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Promega polymerase chain reaction pcr assays
Polymerase Chain Reaction Pcr Assays, supplied by Promega, used in various techniques. Bioz Stars score: 87/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 87 stars, based on 3 article reviews
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polymerase chain reaction pcr assays - by Bioz Stars, 2020-04
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DNA Extraction:

Article Title: Comparative Genomics of Sibling Fungal Pathogenic Taxa Identifies Adaptive Evolution without Divergence in Pathogenicity Genes or Genomic Structure
Article Snippet: DNA extraction was performed by using the Dneasy Plant Mini Kit (QIAGEN, Valencia, CA) on lyophilized fungal cultures that were grown at room temperature for 10 days in a 2% (w/v) malt extract liquid medium (AppliChem GmbH, Darmstadt, Germany). .. Polymerase chain reaction (PCR) assays were performed in a 25 μl volume containing 5× PCR buffer, 1.5 mM of MgCl2 , 0.2 mM of dNTPs mix, 0.5 μM each of the primers, 0.025 U of GoTaq polymerase (Promega, Madison, WI), and 0.1 ng of genomic DNA.

Article Title: Triatoma maculata, the Vector of Trypanosoma cruzi, in Venezuela. Phenotypic and Genotypic Variability as Potential Indicator of Vector Displacement into the Domestic Habitat
Article Snippet: Paragraph title: DNA extraction and molecular marker amplification ... Markers were amplified under standard polymerase chain reaction (PCR) assays as described previously; the reaction was carried out in a final volume of 25 μl containing 12.5 μl cocktail of PCR mix 2X (GoTaq Master Mix, Promega, Madison, WI, USA.

Marker:

Article Title: Triatoma maculata, the Vector of Trypanosoma cruzi, in Venezuela. Phenotypic and Genotypic Variability as Potential Indicator of Vector Displacement into the Domestic Habitat
Article Snippet: Paragraph title: DNA extraction and molecular marker amplification ... Markers were amplified under standard polymerase chain reaction (PCR) assays as described previously; the reaction was carried out in a final volume of 25 μl containing 12.5 μl cocktail of PCR mix 2X (GoTaq Master Mix, Promega, Madison, WI, USA.

Functional Assay:

Article Title: Comparative Genomics of Sibling Fungal Pathogenic Taxa Identifies Adaptive Evolution without Divergence in Pathogenicity Genes or Genomic Structure
Article Snippet: Polymerase chain reaction (PCR) assays were performed in a 25 μl volume containing 5× PCR buffer, 1.5 mM of MgCl2 , 0.2 mM of dNTPs mix, 0.5 μM each of the primers, 0.025 U of GoTaq polymerase (Promega, Madison, WI), and 0.1 ng of genomic DNA. .. The PCR products were sequenced in both directions by the Functional Genomics Laboratory of the University of Berkeley (CA).

Amplification:

Article Title: The Endocannabinoid System in the Baboon (Papio SPP.) as a Complex Framework for Developmental Pharmacology
Article Snippet: Polymerase chain reaction (PCR) assays were carried out in a 25-μl total volume using 1 μl of each primer (10 pM), 5 μl of the RT reaction and 2X PCR Master Mix from Promega (San Luis Obispo, CA, USA). .. Amplification was carried out in a Veriti thermocycler (Applied Biosystems, Foster City, CA, USA) under the conditions described in .

Article Title: Comparative Genomics of Sibling Fungal Pathogenic Taxa Identifies Adaptive Evolution without Divergence in Pathogenicity Genes or Genomic Structure
Article Snippet: To confirm putative H. annosum SNPs/InDels, four loci involved in sabrobic growth, four involved in sporulation, and two related to pathogenicity ( ) were randomly selected to be amplified, sequenced and compared on five additional H. irregulare genotypes and five H. annosum genotypes (listed in supplementary table S1 , Supplementary Material online). .. Polymerase chain reaction (PCR) assays were performed in a 25 μl volume containing 5× PCR buffer, 1.5 mM of MgCl2 , 0.2 mM of dNTPs mix, 0.5 μM each of the primers, 0.025 U of GoTaq polymerase (Promega, Madison, WI), and 0.1 ng of genomic DNA.

