miscript ii rt kit  (Qiagen)


Bioz Verified Symbol Qiagen is a verified supplier
Bioz Manufacturer Symbol Qiagen manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 99
    Name:
    miScript II RT Kit
    Description:
    For reverse transcription of total RNA containing miRNA using the miScript PCR System Kit contents Qiagen miScript II RT Kit 12 rxns SYBR Green based Single step cDNA Synthesis Quantification of miRNA and mRNA For Reverse Transcription of Total RNA Containing miRNA Using the miScript PCR System Includes miScript Reverse Transcriptase Mix 10x miScript Nucleics Mix 5x miScript HiSpec Buffer 5x miScript HiFlex Buffer RNase free Water Benefits cDNA for sensitive and specific miRNA detection A single cDNA enables quantification of multiple miRNAs Quantification of miRNA and mRNA from the same cDNA Quantification of mature and precursor miRNA from the same cDNA Proprietary synthetic RNA to assess reverse transcription efficiency
    Catalog Number:
    218160
    Price:
    133
    Category:
    miScript II RT Kit
    Buy from Supplier


    Structured Review

    Qiagen miscript ii rt kit
    miScript II RT Kit
    For reverse transcription of total RNA containing miRNA using the miScript PCR System Kit contents Qiagen miScript II RT Kit 12 rxns SYBR Green based Single step cDNA Synthesis Quantification of miRNA and mRNA For Reverse Transcription of Total RNA Containing miRNA Using the miScript PCR System Includes miScript Reverse Transcriptase Mix 10x miScript Nucleics Mix 5x miScript HiSpec Buffer 5x miScript HiFlex Buffer RNase free Water Benefits cDNA for sensitive and specific miRNA detection A single cDNA enables quantification of multiple miRNAs Quantification of miRNA and mRNA from the same cDNA Quantification of mature and precursor miRNA from the same cDNA Proprietary synthetic RNA to assess reverse transcription efficiency
    https://www.bioz.com/result/miscript ii rt kit/product/Qiagen
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    miscript ii rt kit - by Bioz Stars, 2021-05
    99/100 stars

    Images

    1) Product Images from "miR-214-Dependent Increase of PHLPP2 Levels Mediates the Impairment of Insulin-Stimulated Akt Activation in Mouse Aortic Endothelial Cells Exposed to Methylglyoxal"

    Article Title: miR-214-Dependent Increase of PHLPP2 Levels Mediates the Impairment of Insulin-Stimulated Akt Activation in Mouse Aortic Endothelial Cells Exposed to Methylglyoxal

    Journal: International Journal of Molecular Sciences

    doi: 10.3390/ijms19020522

    Effect of miR-214 on PHLPP2 regulation. ( a ) MAECs were transfected with a non-targeting control oligonucleotide (Ctr M; white bars) or miR-214 mimic (miR-214 M 50 nmol/L; black bars). Where indicated, MAECs were co-transfected with miScript miR-214 Target Protector (miR-214 TP 0.1 nmol/L). Protein lysates obtained from 48 h transfected MAECs were analyzed by Western blot with anti-PHLPP2 and anti-α-tubulin antibodies. Exposure timing of blots was of 2 min. Protein levels were quantified by the densitometric analysis of at least three independent experiments. Bars in the graphs represent the mean ± SD of the percent (%) over control (Ctr M). ( b ) MAECs were transfected for 24 h with non-targeting control oligonucleotide (Ctr M 50 nmol/L; white bars) or miR-214 mimic (miR-214 M 50 nmol/L; black bars) in the presence of the pEZX-MT06 control reporter vector or the pEZX-MT06/PHLPP2-3′UTR reporter vector. Firefly and Renilla luciferase activities were determined in cell lysates under each experimental condition. Results are normalized to Renilla activity. Bars in the graphs represent the mean ± SD of the fold over control (Ctr M). Statistical analysis was evaluated using Student’s t -test; *** p ≤ 0.001.
    Figure Legend Snippet: Effect of miR-214 on PHLPP2 regulation. ( a ) MAECs were transfected with a non-targeting control oligonucleotide (Ctr M; white bars) or miR-214 mimic (miR-214 M 50 nmol/L; black bars). Where indicated, MAECs were co-transfected with miScript miR-214 Target Protector (miR-214 TP 0.1 nmol/L). Protein lysates obtained from 48 h transfected MAECs were analyzed by Western blot with anti-PHLPP2 and anti-α-tubulin antibodies. Exposure timing of blots was of 2 min. Protein levels were quantified by the densitometric analysis of at least three independent experiments. Bars in the graphs represent the mean ± SD of the percent (%) over control (Ctr M). ( b ) MAECs were transfected for 24 h with non-targeting control oligonucleotide (Ctr M 50 nmol/L; white bars) or miR-214 mimic (miR-214 M 50 nmol/L; black bars) in the presence of the pEZX-MT06 control reporter vector or the pEZX-MT06/PHLPP2-3′UTR reporter vector. Firefly and Renilla luciferase activities were determined in cell lysates under each experimental condition. Results are normalized to Renilla activity. Bars in the graphs represent the mean ± SD of the fold over control (Ctr M). Statistical analysis was evaluated using Student’s t -test; *** p ≤ 0.001.

