pcr mix  (Roche)


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    Structured Review

    Roche pcr mix
    Overview of single-cell RNA-sequencing in Smart-seq3. (a) Library strategy for Smart-seq3. PolyA+ RNA molecules are reverse transcribed and template switching is carried out at the 5’ end. After <t>PCR</t> <t>preamplification,</t> tagmentation via Tn5 introduces near-random cuts in the cDNA, producing 5’ UMI-tagged fragments and internal fragments spanning the whole gene body. ( b ) Gene body coverage averaged over HEK293FT (n = 96) cells sequenced with the Smart-seq3 protocol. Shown is the mean coverage of UMI reads (green) and internal reads (blue) shaded by the standard deviation. ( c ) Effect of tagmentation conditions on the fraction of UMI-containing reads (16 HEK293FT cells per condition). Left panel: varying Tn5 with constant 200 pg cDNA input. Right panel: varying cDNA input with constant 0.5ul Tn5. ( d ) Gene detection sensitivity for Smart-seq2 (44 cells) and Smart-seq3 (88 cells), downsampled to 1 million raw reads per HEK293FT cell. Shown are number of genes detected over 0 or 1 RPKM. P-value was computed as a two-sided t -test. ( e ) Reproducibility in gene expression quantification across HEKF293FT cells for Smart-seq2 (44 cells) and Smart-seq3 (88 cells) at RPKM and UMI level. Shown are adjusted r^2 for all pairwise cell to cell linear model fits in libraries downsampled to 1 million reads per cell. ( f ) Sensitivity to detect RNA molecules in Smart-seq3 shown by summarizing the number of unique error-corrected UMI sequences and genes detected per HEK293FT cell. Colors indicate the per cell downsampling depth ranging from 10.000 (n = 24 cells) to 750.000 (n = 16 cells) UMI-containing sequencing reads. ( g ) Violin plots summarizing the number of molecules detected per cell with Smart-seq2-UMI, Smart-seq3 and using smRNA-FISH for four X chromosomal genes (Hdac6, Igbp1, Mpp1 and Msl3). ( h ) Estimating the percent of smRNA-FISH molecules that were detected in cells using Smart-seq2-UMI and Smart-seq3. Shown are means and 95% confidence intervals.
    Pcr Mix, supplied by Roche, used in various techniques. Bioz Stars score: 94/100, based on 215 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Images

    1) Product Images from "Single-cell RNA counting at allele- and isoform-resolution using Smart-seq3"

    Article Title: Single-cell RNA counting at allele- and isoform-resolution using Smart-seq3

    Journal: bioRxiv

    doi: 10.1101/817924

    Overview of single-cell RNA-sequencing in Smart-seq3. (a) Library strategy for Smart-seq3. PolyA+ RNA molecules are reverse transcribed and template switching is carried out at the 5’ end. After PCR preamplification, tagmentation via Tn5 introduces near-random cuts in the cDNA, producing 5’ UMI-tagged fragments and internal fragments spanning the whole gene body. ( b ) Gene body coverage averaged over HEK293FT (n = 96) cells sequenced with the Smart-seq3 protocol. Shown is the mean coverage of UMI reads (green) and internal reads (blue) shaded by the standard deviation. ( c ) Effect of tagmentation conditions on the fraction of UMI-containing reads (16 HEK293FT cells per condition). Left panel: varying Tn5 with constant 200 pg cDNA input. Right panel: varying cDNA input with constant 0.5ul Tn5. ( d ) Gene detection sensitivity for Smart-seq2 (44 cells) and Smart-seq3 (88 cells), downsampled to 1 million raw reads per HEK293FT cell. Shown are number of genes detected over 0 or 1 RPKM. P-value was computed as a two-sided t -test. ( e ) Reproducibility in gene expression quantification across HEKF293FT cells for Smart-seq2 (44 cells) and Smart-seq3 (88 cells) at RPKM and UMI level. Shown are adjusted r^2 for all pairwise cell to cell linear model fits in libraries downsampled to 1 million reads per cell. ( f ) Sensitivity to detect RNA molecules in Smart-seq3 shown by summarizing the number of unique error-corrected UMI sequences and genes detected per HEK293FT cell. Colors indicate the per cell downsampling depth ranging from 10.000 (n = 24 cells) to 750.000 (n = 16 cells) UMI-containing sequencing reads. ( g ) Violin plots summarizing the number of molecules detected per cell with Smart-seq2-UMI, Smart-seq3 and using smRNA-FISH for four X chromosomal genes (Hdac6, Igbp1, Mpp1 and Msl3). ( h ) Estimating the percent of smRNA-FISH molecules that were detected in cells using Smart-seq2-UMI and Smart-seq3. Shown are means and 95% confidence intervals.
    Figure Legend Snippet: Overview of single-cell RNA-sequencing in Smart-seq3. (a) Library strategy for Smart-seq3. PolyA+ RNA molecules are reverse transcribed and template switching is carried out at the 5’ end. After PCR preamplification, tagmentation via Tn5 introduces near-random cuts in the cDNA, producing 5’ UMI-tagged fragments and internal fragments spanning the whole gene body. ( b ) Gene body coverage averaged over HEK293FT (n = 96) cells sequenced with the Smart-seq3 protocol. Shown is the mean coverage of UMI reads (green) and internal reads (blue) shaded by the standard deviation. ( c ) Effect of tagmentation conditions on the fraction of UMI-containing reads (16 HEK293FT cells per condition). Left panel: varying Tn5 with constant 200 pg cDNA input. Right panel: varying cDNA input with constant 0.5ul Tn5. ( d ) Gene detection sensitivity for Smart-seq2 (44 cells) and Smart-seq3 (88 cells), downsampled to 1 million raw reads per HEK293FT cell. Shown are number of genes detected over 0 or 1 RPKM. P-value was computed as a two-sided t -test. ( e ) Reproducibility in gene expression quantification across HEKF293FT cells for Smart-seq2 (44 cells) and Smart-seq3 (88 cells) at RPKM and UMI level. Shown are adjusted r^2 for all pairwise cell to cell linear model fits in libraries downsampled to 1 million reads per cell. ( f ) Sensitivity to detect RNA molecules in Smart-seq3 shown by summarizing the number of unique error-corrected UMI sequences and genes detected per HEK293FT cell. Colors indicate the per cell downsampling depth ranging from 10.000 (n = 24 cells) to 750.000 (n = 16 cells) UMI-containing sequencing reads. ( g ) Violin plots summarizing the number of molecules detected per cell with Smart-seq2-UMI, Smart-seq3 and using smRNA-FISH for four X chromosomal genes (Hdac6, Igbp1, Mpp1 and Msl3). ( h ) Estimating the percent of smRNA-FISH molecules that were detected in cells using Smart-seq2-UMI and Smart-seq3. Shown are means and 95% confidence intervals.