Article Title: Triatoma maculata, the Vector of Trypanosoma cruzi, in Venezuela. Phenotypic and Genotypic Variability as Potential Indicator of Vector Displacement into the Domestic Habitat
Article Snippet: .. Markers were amplified under standard polymerase chain reaction (PCR) assays as described previously; the reaction was carried out in a final volume of 25 μl containing 12.5 μl cocktail of PCR mix 2X (GoTaq Master Mix, Promega, Madison, WI, USA. .. Cat.# M7122), 0.4 μmol primers (stock 100 μM) and 5 ng total genomic DNA; the PCR reaction was performed in an MJ Research PTC-200 thermocycler.

Polymerase Chain Reaction:

Article Title: The Endocannabinoid System in the Baboon (Papio SPP.) as a Complex Framework for Developmental Pharmacology
Article Snippet: .. Polymerase chain reaction (PCR) assays were carried out in a 25-μl total volume using 1 μl of each primer (10 pM), 5 μl of the RT reaction and 2X PCR Master Mix from Promega (San Luis Obispo, CA, USA). .. Amplification was carried out in a Veriti thermocycler (Applied Biosystems, Foster City, CA, USA) under the conditions described in .

Article Title: Comparative Genomics of Sibling Fungal Pathogenic Taxa Identifies Adaptive Evolution without Divergence in Pathogenicity Genes or Genomic Structure
Article Snippet: .. Polymerase chain reaction (PCR) assays were performed in a 25 μl volume containing 5× PCR buffer, 1.5 mM of MgCl2 , 0.2 mM of dNTPs mix, 0.5 μM each of the primers, 0.025 U of GoTaq polymerase (Promega, Madison, WI), and 0.1 ng of genomic DNA. .. PCR reactions were conducted as follows: an initial cycle with a 96 °C denaturation step of 5 min, followed by 35 cycles, with each cycle consisting of a 96 °C denaturation step of 30 s, an annealing step of 30 s and a 72 °C extension step of 45 s, and one final cycle with a 72 °C extension step of 10 min. Amplicons were digested by ExoSAP-IT (Affymetrix, Santa Clara, CA) at 37 °C for 15 min and then at 80 °C for 15 min.

Article Title: Triatoma maculata, the Vector of Trypanosoma cruzi, in Venezuela. Phenotypic and Genotypic Variability as Potential Indicator of Vector Displacement into the Domestic Habitat
Article Snippet: .. Markers were amplified under standard polymerase chain reaction (PCR) assays as described previously; the reaction was carried out in a final volume of 25 μl containing 12.5 μl cocktail of PCR mix 2X (GoTaq Master Mix, Promega, Madison, WI, USA. .. Cat.# M7122), 0.4 μmol primers (stock 100 μM) and 5 ng total genomic DNA; the PCR reaction was performed in an MJ Research PTC-200 thermocycler.

Article Title: Nitric oxide modulates lipopolysaccharide-induced endothelial platelet endothelial cell adhesion molecule expression via interleukin-10
Article Snippet: .. Trizol reagent was purchased from Invitrogen (Grand Island, NY, USA), and the other reagents used in polymerase chain reaction (PCR) assays were purchased from Promega (Madison, WI, USA). .. SYBR Advantage qPCR Premix was obtained from Clontech (Mountain View, CA, USA).