    Techniques Used: Transfection, Western Blot, Plasmid Preparation, Luciferase, Activity Assay

    2) Product Images from "A Plasma Biomarker Panel of Four MicroRNAs for the Diagnosis of Prostate Cancer"

    Article Title: A Plasma Biomarker Panel of Four MicroRNAs for the Diagnosis of Prostate Cancer

    Journal: Scientific Reports

    doi: 10.1038/s41598-018-24424-w

    Screening of plasma miRNA markers in pooled patient and control samples and validation in the discovery cohort. ( a) Scatter plot of 372 cancer-associated miRNAs in a screening cohort of pooled plasma samples from prostate cancer patients and healthy controls screened using a miScript miRNA PCR array. miRNAs found to be differentially expressed between the patient and control groups are shown in black, and unchanged miRNAs are shown in grey. A fold regulation cut-off of 2.5 was selected for the analysis. (b) Scatter plot showing the 11 differentially regulated miRNAs re-analysed by qRT-PCR in patient samples in the discovery cohort (N = 61). The mean fold regulation of each miRNA across the patient and control samples was taken into account and those that were below the 2 fold regulation cut-off were excluded from further analysis. The selected miRNAs are shown in colour. (c) Relative levels of miR-4289, miR-326, miR-152-3p and miR-98-5p analysed by qRT-PCR as in ( b ) in patients vs healthy controls. Statistically significant differences were assessed using a Mann-Whitney U test; p values are shown after Bonferroni correction for multiple testing. Each data point represents a plasma sample, the horizontal middle line in each data set represents the mean, and the limits of the vertical lines represent the standard deviation.
    Figure Legend Snippet: Screening of plasma miRNA markers in pooled patient and control samples and validation in the discovery cohort. ( a) Scatter plot of 372 cancer-associated miRNAs in a screening cohort of pooled plasma samples from prostate cancer patients and healthy controls screened using a miScript miRNA PCR array. miRNAs found to be differentially expressed between the patient and control groups are shown in black, and unchanged miRNAs are shown in grey. A fold regulation cut-off of 2.5 was selected for the analysis. (b) Scatter plot showing the 11 differentially regulated miRNAs re-analysed by qRT-PCR in patient samples in the discovery cohort (N = 61). The mean fold regulation of each miRNA across the patient and control samples was taken into account and those that were below the 2 fold regulation cut-off were excluded from further analysis. The selected miRNAs are shown in colour. (c) Relative levels of miR-4289, miR-326, miR-152-3p and miR-98-5p analysed by qRT-PCR as in ( b ) in patients vs healthy controls. Statistically significant differences were assessed using a Mann-Whitney U test; p values are shown after Bonferroni correction for multiple testing. Each data point represents a plasma sample, the horizontal middle line in each data set represents the mean, and the limits of the vertical lines represent the standard deviation.

    Techniques Used: Polymerase Chain Reaction, Quantitative RT-PCR, MANN-WHITNEY, Standard Deviation

    3) Product Images from "A Plasma Biomarker Panel of Four MicroRNAs for the Diagnosis of Prostate Cancer"

    Article Title: A Plasma Biomarker Panel of Four MicroRNAs for the Diagnosis of Prostate Cancer