    Techniques Used: RNA Sequencing Assay, Polymerase Chain Reaction, Standard Deviation, Expressing, Sequencing, Fluorescence In Situ Hybridization

    2) Product Images from "Expression of muscarinic receptor subtypes in tree shrew ocular tissues and their regulation during the development of myopia"

    Article Title: Expression of muscarinic receptor subtypes in tree shrew ocular tissues and their regulation during the development of myopia

    Journal: Molecular Vision

    doi:

    Regulation of choroidal muscarinic receptor subtype gene expression during myopia development. The regulation of the five mAChR s was assessed in the tree shrew choroid using real-time PCR and the DNA-binding dye, SYBR green I. Data was calculated relative to the housekeeping gene, HPRT , and expressed relative to the contralateral control eye (for myopic groups) or left/right eyes (for normal groups). Data are shown as the mean percentage change±SEM (n=5 for 1 and 5 day CHRM2 gene data, all other data n=6).
    Figure Legend Snippet: Regulation of choroidal muscarinic receptor subtype gene expression during myopia development. The regulation of the five mAChR s was assessed in the tree shrew choroid using real-time PCR and the DNA-binding dye, SYBR green I. Data was calculated relative to the housekeeping gene, HPRT , and expressed relative to the contralateral control eye (for myopic groups) or left/right eyes (for normal groups). Data are shown as the mean percentage change±SEM (n=5 for 1 and 5 day CHRM2 gene data, all other data n=6).

    Techniques Used: Expressing, Real-time Polymerase Chain Reaction, Binding Assay, SYBR Green Assay

    Regulation of retinal muscarinic receptor subtype gene expression during myopia development. The regulation of the five mAChR s was assessed in the tree shrew retina using real-time PCR and the DNA-binding dye, SYBR green I. Data was calculated relative to the housekeeping gene, HPRT , and expressed relative to the contralateral control eye (for myopic groups) or left/right eyes (for normal groups). Data are shown as the mean percentage change±SEM (n=6).
    Figure Legend Snippet: Regulation of retinal muscarinic receptor subtype gene expression during myopia development. The regulation of the five mAChR s was assessed in the tree shrew retina using real-time PCR and the DNA-binding dye, SYBR green I. Data was calculated relative to the housekeeping gene, HPRT , and expressed relative to the contralateral control eye (for myopic groups) or left/right eyes (for normal groups). Data are shown as the mean percentage change±SEM (n=6).

    Techniques Used: Expressing, Real-time Polymerase Chain Reaction, Binding Assay, SYBR Green Assay

    Regulation of scleral muscarinic receptor subtype gene expression during myopia development. The regulation of the five mAChR s was assessed in the tree shrew sclera using real-time PCR and the DNA-binding dye, SYBR green I. Data was calculated relative to the housekeeping gene, HPRT , and expressed relative to the contralateral control eye (for myopic groups) or left/right eyes (for normal groups). Data are shown as the mean percentage change±SEM (n=5 for 5 day myopia data, 1 day myopia and normal data, n=6).
    Figure Legend Snippet: Regulation of scleral muscarinic receptor subtype gene expression during myopia development. The regulation of the five mAChR s was assessed in the tree shrew sclera using real-time PCR and the DNA-binding dye, SYBR green I. Data was calculated relative to the housekeeping gene, HPRT , and expressed relative to the contralateral control eye (for myopic groups) or left/right eyes (for normal groups). Data are shown as the mean percentage change±SEM (n=5 for 5 day myopia data, 1 day myopia and normal data, n=6).

    Techniques Used: Expressing, Real-time Polymerase Chain Reaction, Binding Assay, SYBR Green Assay

    3) Product Images from "The G-rich Repeats in FMR1 and C9orf72 Loci Are Hotspots for Local Unpairing of DNA"

    Article Title: The G-rich Repeats in FMR1 and C9orf72 Loci Are Hotspots for Local Unpairing of DNA

    Journal: Genetics

    doi: 10.1534/genetics.118.301672

    R-loop enrichment at the FMR1 and C9orf72 genes by DRIP analysis. (A) R-loop enrichments were determined by DRIP-quantitative PCR with the S9.6 antibody in the FMR1 locus (green bars), RPL13A (positive gene control, blue bars), and ERG1 ). DRIP samples were pretreated (light green, light blue, and light red, respectively) or untreated (dark green, dark blue, and dark red, respectively) with RNase H. The addition of RNase H was used as a control for the pulldown assay. DRIP values are presented as percentage of input (paired t -test, * P
    Figure Legend Snippet: R-loop enrichment at the FMR1 and C9orf72 genes by DRIP analysis. (A) R-loop enrichments were determined by DRIP-quantitative PCR with the S9.6 antibody in the FMR1 locus (green bars), RPL13A (positive gene control, blue bars), and ERG1 ). DRIP samples were pretreated (light green, light blue, and light red, respectively) or untreated (dark green, dark blue, and dark red, respectively) with RNase H. The addition of RNase H was used as a control for the pulldown assay. DRIP values are presented as percentage of input (paired t -test, * P

    Techniques Used: Real-time Polymerase Chain Reaction

    4) Product Images from "The G-rich Repeats in FMR1 and C9orf72 Loci Are Hotspots for Local Unpairing of DNA"

    Article Title: The G-rich Repeats in FMR1 and C9orf72 Loci Are Hotspots for Local Unpairing of DNA

    Journal: Genetics

    doi: 10.1534/genetics.118.301672

    R-loop enrichment at the FMR1 and C9orf72 genes by DRIP analysis. (A) R-loop enrichments were determined by DRIP-quantitative PCR with the S9.6 antibody in the FMR1 locus (green bars), RPL13A (positive gene control, blue bars), and ERG1 ). DRIP samples were pretreated (light green, light blue, and light red, respectively) or untreated (dark green, dark blue, and dark red, respectively) with RNase H. The addition of RNase H was used as a control for the pulldown assay. DRIP values are presented as percentage of input (paired t -test, * P
    Figure Legend Snippet: R-loop enrichment at the FMR1 and C9orf72 genes by DRIP analysis. (A) R-loop enrichments were determined by DRIP-quantitative PCR with the S9.6 antibody in the FMR1 locus (green bars), RPL13A (positive gene control, blue bars), and ERG1 ). DRIP samples were pretreated (light green, light blue, and light red, respectively) or untreated (dark green, dark blue, and dark red, respectively) with RNase H. The addition of RNase H was used as a control for the pulldown assay. DRIP values are presented as percentage of input (paired t -test, * P

    Techniques Used: Real-time Polymerase Chain Reaction

    5) Product Images from "Adolescent alcohol exposure alters lysine demethylase 1 (LSD1) expression and histone methylation in the amygdala during adulthood"

    Article Title: Adolescent alcohol exposure alters lysine demethylase 1 (LSD1) expression and histone methylation in the amygdala during adulthood