Isolation:

Article Title: Triatoma maculata, the Vector of Trypanosoma cruzi, in Venezuela. Phenotypic and Genotypic Variability as Potential Indicator of Vector Displacement into the Domestic Habitat
Article Snippet: For direct molecular identification of T. cruzi and T. rangeli , DNA isolated from the intestinal content of insects was used to amplify the variable region of the minicircles of kinetoplast DNA (kDNA) and the non-transcribed spacer region of the mini-exon ( , ). .. Markers were amplified under standard polymerase chain reaction (PCR) assays as described previously; the reaction was carried out in a final volume of 25 μl containing 12.5 μl cocktail of PCR mix 2X (GoTaq Master Mix, Promega, Madison, WI, USA.

Multiplex Assay:

Article Title: Triatoma maculata, the Vector of Trypanosoma cruzi, in Venezuela. Phenotypic and Genotypic Variability as Potential Indicator of Vector Displacement into the Domestic Habitat
Article Snippet: The β-tubulin primers were designed by multiplex alignment of similar genes available in the NCBI Genbank from Triatoma tibiamaculata (KC249297), T. infestans (JK33877), T. braziliensis (EC917343), Rhodnius prolixus (FG544591), Aedes aegypti (XM00165064), and Drosophila melanogaster (AE0135994), and after bioinformatics sequence analysis, the primers sequences TubTmf and TubTmr were selected (Table ). .. Markers were amplified under standard polymerase chain reaction (PCR) assays as described previously; the reaction was carried out in a final volume of 25 μl containing 12.5 μl cocktail of PCR mix 2X (GoTaq Master Mix, Promega, Madison, WI, USA.

Real-time Polymerase Chain Reaction:

Article Title: Nitric oxide modulates lipopolysaccharide-induced endothelial platelet endothelial cell adhesion molecule expression via interleukin-10
Article Snippet: Trizol reagent was purchased from Invitrogen (Grand Island, NY, USA), and the other reagents used in polymerase chain reaction (PCR) assays were purchased from Promega (Madison, WI, USA). .. SYBR Advantage qPCR Premix was obtained from Clontech (Mountain View, CA, USA).

Sequencing:

Article Title: Triatoma maculata, the Vector of Trypanosoma cruzi, in Venezuela. Phenotypic and Genotypic Variability as Potential Indicator of Vector Displacement into the Domestic Habitat
Article Snippet: The β-tubulin primers were designed by multiplex alignment of similar genes available in the NCBI Genbank from Triatoma tibiamaculata (KC249297), T. infestans (JK33877), T. braziliensis (EC917343), Rhodnius prolixus (FG544591), Aedes aegypti (XM00165064), and Drosophila melanogaster (AE0135994), and after bioinformatics sequence analysis, the primers sequences TubTmf and TubTmr were selected (Table ). .. Markers were amplified under standard polymerase chain reaction (PCR) assays as described previously; the reaction was carried out in a final volume of 25 μl containing 12.5 μl cocktail of PCR mix 2X (GoTaq Master Mix, Promega, Madison, WI, USA.

Activity Assay:

Article Title: Nitric oxide modulates lipopolysaccharide-induced endothelial platelet endothelial cell adhesion molecule expression via interleukin-10
Article Snippet: Antibodies against adhesion molecules and anti-IL-10, as well as cytokine detection kits, were purchased from BD Pharmingen (San Diego, CA, USA). l -NAME, LPS ( Escherichia coli ; serotype 026 : B6), Percoll gradients and all reagents used to measure NOS activity and NO levels were obtained from Sigma (St Louis, MO, USA). .. Trizol reagent was purchased from Invitrogen (Grand Island, NY, USA), and the other reagents used in polymerase chain reaction (PCR) assays were purchased from Promega (Madison, WI, USA).

Modification:

Article Title: Nitric oxide modulates lipopolysaccharide-induced endothelial platelet endothelial cell adhesion molecule expression via interleukin-10
Article Snippet: Trizol reagent was purchased from Invitrogen (Grand Island, NY, USA), and the other reagents used in polymerase chain reaction (PCR) assays were purchased from Promega (Madison, WI, USA). .. Calf bovine serum, Dulbecco's modified Eagle's medium (DMEM) and gentamycin were purchased from G ibco (Carlsbad, CA, USA).