    Journal: Scientific Reports

    doi: 10.1038/s41598-018-24424-w

    Screening of plasma miRNA markers in pooled patient and control samples and validation in the discovery cohort. ( a) Scatter plot of 372 cancer-associated miRNAs in a screening cohort of pooled plasma samples from prostate cancer patients and healthy controls screened using a miScript miRNA PCR array. miRNAs found to be differentially expressed between the patient and control groups are shown in black, and unchanged miRNAs are shown in grey. A fold regulation cut-off of 2.5 was selected for the analysis. (b) Scatter plot showing the 11 differentially regulated miRNAs re-analysed by qRT-PCR in patient samples in the discovery cohort (N = 61). The mean fold regulation of each miRNA across the patient and control samples was taken into account and those that were below the 2 fold regulation cut-off were excluded from further analysis. The selected miRNAs are shown in colour. (c) Relative levels of miR-4289, miR-326, miR-152-3p and miR-98-5p analysed by qRT-PCR as in ( b ) in patients vs healthy controls. Statistically significant differences were assessed using a Mann-Whitney U test; p values are shown after Bonferroni correction for multiple testing. Each data point represents a plasma sample, the horizontal middle line in each data set represents the mean, and the limits of the vertical lines represent the standard deviation.
    Figure Legend Snippet: Screening of plasma miRNA markers in pooled patient and control samples and validation in the discovery cohort. ( a) Scatter plot of 372 cancer-associated miRNAs in a screening cohort of pooled plasma samples from prostate cancer patients and healthy controls screened using a miScript miRNA PCR array. miRNAs found to be differentially expressed between the patient and control groups are shown in black, and unchanged miRNAs are shown in grey. A fold regulation cut-off of 2.5 was selected for the analysis. (b) Scatter plot showing the 11 differentially regulated miRNAs re-analysed by qRT-PCR in patient samples in the discovery cohort (N = 61). The mean fold regulation of each miRNA across the patient and control samples was taken into account and those that were below the 2 fold regulation cut-off were excluded from further analysis. The selected miRNAs are shown in colour. (c) Relative levels of miR-4289, miR-326, miR-152-3p and miR-98-5p analysed by qRT-PCR as in ( b ) in patients vs healthy controls. Statistically significant differences were assessed using a Mann-Whitney U test; p values are shown after Bonferroni correction for multiple testing. Each data point represents a plasma sample, the horizontal middle line in each data set represents the mean, and the limits of the vertical lines represent the standard deviation.

    Techniques Used: Polymerase Chain Reaction, Quantitative RT-PCR, MANN-WHITNEY, Standard Deviation

    4) Product Images from "The novel hsa-miR-12528 regulates tumourigenesis and metastasis through hypo-phosphorylation of AKT cascade by targeting IGF-1R in human lung cancer"

    Article Title: The novel hsa-miR-12528 regulates tumourigenesis and metastasis through hypo-phosphorylation of AKT cascade by targeting IGF-1R in human lung cancer

    Journal: Cell Death & Disease

    doi: 10.1038/s41419-018-0535-8

    Basic information, expression profiling and influence on the novel hsa-miR-12528 in lung cancer. Identification and cloning of the miR-12528 from lung cancer cells. The assumed secondary folding structure of miR-12528. Human genomic sequences were found using the RNAfold web-tool. The marked location is a mature miR-12528 sequence. The miR-12528 is located on chromosome 19p.13.3 (649215-649234) ( a ). Maturation of miR-12528 is dependent on the Dicer pathway, as shown via Dicer -knockdown and an miScript miRNA assay ( b ). Mature miR-12528 is conserved at a high rate in other species. These results were determined using the NCBI BLAST tool ( c ). The miR-12528 expression was assessed in both 7 NSCLC cell lines ( d ) and 20 pairs of NSCLC patient tissues ( e ) compared with normal WI-38 and BEAS-2B cells, and matched normal tissues. When classified histologically, cases 1–10 are adenocarcinoma type and cases 11–20 are squamous cell carcinoma type ( e ). The proliferative activity in 7 NSCLC cells was assessed using a XTT reagent 48 h post transfection with 100 nM mimics ( f ). The miR-12528 expression levels were observed using qRT-PCR between absence and presence of FBS stimulation ( g ) and between diploid normal WI-38 and immortalised WI-38 VA13 ( h ). These expression profiling data were normalised to RNU6B and performed in triplicate using an miScript assay on independent samples (bars, mean ± S.E.M.; * p
    Figure Legend Snippet: Basic information, expression profiling and influence on the novel hsa-miR-12528 in lung cancer. Identification and cloning of the miR-12528 from lung cancer cells. The assumed secondary folding structure of miR-12528. Human genomic sequences were found using the RNAfold web-tool. The marked location is a mature miR-12528 sequence. The miR-12528 is located on chromosome 19p.13.3 (649215-649234) ( a ). Maturation of miR-12528 is dependent on the Dicer pathway, as shown via Dicer -knockdown and an miScript miRNA assay ( b ). Mature miR-12528 is conserved at a high rate in other species. These results were determined using the NCBI BLAST tool ( c ). The miR-12528 expression was assessed in both 7 NSCLC cell lines ( d ) and 20 pairs of NSCLC patient tissues ( e ) compared with normal WI-38 and BEAS-2B cells, and matched normal tissues. When classified histologically, cases 1–10 are adenocarcinoma type and cases 11–20 are squamous cell carcinoma type ( e ). The proliferative activity in 7 NSCLC cells was assessed using a XTT reagent 48 h post transfection with 100 nM mimics ( f ). The miR-12528 expression levels were observed using qRT-PCR between absence and presence of FBS stimulation ( g ) and between diploid normal WI-38 and immortalised WI-38 VA13 ( h ). These expression profiling data were normalised to RNU6B and performed in triplicate using an miScript assay on independent samples (bars, mean ± S.E.M.; * p