    Journal: Addiction biology

    doi: 10.1111/adb.12404

    Decreased LSD1 protein and Lsd1+8a mRNA in specific amygdaloid structures of adult rats exposed to AIE A) Photomicrographs (Scale bar = 50 μm) showing the gold immunolabeling of LSD1 in the central (CeA) and medial nucleus of amygdala (MeA) of adolescent intermittent ethanol (AIE) - or saline (AIS)-exposed adult rats. B) Quantification of LSD1 protein levels by gold immunolabeling in the CeA, MeA and basolateral amygdala (BLA) of AIS- and AIE-exposed adult rats. C) Photomicrographs (Scale bar = 50 μm) showing the Lsd1+8a mRNA positive cells (in-situ PCR) in the CeA and MeA of AIE- and AIS-exposed adult rats. D) Quantification of Lsd1+8a mRNA levels by in situ PCR in the CeA, MeA and BLA of AIS- and AIE-exposed adult rats. Values are represented as mean ± SEM of n=5 rats and are significantly different from AIS-exposed group (** p
    Figure Legend Snippet: Decreased LSD1 protein and Lsd1+8a mRNA in specific amygdaloid structures of adult rats exposed to AIE A) Photomicrographs (Scale bar = 50 μm) showing the gold immunolabeling of LSD1 in the central (CeA) and medial nucleus of amygdala (MeA) of adolescent intermittent ethanol (AIE) - or saline (AIS)-exposed adult rats. B) Quantification of LSD1 protein levels by gold immunolabeling in the CeA, MeA and basolateral amygdala (BLA) of AIS- and AIE-exposed adult rats. C) Photomicrographs (Scale bar = 50 μm) showing the Lsd1+8a mRNA positive cells (in-situ PCR) in the CeA and MeA of AIE- and AIS-exposed adult rats. D) Quantification of Lsd1+8a mRNA levels by in situ PCR in the CeA, MeA and BLA of AIS- and AIE-exposed adult rats. Values are represented as mean ± SEM of n=5 rats and are significantly different from AIS-exposed group (** p

    Techniques Used: Immunolabeling, Microelectrode Array, In Situ, Polymerase Chain Reaction

    6) Product Images from "Acquired resistance to PI3K/mTOR inhibition is associated with mitochondrial DNA mutation and glycolysis"

    Article Title: Acquired resistance to PI3K/mTOR inhibition is associated with mitochondrial DNA mutation and glycolysis

    Journal: Oncotarget

    doi: 10.18632/oncotarget.22655

    ( A ) Representation of somatic mitochondrial DNA variants detected only in AQR clones in MT-CO1 gene. ( B ) Validation of the ENST00000361624.2: c.1367T > A variant by pyrosequencing. ( C ) PCR verification of ρ(o) status of H1975ρ(o) cells compared with parental H1975 cells. ( D ) Bar charts showing IC50 values of BEZ235, PI103, KU-0063794 and carboplatin in H1975 (blue columns) and H1975ρ0 (black columns) cells. Also shown are the levels of extracellular lactate and reactive oxygen species in H1975 and H1975ρ0 cells. * p
    Figure Legend Snippet: ( A ) Representation of somatic mitochondrial DNA variants detected only in AQR clones in MT-CO1 gene. ( B ) Validation of the ENST00000361624.2: c.1367T > A variant by pyrosequencing. ( C ) PCR verification of ρ(o) status of H1975ρ(o) cells compared with parental H1975 cells. ( D ) Bar charts showing IC50 values of BEZ235, PI103, KU-0063794 and carboplatin in H1975 (blue columns) and H1975ρ0 (black columns) cells. Also shown are the levels of extracellular lactate and reactive oxygen species in H1975 and H1975ρ0 cells. * p

    Techniques Used: Clone Assay, Variant Assay, Polymerase Chain Reaction

    7) Product Images from "A novel ultra-sensitive method for the detection of FGFR3 mutations in urine of bladder cancer patients – Design of the Urodiag® PCR kit for surveillance of patients with non-muscle-invasive bladder cancer (NMIBC)"

    Article Title: A novel ultra-sensitive method for the detection of FGFR3 mutations in urine of bladder cancer patients – Design of the Urodiag® PCR kit for surveillance of patients with non-muscle-invasive bladder cancer (NMIBC)

    Journal: BMC Medical Genetics

    doi: 10.1186/s12881-020-01050-w

    DNA integrity assessed by PCR amplification of GLOBIN gene. DNA concentrations were determined by fluorometry. The GLOBIN gene was amplified with an amount of urine DNA comprised between 10 and 18 ng (4 μl of DNA sample) from each Filter (F). Amplification curves are shown from Pool 1 to Pool 4, respectively
    Figure Legend Snippet: DNA integrity assessed by PCR amplification of GLOBIN gene. DNA concentrations were determined by fluorometry. The GLOBIN gene was amplified with an amount of urine DNA comprised between 10 and 18 ng (4 μl of DNA sample) from each Filter (F). Amplification curves are shown from Pool 1 to Pool 4, respectively

    Techniques Used: Polymerase Chain Reaction, Amplification

    LoD for the Mutation assay. DNA from FGFR3 mutant plasmids was diluted into the wild-type DNA (standard human DNA). The proportion of mutant DNA was 50, 10, 5, and 1%, respectively. The representative amplification curves ( a , c ) and mean Ct values ( b , d ) are shown in the detection of the FGFR3 S249C/Y375C (a, b) and R248C/G372C ( c , d ) mutations by MASO-PCR
    Figure Legend Snippet: LoD for the Mutation assay. DNA from FGFR3 mutant plasmids was diluted into the wild-type DNA (standard human DNA). The proportion of mutant DNA was 50, 10, 5, and 1%, respectively. The representative amplification curves ( a , c ) and mean Ct values ( b , d ) are shown in the detection of the FGFR3 S249C/Y375C (a, b) and R248C/G372C ( c , d ) mutations by MASO-PCR

    Techniques Used: Mutagenesis, Amplification, Polymerase Chain Reaction

    8) Product Images from "F4-related mutation and expression analysis of the aminopeptidase N gene in pigs 1"

    Article Title: F4-related mutation and expression analysis of the aminopeptidase N gene in pigs 1

    Journal: Journal of Animal Science

    doi: 10.2527/jas.2013-7307

    Agarose gel of Sus scrofa aminopeptidase N +/-3 (primer) ± 3 PCR products using cDNA as a template representing the absence of differential gene expression and alternative splice variants in the small intestine of F4 receptor-positive (F4R+) and F4 receptor-negative (F4R–) pigs. Lane 1: 1Kb Plus DNA ladder (Invitrogen, Carlsbad, CA, USA); lane 2: F4R+ pig 5; lane 3: F4R+ pig 8; lane 4: F4R+ pig 15; lane 5: F4R– pig 23; lane 6: F4R– pig 26; lane 7: water.
    Figure Legend Snippet: Agarose gel of Sus scrofa aminopeptidase N +/-3 (primer) ± 3 PCR products using cDNA as a template representing the absence of differential gene expression and alternative splice variants in the small intestine of F4 receptor-positive (F4R+) and F4 receptor-negative (F4R–) pigs. Lane 1: 1Kb Plus DNA ladder (Invitrogen, Carlsbad, CA, USA); lane 2: F4R+ pig 5; lane 3: F4R+ pig 8; lane 4: F4R+ pig 15; lane 5: F4R– pig 23; lane 6: F4R– pig 26; lane 7: water.