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  • rna  (Promega)
    98
    Promega rna
    FACT Depletion Directly and Indirectly Primes the Transcriptome for Pluripotency (A) Human fibroblasts were used for ChIP-seq analysis or transfected with siRNAs against human FACT subunits and used for ATAC-seq and <t>RNA-seq</t> analysis 48 hr after knockdown without <t>OSKM</t> induction. (B) Correlation of SSRP1 and SUPT16H log 2 -ChIP versus input ratios for TSS windows classified as high for at least one of the two FACT components (see STAR Methods ). Density scale reflects number of TSS windows plotted per hexbin. The Spearman correlation Rho value is given. (C) Correlation of SSRP1 and SUPT16H knockdown RNA-seq log 2 -fold changes for genes that were significantly changed in at least one of the conditions (see STAR Methods ; FDR 0.1). Density scale reflects number of genes plotted per hexbin. The Spearman correlation Rho value is given. (D) Hexbin density scatterplot of SUPT16H knockdown normalized ATAC-seq log 2 -fold changes plotted against SSRP1 knockdown normalized ATAC-seq log 2 -fold changes for ATAC-seq peaks that were significantly differential (FDR 0.01) in at least one of the conditions. Scale shows number of ATAC-seq peaks plotted per hexbin. The Spearman correlation Rho value is given. (E) Log 2 ratio of SUPT16H ChIP versus Input plotted against Fragments Per Kilobase of transcript per Million mapped reads (FPKM) expression values from control RNA-seq for genes classified as FACT high and detected in the RNA-seq. Density scatterplot scale shows number of genes plotted per hexbin. (F) Log 2 -fold changes in expression levels from RNA-seq after SUPT16H knockdown plotted against log 2 ratio of SUPT16H ChIP versus Input for genes classified as FACT high and detected in the RNA-seq. Density scatterplot scale shows number of genes plotted per hexbin. (G) Average positional profiles and heatmaps of library- and input-normalized SUPT16H ChIP-seq signal for genes whose TSS windows classified as FACT high or FACT low (see STAR Methods ). (H) Violin plots of RNA-seq log 2 -fold change distributions following FACT knockdown for genes classified as being FACT high or FACT low. Red jitter dots reflect genes whose expression changed significantly upon FACT knockdown (FDR
    Rna, supplied by Promega, used in various techniques. Bioz Stars score: 98/100, based on 205 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    Promega real time quantitative pcr
    Upregulation of IGF1 , ANG1 , and CSGALNACT1 expression by <t>NTP</t> treatment. Based on the results of the microarray, relative mRNA expression of IGF1 , ANG1 , and CSGALNACT1 was assessed by real-time quantitative <t>PCR.</t> Samples were prepared form one-week-cultured cells, treated as indicated. A-C) Addition of AsAP with or without NTP upregulated expression of these three genes in comparison with control samples. A) The co-treatment with NTP (0.1 or 1.0 mNU/ml) and AsAP tended to increase IGF1 compared to AsAP alone. B) The co-treatment with NTP (0.1 or 1.0 mNU/ml) and AsAP increased ANG1 significantly compared to AsAP alone. C) The co-treatment with NTP (0.1 mNU/ml) and AsAP increased CSGALNACT1 significantly compared to AsAP alone. Data are normalized to G APDH and presented as the average ± standard error (** p
    Real Time Quantitative Pcr, supplied by Promega, used in various techniques. Bioz Stars score: 95/100, based on 54 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Promega quantitative reverse transcription polymerase chain reaction qrt pcr analysis
    Expression analysis of MMP13, miR-140-5p, NF-κB, NF-κB2, Smad3, and TGF-β3 in IL-1β-stimulated MCCs. (A) Assessment of MMP13 mRNA relative expression by <t>qRT-PCR,</t> (B) Assessment of miR-140-5p relative gene expression by qRT-PCR, and (C) Evaluation of NF-κB, NF-κB2, Smad3, TGF-β3 protein expression. β-actin and small nuclear RNA U6 were used as controls. Each group repeated three times ( n = 3). Control group (Con) were without IL-1β stimulation. ∗ P