    Techniques Used: Expressing, Clone Assay, Genomic Sequencing, Sequencing, Activity Assay, Transfection, Quantitative RT-PCR

    Related Articles

    Isolation:

    Article Title: The novel hsa-miR-12528 regulates tumourigenesis and metastasis through hypo-phosphorylation of AKT cascade by targeting IGF-1R in human lung cancer
    Article Snippet: A549 cells were transfected or overexpressed by a miRNA or siRNA mimic (Genolution Pharmaceuticals Inc., Seoul, Korea)-Lipofectamine 2000 (Invitrogen, Carlsbad, CA, USA) mixture, according to the manufacturers’ protocol. .. Total RNA was isolated using the TRIzol reagent (Qiagen, Hilden, Germany) as previously described . cDNA was synthesised using the reverse transcription Kit (Invitrogen) and the miScript II RT Kit (Qiagen), according to the manufacturers’ protocol. .. The results of miRNA or gene profiling were analysed using the quantitative reverse transcription-PCR (RT-PCR) analysis (Bio-Rad, Hercules, CA, USA) and expressed as 2(−delta (delta) threshold cycle) values (2−∆∆(C)T ) with reference previously described .

    Real-time Polymerase Chain Reaction:

    Article Title: RNA-induced epigenetic silencing inhibits HIV-1 reactivation from latency
    Article Snippet: RNA was extracted from all JLat 9.2 cell lines using Monarch Total RNA Miniprep kit as per the manufacturer’s instructions (NEB, Cat# 2010S). .. RNA was quantitated by Nanospectrometer and 1000 ng of RNA was transcribed using the miScript II RT Kit as per the manufacturer’s protocol (Qiagen, Cat# 218161). cDNA was diluted as per kit instructions using nuclease free H2 O (from 20 μL to 500 μL volume). qPCR was then performed using miScript SYBR Green PCR kit (Qiagen Cat# 218073). .. Primer pairs used were a universal loop primer 5′-TTC TGT GAA GCC ACA GAT GGG AA-3′ and the Qiagen universal reverse primer supplied with the miScript kit.

    Article Title: miR-214-Dependent Increase of PHLPP2 Levels Mediates the Impairment of Insulin-Stimulated Akt Activation in Mouse Aortic Endothelial Cells Exposed to Methylglyoxal
    Article Snippet: Total RNA was isolated from the aortic tissue of Glo1-knockdown (Glo1KD) mice [ ], after homogenization in Qiazol using miRNeasy mini kit (QIAGEN, Hilden, Germany), according to manufacturer’s instructions. .. After quantification with NanoDrop 2000 spectrophotometer (Thermo Scientific, Waltham, MA, USA), total RNA was reverse transcribed using the miScript II RT Kit (QIAGEN), and the differential expression of miRNA-214 was analyzed by real time-PCR using the miScript SYBR Green PCR Kit (QIAGEN) and quantified as expression units relative to U6 snRNA, used as housekeeping small RNA. .. Specific primers used for amplification were purchased from QIAGEN: Mm_miR-214_2 miScript Primer Assay, MS00032571 RNU6B_13 miScript Primer Assay, MS00014000

    SYBR Green Assay:

    Article Title: RNA-induced epigenetic silencing inhibits HIV-1 reactivation from latency
    Article Snippet: RNA was extracted from all JLat 9.2 cell lines using Monarch Total RNA Miniprep kit as per the manufacturer’s instructions (NEB, Cat# 2010S). .. RNA was quantitated by Nanospectrometer and 1000 ng of RNA was transcribed using the miScript II RT Kit as per the manufacturer’s protocol (Qiagen, Cat# 218161). cDNA was diluted as per kit instructions using nuclease free H2 O (from 20 μL to 500 μL volume). qPCR was then performed using miScript SYBR Green PCR kit (Qiagen Cat# 218073). .. Primer pairs used were a universal loop primer 5′-TTC TGT GAA GCC ACA GAT GGG AA-3′ and the Qiagen universal reverse primer supplied with the miScript kit.