    Techniques Used: Agarose Gel Electrophoresis, Polymerase Chain Reaction, Expressing

    9) Product Images from "Effects of Surotomycin on Clostridium difficile Viability and Toxin Production In Vitro"

    Article Title: Effects of Surotomycin on Clostridium difficile Viability and Toxin Production In Vitro

    Journal: Antimicrobial Agents and Chemotherapy

    doi: 10.1128/AAC.00275-15

    qRT-PCR analysis of tcdA expression following treatment with surotomycin, metronidazole, or vancomycin at 8× MIC. Strain UK1 was grown in CaCl 2 -supplemented TY medium for ∼12 h as described above, at which time the culture was split and drugs were added at 8× MIC. At 2, 4, 8, and 24 h after addition of drugs, cell samples were harvested, RNA was extracted, and cDNA was synthesized as described in Materials and Methods. cDNA corresponding to tcdA mRNA was quantified by real-time PCR. Reactions were performed in triplicate using cDNA synthesized from each of a minimum of three biological replicates, and results are presented as the means and SEM of the data obtained. Results were calculated using the threshold cycle (2 −ΔΔ CT ) method, in which the amount of target mRNA was normalized to that of an internal control transcript ( rpoC ).
    Figure Legend Snippet: qRT-PCR analysis of tcdA expression following treatment with surotomycin, metronidazole, or vancomycin at 8× MIC. Strain UK1 was grown in CaCl 2 -supplemented TY medium for ∼12 h as described above, at which time the culture was split and drugs were added at 8× MIC. At 2, 4, 8, and 24 h after addition of drugs, cell samples were harvested, RNA was extracted, and cDNA was synthesized as described in Materials and Methods. cDNA corresponding to tcdA mRNA was quantified by real-time PCR. Reactions were performed in triplicate using cDNA synthesized from each of a minimum of three biological replicates, and results are presented as the means and SEM of the data obtained. Results were calculated using the threshold cycle (2 −ΔΔ CT ) method, in which the amount of target mRNA was normalized to that of an internal control transcript ( rpoC ).

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

    10) Product Images from "Acquired resistance to PI3K/mTOR inhibition is associated with mitochondrial DNA mutation and glycolysis"

    Article Title: Acquired resistance to PI3K/mTOR inhibition is associated with mitochondrial DNA mutation and glycolysis

    Journal: Oncotarget

    doi: 10.18632/oncotarget.22655

    ( A ) Representation of somatic mitochondrial DNA variants detected only in AQR clones in MT-CO1 gene. ( B ) Validation of the ENST00000361624.2: c.1367T > A variant by pyrosequencing. ( C ) PCR verification of ρ(o) status of H1975ρ(o) cells compared with parental H1975 cells. ( D ) Bar charts showing IC50 values of BEZ235, PI103, KU-0063794 and carboplatin in H1975 (blue columns) and H1975ρ0 (black columns) cells. Also shown are the levels of extracellular lactate and reactive oxygen species in H1975 and H1975ρ0 cells. * p
    Figure Legend Snippet: ( A ) Representation of somatic mitochondrial DNA variants detected only in AQR clones in MT-CO1 gene. ( B ) Validation of the ENST00000361624.2: c.1367T > A variant by pyrosequencing. ( C ) PCR verification of ρ(o) status of H1975ρ(o) cells compared with parental H1975 cells. ( D ) Bar charts showing IC50 values of BEZ235, PI103, KU-0063794 and carboplatin in H1975 (blue columns) and H1975ρ0 (black columns) cells. Also shown are the levels of extracellular lactate and reactive oxygen species in H1975 and H1975ρ0 cells. * p

    Techniques Used: Clone Assay, Variant Assay, Polymerase Chain Reaction

    11) Product Images from "AMPK Activation Reduces Hepatic Lipid Content by Increasing Fat Oxidation In Vivo"

    Article Title: AMPK Activation Reduces Hepatic Lipid Content by Increasing Fat Oxidation In Vivo

    Journal: International Journal of Molecular Sciences

    doi: 10.3390/ijms19092826

    Effects of the expression of an active form of AMP-activated protein kinase (AMPK) in the liver on body weight and food intake. Ten-week-old male C57BL/6J mice received injections of adenovirus (Ad) expressing the green fluorescent protein (GFP) or a constitutively active form of AMPKα2 (AMPK-CA) and were studied for the indicated times after adenovirus injection and in the indicated nutritional state. ( A ) Western blot analysis of liver lysates with antibodies raised against pan-AMPKα and myc-tagged AMPK-CA was performed on days 2 and day 8 after adenovirus administration; ( B ) Western blot analysis of liver lysates from fed mice 48 h after the injection of Ad GFP or Ad AMPK-CA, with the antibodies indicated. Each lane represents a liver sample from an individual mouse. The panel on the right shows Ser79 phosphorylated acetyl CoA carboxylase/ total acetyl CoA carboxylase (P-ACC/ACC) ratios from the quantification of immunoblot images ( n = 5); ( C ) Hepatic malonyl-CoA levels in 8 h-fasted mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 5); ( D ) Effect of AMPK activation in the liver on the expression of Cpt1a and Cpt 2 genes. Total RNA was isolated from the liver of 24 h-fasted mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 6). The expression of Cpt1a and Cpt2 genes was assessed by real-time quantitative RT-PCR. Relative mRNA levels are expressed as fold-activation relative to levels in Ad GFP livers; ( E ) Body weight changes and ( F ) cumulative food intake measured for 8 days after adenovirus administration ( n = 11–12 per group). Data are means ± standard error of mean (SEM). * p
    Figure Legend Snippet: Effects of the expression of an active form of AMP-activated protein kinase (AMPK) in the liver on body weight and food intake. Ten-week-old male C57BL/6J mice received injections of adenovirus (Ad) expressing the green fluorescent protein (GFP) or a constitutively active form of AMPKα2 (AMPK-CA) and were studied for the indicated times after adenovirus injection and in the indicated nutritional state. ( A ) Western blot analysis of liver lysates with antibodies raised against pan-AMPKα and myc-tagged AMPK-CA was performed on days 2 and day 8 after adenovirus administration; ( B ) Western blot analysis of liver lysates from fed mice 48 h after the injection of Ad GFP or Ad AMPK-CA, with the antibodies indicated. Each lane represents a liver sample from an individual mouse. The panel on the right shows Ser79 phosphorylated acetyl CoA carboxylase/ total acetyl CoA carboxylase (P-ACC/ACC) ratios from the quantification of immunoblot images ( n = 5); ( C ) Hepatic malonyl-CoA levels in 8 h-fasted mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 5); ( D ) Effect of AMPK activation in the liver on the expression of Cpt1a and Cpt 2 genes. Total RNA was isolated from the liver of 24 h-fasted mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 6). The expression of Cpt1a and Cpt2 genes was assessed by real-time quantitative RT-PCR. Relative mRNA levels are expressed as fold-activation relative to levels in Ad GFP livers; ( E ) Body weight changes and ( F ) cumulative food intake measured for 8 days after adenovirus administration ( n = 11–12 per group). Data are means ± standard error of mean (SEM). * p