    Quantitative Reverse Transcription Polymerase Chain Reaction Qrt Pcr Analysis, supplied by Promega, 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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    92
    Promega proteasome assay proteasome activity
    Effect of LAMP-2C expression on CMA substrates. (A) Cellular levels of CMA substrates Chk1, IκBα, and p-IκBα in DM331 pCMV and DM331 2C myc cells were examined by western blotting. Relative protein levels were calculated by setting the normalized expression to one for DM331 pCMV cells. (B) mRNA levels of CHEK1 and NFKBIA transcripts were analyzed by qPCR and normalized to ACTB expression. mRNA levels in DM331 pCMV cells were normalized and set to one. (C) <t>Proteasome</t> activity was measured using the Proteasome-Glo Chymotrypsin-Like Cell-Based Assay. Cells were incubated with a substrate succinyl-LLVY-aminoluciferin which penetrates into the cytoplasm. This substrate is cleaved by the proteasome to release aminoluciferin which is released from cells. Luciferase is added to these cells, cleaving aminoluciferin to a luminescent product detectable using a luminometer. Data were analyzed by two-way ANOVA or by two-tailed, unpaired Student’s t -test. ∗ p
    Proteasome Assay Proteasome Activity, supplied by Promega, used in various techniques. Bioz Stars score: 92/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    FACT Depletion Directly and Indirectly Primes the Transcriptome for Pluripotency (A) Human fibroblasts were used for ChIP-seq analysis or transfected with siRNAs against human FACT subunits and used for ATAC-seq and RNA-seq analysis 48 hr after knockdown without OSKM induction. (B) Correlation of SSRP1 and SUPT16H log 2 -ChIP versus input ratios for TSS windows classified as high for at least one of the two FACT components (see STAR Methods ). Density scale reflects number of TSS windows plotted per hexbin. The Spearman correlation Rho value is given. (C) Correlation of SSRP1 and SUPT16H knockdown RNA-seq log 2 -fold changes for genes that were significantly changed in at least one of the conditions (see STAR Methods ; FDR 0.1). Density scale reflects number of genes plotted per hexbin. The Spearman correlation Rho value is given. (D) Hexbin density scatterplot of SUPT16H knockdown normalized ATAC-seq log 2 -fold changes plotted against SSRP1 knockdown normalized ATAC-seq log 2 -fold changes for ATAC-seq peaks that were significantly differential (FDR 0.01) in at least one of the conditions. Scale shows number of ATAC-seq peaks plotted per hexbin. The Spearman correlation Rho value is given. (E) Log 2 ratio of SUPT16H ChIP versus Input plotted against Fragments Per Kilobase of transcript per Million mapped reads (FPKM) expression values from control RNA-seq for genes classified as FACT high and detected in the RNA-seq. Density scatterplot scale shows number of genes plotted per hexbin. (F) Log 2 -fold changes in expression levels from RNA-seq after SUPT16H knockdown plotted against log 2 ratio of SUPT16H ChIP versus Input for genes classified as FACT high and detected in the RNA-seq. Density scatterplot scale shows number of genes plotted per hexbin. (G) Average positional profiles and heatmaps of library- and input-normalized SUPT16H ChIP-seq signal for genes whose TSS windows classified as FACT high or FACT low (see STAR Methods ). (H) Violin plots of RNA-seq log 2 -fold change distributions following FACT knockdown for genes classified as being FACT high or FACT low. Red jitter dots reflect genes whose expression changed significantly upon FACT knockdown (FDR