    Article Title: A Plasma Biomarker Panel of Four MicroRNAs for the Diagnosis of Prostate Cancer
    Article Snippet: The expression of the significantly deregulated miRNAs in prostate tumour tissues was evaluated in a panel of prostate cancer cell lines by qRT-PCR. .. RNA was extracted from cell lysates using the miRNeasy Micro kit (Qiagen) according to the manufacturer’s instructions followed by cDNA synthesis using the miScript II RT kit with HiSpec Buffer (Qiagen). qRT-PCR using the miScript primer assays (Qiagen) and QuantiTect SYBR Green PCR Master Mix (Qiagen) was performed as described previously. .. The LNCaP and PC3 prostate cancer cell lines were obtained from the American Type Culture Collection (ATCC, Rockville, MD, USA) and cultured in RPMI-1640 (Gibco) supplemented with 5% Fetal Bovine Serum (FBS) (Life Technologies).

    Article Title: miR-214-Dependent Increase of PHLPP2 Levels Mediates the Impairment of Insulin-Stimulated Akt Activation in Mouse Aortic Endothelial Cells Exposed to Methylglyoxal
    Article Snippet: Total RNA was isolated from the aortic tissue of Glo1-knockdown (Glo1KD) mice [ ], after homogenization in Qiazol using miRNeasy mini kit (QIAGEN, Hilden, Germany), according to manufacturer’s instructions. .. After quantification with NanoDrop 2000 spectrophotometer (Thermo Scientific, Waltham, MA, USA), total RNA was reverse transcribed using the miScript II RT Kit (QIAGEN), and the differential expression of miRNA-214 was analyzed by real time-PCR using the miScript SYBR Green PCR Kit (QIAGEN) and quantified as expression units relative to U6 snRNA, used as housekeeping small RNA. .. Specific primers used for amplification were purchased from QIAGEN: Mm_miR-214_2 miScript Primer Assay, MS00032571 RNU6B_13 miScript Primer Assay, MS00014000

    Polymerase Chain Reaction:

    Article Title: RNA-induced epigenetic silencing inhibits HIV-1 reactivation from latency
    Article Snippet: RNA was extracted from all JLat 9.2 cell lines using Monarch Total RNA Miniprep kit as per the manufacturer’s instructions (NEB, Cat# 2010S). .. RNA was quantitated by Nanospectrometer and 1000 ng of RNA was transcribed using the miScript II RT Kit as per the manufacturer’s protocol (Qiagen, Cat# 218161). cDNA was diluted as per kit instructions using nuclease free H2 O (from 20 μL to 500 μL volume). qPCR was then performed using miScript SYBR Green PCR kit (Qiagen Cat# 218073). .. Primer pairs used were a universal loop primer 5′-TTC TGT GAA GCC ACA GAT GGG AA-3′ and the Qiagen universal reverse primer supplied with the miScript kit.

    Article Title: A Plasma Biomarker Panel of Four MicroRNAs for the Diagnosis of Prostate Cancer
    Article Snippet: The expression of the significantly deregulated miRNAs in prostate tumour tissues was evaluated in a panel of prostate cancer cell lines by qRT-PCR. .. RNA was extracted from cell lysates using the miRNeasy Micro kit (Qiagen) according to the manufacturer’s instructions followed by cDNA synthesis using the miScript II RT kit with HiSpec Buffer (Qiagen). qRT-PCR using the miScript primer assays (Qiagen) and QuantiTect SYBR Green PCR Master Mix (Qiagen) was performed as described previously. .. The LNCaP and PC3 prostate cancer cell lines were obtained from the American Type Culture Collection (ATCC, Rockville, MD, USA) and cultured in RPMI-1640 (Gibco) supplemented with 5% Fetal Bovine Serum (FBS) (Life Technologies).

    Article Title: miR-214-Dependent Increase of PHLPP2 Levels Mediates the Impairment of Insulin-Stimulated Akt Activation in Mouse Aortic Endothelial Cells Exposed to Methylglyoxal
    Article Snippet: Total RNA was isolated from the aortic tissue of Glo1-knockdown (Glo1KD) mice [ ], after homogenization in Qiazol using miRNeasy mini kit (QIAGEN, Hilden, Germany), according to manufacturer’s instructions. .. After quantification with NanoDrop 2000 spectrophotometer (Thermo Scientific, Waltham, MA, USA), total RNA was reverse transcribed using the miScript II RT Kit (QIAGEN), and the differential expression of miRNA-214 was analyzed by real time-PCR using the miScript SYBR Green PCR Kit (QIAGEN) and quantified as expression units relative to U6 snRNA, used as housekeeping small RNA. .. Specific primers used for amplification were purchased from QIAGEN: Mm_miR-214_2 miScript Primer Assay, MS00032571 RNU6B_13 miScript Primer Assay, MS00014000

    Quantitative RT-PCR:

    Article Title: Expressing MicroRNA Bantam Sponge Drastically Improves the Insecticidal Activity of Baculovirus via Increasing the Level of Ecdysteroid Hormone in Spodoptera exigua Larvae
    Article Snippet: To determine the level of bantam in Sf9 cells, total small RNAs ( < 200 nt) were harvested from Sf9 cells and reverse-transcribed using miRVana microRNA Isolation Kit (Ambion, Life Technologies, Carlsbad, CA, Unites States). .. RT-qPCR detection of bantam was performed using miScript II RT Kit (Qiagen, Venlo, Netherlands) according to the manufacturer’s instructions. ..