    Techniques Used: Expressing, Mouse Assay, Injection, Western Blot, Activation Assay, Cycling Probe Technology, Isolation, Quantitative RT-PCR

    Long-term adenovirus-mediated expression of an active form of AMPK in the liver increases hepatic lipid oxidation and fatty acid uptake. Ten-week-old male C57BL/6J mice received injections of Ad GFP or Ad AMPK CA and were studied at the indicated times after adenovirus injection and in the indicated nutritional state. ( A ) Hepatic [1- 14 C]-palmitate oxidation in fed mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 4); ( B ) Plasma β-hydroxybutyrate levels in 24 h-fasted mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 6); ( C ) Plasma triglyceride (TG) and ( D ) plasma free fatty acid (FFA) levels in overnight-fasted mice 8 days after the injection of Ad GFP or Ad AMPK-CA ( n = 12); ( E ) Hepatic [1- 14 C]-palmitate uptake in 24 h-fasted mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 5); ( F ) Effect of AMPK activation in the liver on the expression of the fatty acid transporters. Total RNA was isolated from the liver of 24 h-fasted mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 5). The expression of Slc27a4 ( Fatp4 ), Cd36 , and Fabp4 genes was assessed by real-time quantitative RT-PCR. Relative mRNA levels are expressed as fold-activation relative to levels in Ad GFP livers. Data are means ± SEM. * p
    Figure Legend Snippet: Long-term adenovirus-mediated expression of an active form of AMPK in the liver increases hepatic lipid oxidation and fatty acid uptake. Ten-week-old male C57BL/6J mice received injections of Ad GFP or Ad AMPK CA and were studied at the indicated times after adenovirus injection and in the indicated nutritional state. ( A ) Hepatic [1- 14 C]-palmitate oxidation in fed mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 4); ( B ) Plasma β-hydroxybutyrate levels in 24 h-fasted mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 6); ( C ) Plasma triglyceride (TG) and ( D ) plasma free fatty acid (FFA) levels in overnight-fasted mice 8 days after the injection of Ad GFP or Ad AMPK-CA ( n = 12); ( E ) Hepatic [1- 14 C]-palmitate uptake in 24 h-fasted mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 5); ( F ) Effect of AMPK activation in the liver on the expression of the fatty acid transporters. Total RNA was isolated from the liver of 24 h-fasted mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 5). The expression of Slc27a4 ( Fatp4 ), Cd36 , and Fabp4 genes was assessed by real-time quantitative RT-PCR. Relative mRNA levels are expressed as fold-activation relative to levels in Ad GFP livers. Data are means ± SEM. * p

    Techniques Used: Expressing, Mouse Assay, Injection, Activation Assay, Isolation, Quantitative RT-PCR

    12) Product Images from "AMPK Activation Reduces Hepatic Lipid Content by Increasing Fat Oxidation In Vivo"

    Article Title: AMPK Activation Reduces Hepatic Lipid Content by Increasing Fat Oxidation In Vivo

    Journal: International Journal of Molecular Sciences

    doi: 10.3390/ijms19092826

    Effects of the expression of an active form of AMP-activated protein kinase (AMPK) in the liver on body weight and food intake. Ten-week-old male C57BL/6J mice received injections of adenovirus (Ad) expressing the green fluorescent protein (GFP) or a constitutively active form of AMPKα2 (AMPK-CA) and were studied for the indicated times after adenovirus injection and in the indicated nutritional state. ( A ) Western blot analysis of liver lysates with antibodies raised against pan-AMPKα and myc-tagged AMPK-CA was performed on days 2 and day 8 after adenovirus administration; ( B ) Western blot analysis of liver lysates from fed mice 48 h after the injection of Ad GFP or Ad AMPK-CA, with the antibodies indicated. Each lane represents a liver sample from an individual mouse. The panel on the right shows Ser79 phosphorylated acetyl CoA carboxylase/ total acetyl CoA carboxylase (P-ACC/ACC) ratios from the quantification of immunoblot images ( n = 5); ( C ) Hepatic malonyl-CoA levels in 8 h-fasted mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 5); ( D ) Effect of AMPK activation in the liver on the expression of Cpt1a and Cpt 2 genes. Total RNA was isolated from the liver of 24 h-fasted mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 6). The expression of Cpt1a and Cpt2 genes was assessed by real-time quantitative RT-PCR. Relative mRNA levels are expressed as fold-activation relative to levels in Ad GFP livers; ( E ) Body weight changes and ( F ) cumulative food intake measured for 8 days after adenovirus administration ( n = 11–12 per group). Data are means ± standard error of mean (SEM). * p
    Figure Legend Snippet: Effects of the expression of an active form of AMP-activated protein kinase (AMPK) in the liver on body weight and food intake. Ten-week-old male C57BL/6J mice received injections of adenovirus (Ad) expressing the green fluorescent protein (GFP) or a constitutively active form of AMPKα2 (AMPK-CA) and were studied for the indicated times after adenovirus injection and in the indicated nutritional state. ( A ) Western blot analysis of liver lysates with antibodies raised against pan-AMPKα and myc-tagged AMPK-CA was performed on days 2 and day 8 after adenovirus administration; ( B ) Western blot analysis of liver lysates from fed mice 48 h after the injection of Ad GFP or Ad AMPK-CA, with the antibodies indicated. Each lane represents a liver sample from an individual mouse. The panel on the right shows Ser79 phosphorylated acetyl CoA carboxylase/ total acetyl CoA carboxylase (P-ACC/ACC) ratios from the quantification of immunoblot images ( n = 5); ( C ) Hepatic malonyl-CoA levels in 8 h-fasted mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 5); ( D ) Effect of AMPK activation in the liver on the expression of Cpt1a and Cpt 2 genes. Total RNA was isolated from the liver of 24 h-fasted mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 6). The expression of Cpt1a and Cpt2 genes was assessed by real-time quantitative RT-PCR. Relative mRNA levels are expressed as fold-activation relative to levels in Ad GFP livers; ( E ) Body weight changes and ( F ) cumulative food intake measured for 8 days after adenovirus administration ( n = 11–12 per group). Data are means ± standard error of mean (SEM). * p

    Techniques Used: Expressing, Mouse Assay, Injection, Western Blot, Activation Assay, Cycling Probe Technology, Isolation, Quantitative RT-PCR