    Journal: Developmental Cell

    Article Title: FACT Sets a Barrier for Cell Fate Reprogramming in Caenorhabditis elegans and Human Cells

    doi: 10.1016/j.devcel.2018.07.006

    Figure Lengend Snippet: FACT Depletion Directly and Indirectly Primes the Transcriptome for Pluripotency (A) Human fibroblasts were used for ChIP-seq analysis or transfected with siRNAs against human FACT subunits and used for ATAC-seq and RNA-seq analysis 48 hr after knockdown without OSKM induction. (B) Correlation of SSRP1 and SUPT16H log 2 -ChIP versus input ratios for TSS windows classified as high for at least one of the two FACT components (see STAR Methods ). Density scale reflects number of TSS windows plotted per hexbin. The Spearman correlation Rho value is given. (C) Correlation of SSRP1 and SUPT16H knockdown RNA-seq log 2 -fold changes for genes that were significantly changed in at least one of the conditions (see STAR Methods ; FDR 0.1). Density scale reflects number of genes plotted per hexbin. The Spearman correlation Rho value is given. (D) Hexbin density scatterplot of SUPT16H knockdown normalized ATAC-seq log 2 -fold changes plotted against SSRP1 knockdown normalized ATAC-seq log 2 -fold changes for ATAC-seq peaks that were significantly differential (FDR 0.01) in at least one of the conditions. Scale shows number of ATAC-seq peaks plotted per hexbin. The Spearman correlation Rho value is given. (E) Log 2 ratio of SUPT16H ChIP versus Input plotted against Fragments Per Kilobase of transcript per Million mapped reads (FPKM) expression values from control RNA-seq for genes classified as FACT high and detected in the RNA-seq. Density scatterplot scale shows number of genes plotted per hexbin. (F) Log 2 -fold changes in expression levels from RNA-seq after SUPT16H knockdown plotted against log 2 ratio of SUPT16H ChIP versus Input for genes classified as FACT high and detected in the RNA-seq. Density scatterplot scale shows number of genes plotted per hexbin. (G) Average positional profiles and heatmaps of library- and input-normalized SUPT16H ChIP-seq signal for genes whose TSS windows classified as FACT high or FACT low (see STAR Methods ). (H) Violin plots of RNA-seq log 2 -fold change distributions following FACT knockdown for genes classified as being FACT high or FACT low. Red jitter dots reflect genes whose expression changed significantly upon FACT knockdown (FDR

    Article Snippet: To control for OSKM expression after SSRP1 and SUPT16H depletion, RNA was isolated from siRNA transfected cells 48h after knockdown without prior Doxycycline treatment. cDNA was synthesized with oligo dT primers using the GoScript Reverse Transcriptase (Promega). cDNA was used for qPCR with the Maxima SYBR Green qPCR Master Mix (2X) (Thermo Scientific) according to the manufacturer’s instructions on an ABI PRISM 7700 system (Applied Biosystems).

    Techniques: Chromatin Immunoprecipitation, Transfection, RNA Sequencing Assay, Expressing

    Upregulation of IGF1 , ANG1 , and CSGALNACT1 expression by NTP treatment. Based on the results of the microarray, relative mRNA expression of IGF1 , ANG1 , and CSGALNACT1 was assessed by real-time quantitative PCR. Samples were prepared form one-week-cultured cells, treated as indicated. A-C) Addition of AsAP with or without NTP upregulated expression of these three genes in comparison with control samples. A) The co-treatment with NTP (0.1 or 1.0 mNU/ml) and AsAP tended to increase IGF1 compared to AsAP alone. B) The co-treatment with NTP (0.1 or 1.0 mNU/ml) and AsAP increased ANG1 significantly compared to AsAP alone. C) The co-treatment with NTP (0.1 mNU/ml) and AsAP increased CSGALNACT1 significantly compared to AsAP alone. Data are normalized to G APDH and presented as the average ± standard error (** p