    Article Title: A Plasma Biomarker Panel of Four MicroRNAs for the Diagnosis of Prostate Cancer
    Article Snippet: The expression of the significantly deregulated miRNAs in prostate tumour tissues was evaluated in a panel of prostate cancer cell lines by qRT-PCR. .. RNA was extracted from cell lysates using the miRNeasy Micro kit (Qiagen) according to the manufacturer’s instructions followed by cDNA synthesis using the miScript II RT kit with HiSpec Buffer (Qiagen). qRT-PCR using the miScript primer assays (Qiagen) and QuantiTect SYBR Green PCR Master Mix (Qiagen) was performed as described previously. .. The LNCaP and PC3 prostate cancer cell lines were obtained from the American Type Culture Collection (ATCC, Rockville, MD, USA) and cultured in RPMI-1640 (Gibco) supplemented with 5% Fetal Bovine Serum (FBS) (Life Technologies).

    Homogenization:

    Article Title: Ursodeoxycholic acid improves liver function via phenylalanine/tyrosine pathway and microbiome remodelling in patients with liver dysfunction
    Article Snippet: Total RNA was extracted from serum (100 μL) for quantitative analysis of miRNAs using an miRNeasy mini kit (Qiagen, Hilden, Germany) according to the manufacturer’s recommendations. .. A synthetic miRNA from Caenorhabditis elegans was included in the sample after its homogenisation in the QIAzol lysis reagent (Qiagen) to validate the RNA extraction efficiency. cDNA was prepared using a miScript reverse transcription kit II (Qiagen). .. The polymerase chain reaction (PCR; 20 µL) contained 10 μL of the miScript SYBR Green PCR master mix (Qiagen), 4 μL of nuclease-free water, 2 μL of 10 × miScript primer assay, 2 μL of 10 × miScript universal primer, and 2 μL of the cDNA template.

    Lysis:

    Article Title: Ursodeoxycholic acid improves liver function via phenylalanine/tyrosine pathway and microbiome remodelling in patients with liver dysfunction
    Article Snippet: Total RNA was extracted from serum (100 μL) for quantitative analysis of miRNAs using an miRNeasy mini kit (Qiagen, Hilden, Germany) according to the manufacturer’s recommendations. .. A synthetic miRNA from Caenorhabditis elegans was included in the sample after its homogenisation in the QIAzol lysis reagent (Qiagen) to validate the RNA extraction efficiency. cDNA was prepared using a miScript reverse transcription kit II (Qiagen). .. The polymerase chain reaction (PCR; 20 µL) contained 10 μL of the miScript SYBR Green PCR master mix (Qiagen), 4 μL of nuclease-free water, 2 μL of 10 × miScript primer assay, 2 μL of 10 × miScript universal primer, and 2 μL of the cDNA template.

    RNA Extraction:

    Article Title: Ursodeoxycholic acid improves liver function via phenylalanine/tyrosine pathway and microbiome remodelling in patients with liver dysfunction
    Article Snippet: Total RNA was extracted from serum (100 μL) for quantitative analysis of miRNAs using an miRNeasy mini kit (Qiagen, Hilden, Germany) according to the manufacturer’s recommendations. .. A synthetic miRNA from Caenorhabditis elegans was included in the sample after its homogenisation in the QIAzol lysis reagent (Qiagen) to validate the RNA extraction efficiency. cDNA was prepared using a miScript reverse transcription kit II (Qiagen). .. The polymerase chain reaction (PCR; 20 µL) contained 10 μL of the miScript SYBR Green PCR master mix (Qiagen), 4 μL of nuclease-free water, 2 μL of 10 × miScript primer assay, 2 μL of 10 × miScript universal primer, and 2 μL of the cDNA template.