    Long-term adenovirus-mediated expression of an active form of AMPK in the liver increases hepatic lipid oxidation and fatty acid uptake. Ten-week-old male C57BL/6J mice received injections of Ad GFP or Ad AMPK CA and were studied at the indicated times after adenovirus injection and in the indicated nutritional state. ( A ) Hepatic [1- 14 C]-palmitate oxidation in fed mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 4); ( B ) Plasma β-hydroxybutyrate levels in 24 h-fasted mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 6); ( C ) Plasma triglyceride (TG) and ( D ) plasma free fatty acid (FFA) levels in overnight-fasted mice 8 days after the injection of Ad GFP or Ad AMPK-CA ( n = 12); ( E ) Hepatic [1- 14 C]-palmitate uptake in 24 h-fasted mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 5); ( F ) Effect of AMPK activation in the liver on the expression of the fatty acid transporters. Total RNA was isolated from the liver of 24 h-fasted mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 5). The expression of Slc27a4 ( Fatp4 ), Cd36 , and Fabp4 genes was assessed by real-time quantitative RT-PCR. Relative mRNA levels are expressed as fold-activation relative to levels in Ad GFP livers. Data are means ± SEM. * p
    Figure Legend Snippet: Long-term adenovirus-mediated expression of an active form of AMPK in the liver increases hepatic lipid oxidation and fatty acid uptake. Ten-week-old male C57BL/6J mice received injections of Ad GFP or Ad AMPK CA and were studied at the indicated times after adenovirus injection and in the indicated nutritional state. ( A ) Hepatic [1- 14 C]-palmitate oxidation in fed mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 4); ( B ) Plasma β-hydroxybutyrate levels in 24 h-fasted mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 6); ( C ) Plasma triglyceride (TG) and ( D ) plasma free fatty acid (FFA) levels in overnight-fasted mice 8 days after the injection of Ad GFP or Ad AMPK-CA ( n = 12); ( E ) Hepatic [1- 14 C]-palmitate uptake in 24 h-fasted mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 5); ( F ) Effect of AMPK activation in the liver on the expression of the fatty acid transporters. Total RNA was isolated from the liver of 24 h-fasted mice 48 h after the injection of Ad GFP or Ad AMPK-CA ( n = 5). The expression of Slc27a4 ( Fatp4 ), Cd36 , and Fabp4 genes was assessed by real-time quantitative RT-PCR. Relative mRNA levels are expressed as fold-activation relative to levels in Ad GFP livers. Data are means ± SEM. * p

    Techniques Used: Expressing, Mouse Assay, Injection, Activation Assay, Isolation, Quantitative RT-PCR

    Related Articles

    Polymerase Chain Reaction:

    Article Title: The G-rich Repeats in FMR1 and C9orf72 Loci Are Hotspots for Local Unpairing of DNA
    Article Snippet: .. For ssDNA displacement at FMR1 , GC-RICH solution was added to the PCR mix, according to the manufacturer’s instructions (Roche). .. Amplified products were cloned, and single colonies were analyzed for cytosine conversions by direct sequencing (ABI 3130), using the BigDye Terminator v3.1 Cycle Sequencing Kit.

    Article Title: Single-cell RNA counting at allele- and isoform-resolution using Smart-seq3
    Article Snippet: .. The reaction was terminated by incubating at 85 degrees for 5 min. PCR preamplification was performed directly after reverse transcription by adding 6 μL of PCR mix, bringing reaction concentrations to 1x KAPA HiFi PCR buffer (contains 2mM MgCl2 at 1×) (Roche), 0.02u/μl DNA polymerase (Roche), 0.3mM dNTPs, 0.1μM Smartseq3 Forward PCR primer (5’-TCGTCGGCAGCGTCAGATGTGTATAAGAGACAGATTGCGCAATG-3’; IDT), 0.1μM Smartseq3 Reverse PCR primer (5’-ACGAGCATCAGCAGCATACGA-3’; IDT). ..

    Article Title: A novel ultra-sensitive method for the detection of FGFR3 mutations in urine of bladder cancer patients – Design of the Urodiag® PCR kit for surveillance of patients with non-muscle-invasive bladder cancer (NMIBC)
    Article Snippet: .. All reactions were performed with 4 μl of bisulfite-converted positive control DNA (100% methylated) and 16 μl of PCR mix containing 1x KAPA PROBE FAST qPCR Master Mix (KAPA Biosystems), 400 nM primers (Eurogentec) and 250 nM TaqMan-mgb probes (Thermo Fischer Scientific). .. ALBUMIN sequence has been designed without CpG site and used for normalizing the DNA amounts.

    Article Title: Expression of muscarinic receptor subtypes in tree shrew ocular tissues and their regulation during the development of myopia
    Article Snippet: .. Total RNA (0.5 μg) was reverse transcribed as described above, and amplifications were performed using a commercial PCR mix incorporating SYBR green 1 (FastStart DNA Master Mix; Roche). .. The protocol for each receptor subtype was optimized to ensure that a single, specific product was produced ( ).

    Article Title: The G-rich Repeats in FMR1 and C9orf72 Loci Are Hotspots for Local Unpairing of DNA
    Article Snippet: .. For ssDNA displacement at FMR1 , GC-RICH solution was added to the PCR mix, according to the manufacturer’s instructions (Roche). .. Amplified products were cloned, and single colonies were analyzed for cytosine conversions by direct sequencing (ABI 3130), using the BigDye Terminator v3.1 Cycle Sequencing Kit.

    Article Title: Adolescent alcohol exposure alters lysine demethylase 1 (LSD1) expression and histone methylation in the amygdala during adulthood
    Article Snippet: .. Sections were transferred to a clean tube with a PCR mix containing 100 pmol of Lsd1+8a mRNA primers (same as those used for real-time PCR above; ) and digoxigenin (DIG)-11-dUTP (Roche Diagnostics, Indianapolis, IN, USA) rather than dTTP. .. After PCR cycling, sections were mounted and incubated with anti-DIG antibody conjugated to alkaline phosphotase (Roche).

    Article Title: Cloning and Characterization of a Family B DNA Polymerase from the Hyperthermophilic Crenarchaeon Pyrobaculum islandicum †
    Article Snippet: .. Approximately 1 ng of P. islandicum DNA and 100 pmol of each primer were added to a PCR mixture containing 200 μM deoxyribonucleoside triphosphates and 2 U of Expand High Fidelity enzyme (Roche Diagnostics). .. After an initial denaturation step at 94°C for 2 min, 30 cycles with a temperature profile of 10 s at 94°C, 30 s at 46°C, and 90 s at 68°C were performed with a DNA thermal cycler (GeneAmp PCR System 2400; Perkin-Elmer).

    Article Title: Acquired resistance to PI3K/mTOR inhibition is associated with mitochondrial DNA mutation and glycolysis
    Article Snippet: .. The PCR mix comprised 100 ng DNA, 2.5 μl of PCR buffer with MgCl2 , 0.5 μl of DNTP mix, 0.2 μl of Taq Polymerase (Roche), and 2.5 μl of forward and reverse primer (IDT) in a total volume of 25μl. .. The primers were as follows: Nuclear DNA Forward: 5′-CAT TGC TCC TCC TGA GCG CAA-3′, Nuclear DNA Reverse: 5′-GCT GTC ACC TTC ACC GTT CCA-3′, Mitochondrial DNA Forward: 5′-ACA ATA GCT AAG ACC CAA ACT GGG-3′, Mitochondrial DNA Reverse: 5′-CCA TTT CTT GCC ACC TCA TGG GC-3′.