    Journal: PLoS ONE

    Article Title: Upregulation of glycosaminoglycan synthesis by Neurotropin in nucleus pulposus cells via stimulation of chondroitin sulfate N-acetylgalactosaminyltransferase 1: A new approach to attenuation of intervertebral disc degeneration

    doi: 10.1371/journal.pone.0202640

    Figure Lengend Snippet: Upregulation of IGF1 , ANG1 , and CSGALNACT1 expression by NTP treatment. Based on the results of the microarray, relative mRNA expression of IGF1 , ANG1 , and CSGALNACT1 was assessed by real-time quantitative PCR. Samples were prepared form one-week-cultured cells, treated as indicated. A-C) Addition of AsAP with or without NTP upregulated expression of these three genes in comparison with control samples. A) The co-treatment with NTP (0.1 or 1.0 mNU/ml) and AsAP tended to increase IGF1 compared to AsAP alone. B) The co-treatment with NTP (0.1 or 1.0 mNU/ml) and AsAP increased ANG1 significantly compared to AsAP alone. C) The co-treatment with NTP (0.1 mNU/ml) and AsAP increased CSGALNACT1 significantly compared to AsAP alone. Data are normalized to G APDH and presented as the average ± standard error (** p

    Article Snippet: Real-time quantitative PCR After a week in culture with NTP, cells were harvested (N = 5) homogenized in lysis buffer, and total RNA was prepared using an SV Total RNA Isolation System (Z3100, Promega, Madison, WI).

    Techniques: Expressing, Microarray, Real-time Polymerase Chain Reaction, Cell Culture

    Effect of Neurotropin (NTP) on production of sulfated glycosaminoglycans. A) After two weeks of culture, glycosaminoglycan (GAG) content per plate increased in cultures with added AsAP, with or without the addition of NTP. B) DNA content increased in cultures with added AsAP, with or without the addition of NTP. C) GAG content normalized to DNA content was significantly upregulated by addition of 0.1 mNU/ml NTP with AsAP. D) After one week of culture, the effect of NTP on aggrecan ( ACAN ) mRNA expression was measured by real-time quantitative PCR. Gene expression of ACAN normalized to GAPDH showed a tendency to increase with increasing concentrations of NTP. Data are presented as the average ± standard error (** p

    Journal: PLoS ONE

    Article Title: Upregulation of glycosaminoglycan synthesis by Neurotropin in nucleus pulposus cells via stimulation of chondroitin sulfate N-acetylgalactosaminyltransferase 1: A new approach to attenuation of intervertebral disc degeneration

    doi: 10.1371/journal.pone.0202640

    Figure Lengend Snippet: Effect of Neurotropin (NTP) on production of sulfated glycosaminoglycans. A) After two weeks of culture, glycosaminoglycan (GAG) content per plate increased in cultures with added AsAP, with or without the addition of NTP. B) DNA content increased in cultures with added AsAP, with or without the addition of NTP. C) GAG content normalized to DNA content was significantly upregulated by addition of 0.1 mNU/ml NTP with AsAP. D) After one week of culture, the effect of NTP on aggrecan ( ACAN ) mRNA expression was measured by real-time quantitative PCR. Gene expression of ACAN normalized to GAPDH showed a tendency to increase with increasing concentrations of NTP. Data are presented as the average ± standard error (** p

    Article Snippet: Real-time quantitative PCR After a week in culture with NTP, cells were harvested (N = 5) homogenized in lysis buffer, and total RNA was prepared using an SV Total RNA Isolation System (Z3100, Promega, Madison, WI).

    Techniques: Expressing, Real-time Polymerase Chain Reaction

    Expression analysis of MMP13, miR-140-5p, NF-κB, NF-κB2, Smad3, and TGF-β3 in IL-1β-stimulated MCCs. (A) Assessment of MMP13 mRNA relative expression by qRT-PCR, (B) Assessment of miR-140-5p relative gene expression by qRT-PCR, and (C) Evaluation of NF-κB, NF-κB2, Smad3, TGF-β3 protein expression. β-actin and small nuclear RNA U6 were used as controls. Each group repeated three times ( n = 3). Control group (Con) were without IL-1β stimulation. ∗ P

