (ABI Thermo Fisher Scientific, Foster City, CA, USA) on a StepOne 48-well real-time PCR system (ABI/Thermo Fisher Scientific).

    Techniques: Expressing, Activation Assay, Transfection, Plasmid Preparation, Luciferase, Activity Assay, Isolation, Quantitative RT-PCR

    MD001 promotes glucose consumption by induction of GLUT expression. HepG2 ( A ), differentiated 3T3-L1 adipocytes ( B ), and differentiated C2C12 myotubes ( C ) were treated with vehicle, MD001 (10, 50 μM), or rosiglitazone (rosi, 10 μM) for 24 h; glucose consumption was examined as described in the Methods section. Cells were harvested for total RNA isolation. Relative expression levels of GLUT2 ( D ) and GLUT4 ( E , F ) were quantitated using qRT-PCR. *, **, and † vs. vehicle. Data represent the mean ± SD of three independent experiments. * P

    Journal: Scientific Reports

    Article Title: MD001, a Novel Peroxisome Proliferator-activated Receptor α/γ Agonist, Improves Glucose and Lipid Metabolism

    doi: 10.1038/s41598-018-38281-0

    Figure Lengend Snippet: MD001 promotes glucose consumption by induction of GLUT expression. HepG2 ( A ), differentiated 3T3-L1 adipocytes ( B ), and differentiated C2C12 myotubes ( C ) were treated with vehicle, MD001 (10, 50 μM), or rosiglitazone (rosi, 10 μM) for 24 h; glucose consumption was examined as described in the Methods section. Cells were harvested for total RNA isolation. Relative expression levels of GLUT2 ( D ) and GLUT4 ( E , F ) were quantitated using qRT-PCR. *, **, and † vs. vehicle. Data represent the mean ± SD of three independent experiments. * P

    Article Snippet: Complementary DNA (cDNA) was synthesised by reverse transcription using 0.5 μg of total RNA, and quantitative RT-PCR was performed using Power SYBR Green PCR Master Mix (ABI Thermo Fisher Scientific, Foster City, CA, USA) on a StepOne 48-well real-time PCR system (ABI/Thermo Fisher Scientific).

    Techniques: Expressing, Isolation, Quantitative RT-PCR

    Pcf11 Expression Is Regulated by IPA (A) Schematic of mouse Pcf11 gene. Multiple PASs are indicated. EPA, internal exonic polyadenylation; IPA, intronic polyadenylation; TPA, 3′ terminal exon polyadenylation. 3′READS data of brain, heart, and testis are shown. (B) Features of intron 1 compared with other introns in the mouse genome. Each feature has ten bins, and the bin containing intron 1 of Pcf11 is highlighted in red. Distance to TSS is based on all IPA sites in PolyA_DB. (C) RT-qPCR analysis of IPA and TPA isoform expression in siPcf11 versus siCtrl cells. Error bars are SD on the basis of three replicates. (D) Top: schematic showing two sgRNAs used to remove a region flanking the IPA site. Bottom: validation of IPA site knockout (IPA Pcf11 -KO) by genomic DNA PCR using primers flanking the KO region. WT, wild-type 4T1 cells. (E) RT-qPCR analysis of Pcf11 isoform expression in WT and IPA Pcf11 -KO cells. Error bars are SD on the basis of three replicates. (F) Western blot analysis of PCF11 in WT and IPA Pcf11 -KO cells. (G) 3′UTR APA changes in IPA Pcf11 -KO cells. (H) IPA changes in IPA Pcf11 -KO cells. IPA isoforms (all combined) were compared with TPA isoforms (all combined) in analysis. (I) IPA distribution maps of activated IPA events in IPA Pcf11 -KO cells. (J) Left: schematic of PASs around the transcription start site (TSS); right: metagene analysis of PAS usage around the TSS in WT and IPA Pcf1 1 -KO cells. (K) Gene expression changes in IPA Pcf11 -KO cells for different groups of genes on the basis of IPA site frequency in PolyA_DB. ***p

    Journal: Cell reports

    Article Title: Regulation of Intronic Polyadenylation by PCF11 Impacts mRNA Expression of Long Genes

    doi: 10.1016/j.celrep.2019.02.049

    Figure Lengend Snippet: Pcf11 Expression Is Regulated by IPA (A) Schematic of mouse Pcf11 gene. Multiple PASs are indicated. EPA, internal exonic polyadenylation; IPA, intronic polyadenylation; TPA, 3′ terminal exon polyadenylation. 3′READS data of brain, heart, and testis are shown. (B) Features of intron 1 compared with other introns in the mouse genome. Each feature has ten bins, and the bin containing intron 1 of Pcf11 is highlighted in red. Distance to TSS is based on all IPA sites in PolyA_DB. (C) RT-qPCR analysis of IPA and TPA isoform expression in siPcf11 versus siCtrl cells. Error bars are SD on the basis of three replicates. (D) Top: schematic showing two sgRNAs used to remove a region flanking the IPA site. Bottom: validation of IPA site knockout (IPA Pcf11 -KO) by genomic DNA PCR using primers flanking the KO region. WT, wild-type 4T1 cells. (E) RT-qPCR analysis of Pcf11 isoform expression in WT and IPA Pcf11 -KO cells. Error bars are SD on the basis of three replicates. (F) Western blot analysis of PCF11 in WT and IPA Pcf11 -KO cells. (G) 3′UTR APA changes in IPA Pcf11 -KO cells. (H) IPA changes in IPA Pcf11 -KO cells. IPA isoforms (all combined) were compared with TPA isoforms (all combined) in analysis. (I) IPA distribution maps of activated IPA events in IPA Pcf11 -KO cells. (J) Left: schematic of PASs around the transcription start site (TSS); right: metagene analysis of PAS usage around the TSS in WT and IPA Pcf1 1 -KO cells. (K) Gene expression changes in IPA Pcf11 -KO cells for different groups of genes on the basis of IPA site frequency in PolyA_DB. ***p

    Article Snippet: Real-time quantitative PCR (qPCR) Total RNA was extracted using TRIzol (Thermo Fisher) and treated with Turbo DNase (Thermo Fisher). cDNA was synthesized using oligo(dT) and M-MLV reverse transcriptase with 1.5–2 μg of RNA. cDNA was then mixed with real-time PCR primers and Luna qPCR master mix (NEB).

    Techniques: Expressing, Indirect Immunoperoxidase Assay, Quantitative RT-PCR, Knock-Out, Polymerase Chain Reaction, Western Blot