    Real-time Polymerase Chain Reaction:

    Article Title: A novel ultra-sensitive method for the detection of FGFR3 mutations in urine of bladder cancer patients – Design of the Urodiag® PCR kit for surveillance of patients with non-muscle-invasive bladder cancer (NMIBC)
    Article Snippet: .. All reactions were performed with 4 μl of bisulfite-converted positive control DNA (100% methylated) and 16 μl of PCR mix containing 1x KAPA PROBE FAST qPCR Master Mix (KAPA Biosystems), 400 nM primers (Eurogentec) and 250 nM TaqMan-mgb probes (Thermo Fischer Scientific). .. ALBUMIN sequence has been designed without CpG site and used for normalizing the DNA amounts.

    Article Title: Adolescent alcohol exposure alters lysine demethylase 1 (LSD1) expression and histone methylation in the amygdala during adulthood
    Article Snippet: .. Sections were transferred to a clean tube with a PCR mix containing 100 pmol of Lsd1+8a mRNA primers (same as those used for real-time PCR above; ) and digoxigenin (DIG)-11-dUTP (Roche Diagnostics, Indianapolis, IN, USA) rather than dTTP. .. After PCR cycling, sections were mounted and incubated with anti-DIG antibody conjugated to alkaline phosphotase (Roche).

    SYBR Green Assay:

    Article Title: Expression of muscarinic receptor subtypes in tree shrew ocular tissues and their regulation during the development of myopia
    Article Snippet: .. Total RNA (0.5 μg) was reverse transcribed as described above, and amplifications were performed using a commercial PCR mix incorporating SYBR green 1 (FastStart DNA Master Mix; Roche). .. The protocol for each receptor subtype was optimized to ensure that a single, specific product was produced ( ).

    Positive Control:

    Article Title: A novel ultra-sensitive method for the detection of FGFR3 mutations in urine of bladder cancer patients – Design of the Urodiag® PCR kit for surveillance of patients with non-muscle-invasive bladder cancer (NMIBC)
    Article Snippet: .. All reactions were performed with 4 μl of bisulfite-converted positive control DNA (100% methylated) and 16 μl of PCR mix containing 1x KAPA PROBE FAST qPCR Master Mix (KAPA Biosystems), 400 nM primers (Eurogentec) and 250 nM TaqMan-mgb probes (Thermo Fischer Scientific). .. ALBUMIN sequence has been designed without CpG site and used for normalizing the DNA amounts.

    Methylation:

    Article Title: A novel ultra-sensitive method for the detection of FGFR3 mutations in urine of bladder cancer patients – Design of the Urodiag® PCR kit for surveillance of patients with non-muscle-invasive bladder cancer (NMIBC)
    Article Snippet: .. All reactions were performed with 4 μl of bisulfite-converted positive control DNA (100% methylated) and 16 μl of PCR mix containing 1x KAPA PROBE FAST qPCR Master Mix (KAPA Biosystems), 400 nM primers (Eurogentec) and 250 nM TaqMan-mgb probes (Thermo Fischer Scientific). .. ALBUMIN sequence has been designed without CpG site and used for normalizing the DNA amounts.

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    Roche lc pcr master mix
    Detection of HSV <t>DNA</t> from clinical samples by LightCycler <t>PCR.</t>
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    Regulation of choroidal muscarinic receptor subtype gene expression during myopia development. The regulation of the five mAChR s was assessed in the tree shrew choroid using real-time <t>PCR</t> and the DNA-binding dye, <t>SYBR</t> green I. Data was calculated relative to the housekeeping gene, HPRT , and expressed relative to the contralateral control eye (for myopic groups) or left/right eyes (for normal groups). Data are shown as the mean percentage change±SEM (n=5 for 1 and 5 day CHRM2 gene data, all other data n=6).
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    Roche runx2 pcr mixtures
    Distribution of <t>RUNX2</t> gene expression (A) on patient’s population and statistically significant correlations between ovulation induction factors, such as number of follicles, oocytes, fertilized oocytes (B) and hormone serum levels for E2 (C) and LH (D) respectively. RUNX2 gene expression was determined by real-time <t>PCR</t> for 41 patients and categorized into two groups of patients, with expression and without expression. Expression ratios (RUNX2 copies/G6PDH copies) were used for the evaluation of RUNX2 gene expression.
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    Roche alu ltr pcr mixture
    Analysis of transcripts and nuclear HIV-1 DNA in FcγR-stimulated or control MDM. MDM (10 6 cells/well of a 12-well plate) were activated with hIgG and immediately infected with HIV-1 BaL (5 × 10 2 TCID 50 /10 6 cells) for 1 h at 37°C. Cells were lysed at the indicated time points after infection, and 5 μl of cell lysate (corresponding to approximately 20,000 cells) was analyzed by <t>PCR.</t> Primers specific for intermediate ( pol ) and late (U3/ gag ) reverse transcripts, nuclear 2LTR circles, or integrated provirus ( <t>Alu</t> -LTR) were used. PCR products were analyzed by gel electrophoresis. Four independent experiments performed with MDM from different donors showed similar PCR amplification profiles despite signal intensity variations. NI, noninfected; −, absence of hIgG; +, presence of hIgG.
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    Image Search Results


    Detection of HSV DNA from clinical samples by LightCycler PCR.

    Journal: BMC Microbiology

    Article Title: Detection and subtyping of Herpes simplex virus in clinical samples by LightCycler PCR, enzyme immunoassay and cell culture

    doi: 10.1186/1471-2180-2-12

    Figure Lengend Snippet: Detection of HSV DNA from clinical samples by LightCycler PCR.

    Article Snippet: The LC-PCR master mix contained the following: 1× FastStart Taq DNA polymerase reaction buffer (Roche Molecular Diagnostics) which included a dNTP mix (containing dUTP instead of dTTP), 3 mM MgCl2 , 0.5 μM of each primer, 0.2 μM HSV-2 FLU and 0.4 μM HSV-2 LCR.

    Techniques: Polymerase Chain Reaction

    Regulation of choroidal muscarinic receptor subtype gene expression during myopia development. The regulation of the five mAChR s was assessed in the tree shrew choroid using real-time PCR and the DNA-binding dye, SYBR green I. Data was calculated relative to the housekeeping gene, HPRT , and expressed relative to the contralateral control eye (for myopic groups) or left/right eyes (for normal groups). Data are shown as the mean percentage change±SEM (n=5 for 1 and 5 day CHRM2 gene data, all other data n=6).

    Journal: Molecular Vision

    Article Title: Expression of muscarinic receptor subtypes in tree shrew ocular tissues and their regulation during the development of myopia

    doi:

    Figure Lengend Snippet: Regulation of choroidal muscarinic receptor subtype gene expression during myopia development. The regulation of the five mAChR s was assessed in the tree shrew choroid using real-time PCR and the DNA-binding dye, SYBR green I. Data was calculated relative to the housekeeping gene, HPRT , and expressed relative to the contralateral control eye (for myopic groups) or left/right eyes (for normal groups). Data are shown as the mean percentage change±SEM (n=5 for 1 and 5 day CHRM2 gene data, all other data n=6).