    Journal: Frontiers in Pharmacology

    Article Title: Potential Novel Prediction of TMJ-OA: MiR-140-5p Regulates Inflammation Through Smad/TGF-β Signaling

    doi: 10.3389/fphar.2019.00015

    Figure Lengend Snippet: Expression analysis of MMP13, miR-140-5p, NF-κB, NF-κB2, Smad3, and TGF-β3 in IL-1β-stimulated MCCs. (A) Assessment of MMP13 mRNA relative expression by qRT-PCR, (B) Assessment of miR-140-5p relative gene expression by qRT-PCR, and (C) Evaluation of NF-κB, NF-κB2, Smad3, TGF-β3 protein expression. β-actin and small nuclear RNA U6 were used as controls. Each group repeated three times ( n = 3). Control group (Con) were without IL-1β stimulation. ∗ P

    Article Snippet: Quantitative Reverse-Transcription Polymerase Chain Reaction (qRT-PCR) Analysis Following to the manufacturer’s instructions, 1 μg for total RNA were extracted by Eastep® Super Total RNA Extraction Kit (Promega, Madison, WI, United States) then reversed transcribed into cDNA by A5000 GoScriptTM Reverse Transcription System (Promega, Madison, WI, United States).

    Techniques: Expressing, Quantitative RT-PCR

    Effect of LAMP-2C expression on CMA substrates. (A) Cellular levels of CMA substrates Chk1, IκBα, and p-IκBα in DM331 pCMV and DM331 2C myc cells were examined by western blotting. Relative protein levels were calculated by setting the normalized expression to one for DM331 pCMV cells. (B) mRNA levels of CHEK1 and NFKBIA transcripts were analyzed by qPCR and normalized to ACTB expression. mRNA levels in DM331 pCMV cells were normalized and set to one. (C) Proteasome activity was measured using the Proteasome-Glo Chymotrypsin-Like Cell-Based Assay. Cells were incubated with a substrate succinyl-LLVY-aminoluciferin which penetrates into the cytoplasm. This substrate is cleaved by the proteasome to release aminoluciferin which is released from cells. Luciferase is added to these cells, cleaving aminoluciferin to a luminescent product detectable using a luminometer. Data were analyzed by two-way ANOVA or by two-tailed, unpaired Student’s t -test. ∗ p

    Journal: Frontiers in Cell and Developmental Biology

    Article Title: Melanoma LAMP-2C Modulates Tumor Growth and Autophagy

    doi: 10.3389/fcell.2018.00101

    Figure Lengend Snippet: Effect of LAMP-2C expression on CMA substrates. (A) Cellular levels of CMA substrates Chk1, IκBα, and p-IκBα in DM331 pCMV and DM331 2C myc cells were examined by western blotting. Relative protein levels were calculated by setting the normalized expression to one for DM331 pCMV cells. (B) mRNA levels of CHEK1 and NFKBIA transcripts were analyzed by qPCR and normalized to ACTB expression. mRNA levels in DM331 pCMV cells were normalized and set to one. (C) Proteasome activity was measured using the Proteasome-Glo Chymotrypsin-Like Cell-Based Assay. Cells were incubated with a substrate succinyl-LLVY-aminoluciferin which penetrates into the cytoplasm. This substrate is cleaved by the proteasome to release aminoluciferin which is released from cells. Luciferase is added to these cells, cleaving aminoluciferin to a luminescent product detectable using a luminometer. Data were analyzed by two-way ANOVA or by two-tailed, unpaired Student’s t -test. ∗ p

    Article Snippet: Proteasome Assay Proteasome activity was determined using Proteasome-Glo Chymotrypsin-Like Cell-Based Assay (Promega, Madison, WI, United States) ( ).

    Techniques: Expressing, Western Blot, Real-time Polymerase Chain Reaction, Activity Assay, Cell Based Assay, Incubation, Luciferase, Two Tailed Test