    Spectrophotometry:

    Article Title: miR-214-Dependent Increase of PHLPP2 Levels Mediates the Impairment of Insulin-Stimulated Akt Activation in Mouse Aortic Endothelial Cells Exposed to Methylglyoxal
    Article Snippet: Total RNA was isolated from the aortic tissue of Glo1-knockdown (Glo1KD) mice [ ], after homogenization in Qiazol using miRNeasy mini kit (QIAGEN, Hilden, Germany), according to manufacturer’s instructions. .. After quantification with NanoDrop 2000 spectrophotometer (Thermo Scientific, Waltham, MA, USA), total RNA was reverse transcribed using the miScript II RT Kit (QIAGEN), and the differential expression of miRNA-214 was analyzed by real time-PCR using the miScript SYBR Green PCR Kit (QIAGEN) and quantified as expression units relative to U6 snRNA, used as housekeeping small RNA. .. Specific primers used for amplification were purchased from QIAGEN: Mm_miR-214_2 miScript Primer Assay, MS00032571 RNU6B_13 miScript Primer Assay, MS00014000

    Expressing:

    Article Title: miR-214-Dependent Increase of PHLPP2 Levels Mediates the Impairment of Insulin-Stimulated Akt Activation in Mouse Aortic Endothelial Cells Exposed to Methylglyoxal
    Article Snippet: Total RNA was isolated from the aortic tissue of Glo1-knockdown (Glo1KD) mice [ ], after homogenization in Qiazol using miRNeasy mini kit (QIAGEN, Hilden, Germany), according to manufacturer’s instructions. .. After quantification with NanoDrop 2000 spectrophotometer (Thermo Scientific, Waltham, MA, USA), total RNA was reverse transcribed using the miScript II RT Kit (QIAGEN), and the differential expression of miRNA-214 was analyzed by real time-PCR using the miScript SYBR Green PCR Kit (QIAGEN) and quantified as expression units relative to U6 snRNA, used as housekeeping small RNA. .. Specific primers used for amplification were purchased from QIAGEN: Mm_miR-214_2 miScript Primer Assay, MS00032571 RNU6B_13 miScript Primer Assay, MS00014000

    Similar Products

  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 99
    Qiagen miscript ii rt kit
    Effect of miR-214 on PHLPP2 regulation. ( a ) MAECs were transfected with a non-targeting control oligonucleotide (Ctr M; white bars) or miR-214 mimic (miR-214 M 50 nmol/L; black bars). Where indicated, MAECs were co-transfected with <t>miScript</t> miR-214 Target Protector (miR-214 TP 0.1 nmol/L). Protein lysates obtained from 48 h transfected MAECs were analyzed by Western blot with anti-PHLPP2 and anti-α-tubulin antibodies. Exposure timing of blots was of 2 min. Protein levels were quantified by the densitometric analysis of at least three independent experiments. Bars in the graphs represent the mean ± SD of the percent (%) over control (Ctr M). ( b ) MAECs were transfected for 24 h with non-targeting control oligonucleotide (Ctr M 50 nmol/L; white bars) or miR-214 mimic (miR-214 M 50 nmol/L; black bars) in the presence of the pEZX-MT06 control reporter vector or the pEZX-MT06/PHLPP2-3′UTR reporter vector. Firefly and Renilla luciferase activities were determined in cell lysates under each experimental condition. Results are normalized to Renilla activity. Bars in the graphs represent the mean ± SD of the fold over control (Ctr M). Statistical analysis was evaluated using Student’s t -test; *** p ≤ 0.001.
    Miscript Ii Rt Kit, supplied by Qiagen, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/miscript ii rt kit/product/Qiagen
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    miscript ii rt kit - by Bioz Stars, 2021-05
    99/100 stars
      Buy from Supplier

    86
    Qiagen rt qpcr quantification
    O-GlcNAc regulates DI splicing to control both O-GlcNAc homeostasis and cell state transitions. (A) Volcano plot of fold changes (log 2 ) in DIs of polyadenylated transcripts isolated from cells treated with OGT inhibitor for 6 h (10 μM OSMI-2 relative to DMSO) showing a global decrease in DI abundance; ( B ) A similar volcano plot as in (A), but generated from cells treated with OGA inhibitor for 2 h (5 uM TMG), showing a global shift toward increased intron detention. (C) <t>RT-Qpcr</t> analysis comparing the abundance of DI-containing transcripts in the nuclear and the cytoplasmic fractions for a panel of genes. <t>mRNA</t> was isolated from subcellular fractions obtained from HEK293T cells. The transcript for actin acts as a cytoplasmic localization control, and the transcript for MALAT1 acts as a nuclear localization control. (D) RT-qPCR analysis of the abundance of a panel of DI-containing transcripts from HEK293T cells that were treated with either DMSO or OSMI-2 for 6 h. Transcript abundances are normalized to DMSO control. (E) Changes in alternative splicing events after 6 h treatment with OGT inhibitor (10 uM OSMI-2, left) or OGA inhibitor (5 uM TMG, right) represented as boxplots. Color coded dots represent splicing events that meet the statistical cutoff. The black dotted line denotes no change compared to the DMSO control.
    Rt Qpcr Quantification, supplied by Qiagen, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rt qpcr quantification/product/Qiagen
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rt qpcr quantification - by Bioz Stars, 2021-05
    86/100 stars
      Buy from Supplier