    Article Snippet: Total RNA (0.5 μg) was reverse transcribed as described above, and amplifications were performed using a commercial PCR mix incorporating SYBR green 1 (FastStart DNA Master Mix; Roche).

    Techniques: Expressing, Real-time Polymerase Chain Reaction, Binding Assay, SYBR Green Assay

    Regulation of retinal muscarinic receptor subtype gene expression during myopia development. The regulation of the five mAChR s was assessed in the tree shrew retina using real-time PCR and the DNA-binding dye, SYBR green I. Data was calculated relative to the housekeeping gene, HPRT , and expressed relative to the contralateral control eye (for myopic groups) or left/right eyes (for normal groups). Data are shown as the mean percentage change±SEM (n=6).

    Journal: Molecular Vision

    Article Title: Expression of muscarinic receptor subtypes in tree shrew ocular tissues and their regulation during the development of myopia

    doi:

    Figure Lengend Snippet: Regulation of retinal muscarinic receptor subtype gene expression during myopia development. The regulation of the five mAChR s was assessed in the tree shrew retina using real-time PCR and the DNA-binding dye, SYBR green I. Data was calculated relative to the housekeeping gene, HPRT , and expressed relative to the contralateral control eye (for myopic groups) or left/right eyes (for normal groups). Data are shown as the mean percentage change±SEM (n=6).

    Article Snippet: Total RNA (0.5 μg) was reverse transcribed as described above, and amplifications were performed using a commercial PCR mix incorporating SYBR green 1 (FastStart DNA Master Mix; Roche).

    Techniques: Expressing, Real-time Polymerase Chain Reaction, Binding Assay, SYBR Green Assay

    Regulation of scleral muscarinic receptor subtype gene expression during myopia development. The regulation of the five mAChR s was assessed in the tree shrew sclera using real-time PCR and the DNA-binding dye, SYBR green I. Data was calculated relative to the housekeeping gene, HPRT , and expressed relative to the contralateral control eye (for myopic groups) or left/right eyes (for normal groups). Data are shown as the mean percentage change±SEM (n=5 for 5 day myopia data, 1 day myopia and normal data, n=6).

    Journal: Molecular Vision

    Article Title: Expression of muscarinic receptor subtypes in tree shrew ocular tissues and their regulation during the development of myopia

    doi:

    Figure Lengend Snippet: Regulation of scleral muscarinic receptor subtype gene expression during myopia development. The regulation of the five mAChR s was assessed in the tree shrew sclera using real-time PCR and the DNA-binding dye, SYBR green I. Data was calculated relative to the housekeeping gene, HPRT , and expressed relative to the contralateral control eye (for myopic groups) or left/right eyes (for normal groups). Data are shown as the mean percentage change±SEM (n=5 for 5 day myopia data, 1 day myopia and normal data, n=6).

    Article Snippet: Total RNA (0.5 μg) was reverse transcribed as described above, and amplifications were performed using a commercial PCR mix incorporating SYBR green 1 (FastStart DNA Master Mix; Roche).

    Techniques: Expressing, Real-time Polymerase Chain Reaction, Binding Assay, SYBR Green Assay

    Distribution of RUNX2 gene expression (A) on patient’s population and statistically significant correlations between ovulation induction factors, such as number of follicles, oocytes, fertilized oocytes (B) and hormone serum levels for E2 (C) and LH (D) respectively. RUNX2 gene expression was determined by real-time PCR for 41 patients and categorized into two groups of patients, with expression and without expression. Expression ratios (RUNX2 copies/G6PDH copies) were used for the evaluation of RUNX2 gene expression.

    Journal: Reproductive Biology and Endocrinology : RB & E

    Article Title: Detection of RUNX2 gene expression in cumulus cells in women undergoing controlled ovarian stimulation

    doi: 10.1186/1477-7827-10-99

    Figure Lengend Snippet: Distribution of RUNX2 gene expression (A) on patient’s population and statistically significant correlations between ovulation induction factors, such as number of follicles, oocytes, fertilized oocytes (B) and hormone serum levels for E2 (C) and LH (D) respectively. RUNX2 gene expression was determined by real-time PCR for 41 patients and categorized into two groups of patients, with expression and without expression. Expression ratios (RUNX2 copies/G6PDH copies) were used for the evaluation of RUNX2 gene expression.

    Article Snippet: RUNX2 PCR mixtures contained 5 μl cDNA, 5x master mix (LightCycler 480 Genotyping Master, Roche), 20 μM of each primer, 20 μM of each probe and PCR-grade water (LightCycler 480 Genotyping Master, Roche) to a total volume of 20 μl.

    Techniques: Expressing, Real-time Polymerase Chain Reaction

    Analysis of transcripts and nuclear HIV-1 DNA in FcγR-stimulated or control MDM. MDM (10 6 cells/well of a 12-well plate) were activated with hIgG and immediately infected with HIV-1 BaL (5 × 10 2 TCID 50 /10 6 cells) for 1 h at 37°C. Cells were lysed at the indicated time points after infection, and 5 μl of cell lysate (corresponding to approximately 20,000 cells) was analyzed by PCR. Primers specific for intermediate ( pol ) and late (U3/ gag ) reverse transcripts, nuclear 2LTR circles, or integrated provirus ( Alu -LTR) were used. PCR products were analyzed by gel electrophoresis. Four independent experiments performed with MDM from different donors showed similar PCR amplification profiles despite signal intensity variations. NI, noninfected; −, absence of hIgG; +, presence of hIgG.

    Journal: Journal of Virology

    Article Title: Fc? Receptor-Mediated Suppression of Human Immunodeficiency Virus Type 1 Replication in Primary Human Macrophages

    doi: 10.1128/JVI.77.7.4081-4094.2003

    Figure Lengend Snippet: Analysis of transcripts and nuclear HIV-1 DNA in FcγR-stimulated or control MDM. MDM (10 6 cells/well of a 12-well plate) were activated with hIgG and immediately infected with HIV-1 BaL (5 × 10 2 TCID 50 /10 6 cells) for 1 h at 37°C. Cells were lysed at the indicated time points after infection, and 5 μl of cell lysate (corresponding to approximately 20,000 cells) was analyzed by PCR. Primers specific for intermediate ( pol ) and late (U3/ gag ) reverse transcripts, nuclear 2LTR circles, or integrated provirus ( Alu -LTR) were used. PCR products were analyzed by gel electrophoresis. Four independent experiments performed with MDM from different donors showed similar PCR amplification profiles despite signal intensity variations. NI, noninfected; −, absence of hIgG; +, presence of hIgG.

    Article Snippet: The Alu -LTR PCR mixture contained 5 μl of lysate, 5 μl of 10× PCR buffer (Roche Diagnostics GmbH), 1.75 μl of 10 nM dNTP, 1.5 μl each of sense and antisense primers (from a 10 μM solution), and 0.75 μl of Long Expand Template Taq DNA polymerase (Roche Diagnostics GmbH).

    Techniques: Infection, Polymerase Chain Reaction, Nucleic Acid Electrophoresis, Amplification