    Image Search Results


    Effect of miR-214 on PHLPP2 regulation. ( a ) MAECs were transfected with a non-targeting control oligonucleotide (Ctr M; white bars) or miR-214 mimic (miR-214 M 50 nmol/L; black bars). Where indicated, MAECs were co-transfected with miScript miR-214 Target Protector (miR-214 TP 0.1 nmol/L). Protein lysates obtained from 48 h transfected MAECs were analyzed by Western blot with anti-PHLPP2 and anti-α-tubulin antibodies. Exposure timing of blots was of 2 min. Protein levels were quantified by the densitometric analysis of at least three independent experiments. Bars in the graphs represent the mean ± SD of the percent (%) over control (Ctr M). ( b ) MAECs were transfected for 24 h with non-targeting control oligonucleotide (Ctr M 50 nmol/L; white bars) or miR-214 mimic (miR-214 M 50 nmol/L; black bars) in the presence of the pEZX-MT06 control reporter vector or the pEZX-MT06/PHLPP2-3′UTR reporter vector. Firefly and Renilla luciferase activities were determined in cell lysates under each experimental condition. Results are normalized to Renilla activity. Bars in the graphs represent the mean ± SD of the fold over control (Ctr M). Statistical analysis was evaluated using Student’s t -test; *** p ≤ 0.001.

    Journal: International Journal of Molecular Sciences

    Article Title: miR-214-Dependent Increase of PHLPP2 Levels Mediates the Impairment of Insulin-Stimulated Akt Activation in Mouse Aortic Endothelial Cells Exposed to Methylglyoxal

    doi: 10.3390/ijms19020522

    Figure Lengend Snippet: Effect of miR-214 on PHLPP2 regulation. ( a ) MAECs were transfected with a non-targeting control oligonucleotide (Ctr M; white bars) or miR-214 mimic (miR-214 M 50 nmol/L; black bars). Where indicated, MAECs were co-transfected with miScript miR-214 Target Protector (miR-214 TP 0.1 nmol/L). Protein lysates obtained from 48 h transfected MAECs were analyzed by Western blot with anti-PHLPP2 and anti-α-tubulin antibodies. Exposure timing of blots was of 2 min. Protein levels were quantified by the densitometric analysis of at least three independent experiments. Bars in the graphs represent the mean ± SD of the percent (%) over control (Ctr M). ( b ) MAECs were transfected for 24 h with non-targeting control oligonucleotide (Ctr M 50 nmol/L; white bars) or miR-214 mimic (miR-214 M 50 nmol/L; black bars) in the presence of the pEZX-MT06 control reporter vector or the pEZX-MT06/PHLPP2-3′UTR reporter vector. Firefly and Renilla luciferase activities were determined in cell lysates under each experimental condition. Results are normalized to Renilla activity. Bars in the graphs represent the mean ± SD of the fold over control (Ctr M). Statistical analysis was evaluated using Student’s t -test; *** p ≤ 0.001.

    Article Snippet: After quantification with NanoDrop 2000 spectrophotometer (Thermo Scientific, Waltham, MA, USA), total RNA was reverse transcribed using the miScript II RT Kit (QIAGEN), and the differential expression of miRNA-214 was analyzed by real time-PCR using the miScript SYBR Green PCR Kit (QIAGEN) and quantified as expression units relative to U6 snRNA, used as housekeeping small RNA.

    Techniques: Transfection, Western Blot, Plasmid Preparation, Luciferase, Activity Assay

    Screening of plasma miRNA markers in pooled patient and control samples and validation in the discovery cohort. ( a) Scatter plot of 372 cancer-associated miRNAs in a screening cohort of pooled plasma samples from prostate cancer patients and healthy controls screened using a miScript miRNA PCR array. miRNAs found to be differentially expressed between the patient and control groups are shown in black, and unchanged miRNAs are shown in grey. A fold regulation cut-off of 2.5 was selected for the analysis. (b) Scatter plot showing the 11 differentially regulated miRNAs re-analysed by qRT-PCR in patient samples in the discovery cohort (N = 61). The mean fold regulation of each miRNA across the patient and control samples was taken into account and those that were below the 2 fold regulation cut-off were excluded from further analysis. The selected miRNAs are shown in colour. (c) Relative levels of miR-4289, miR-326, miR-152-3p and miR-98-5p analysed by qRT-PCR as in ( b ) in patients vs healthy controls. Statistically significant differences were assessed using a Mann-Whitney U test; p values are shown after Bonferroni correction for multiple testing. Each data point represents a plasma sample, the horizontal middle line in each data set represents the mean, and the limits of the vertical lines represent the standard deviation.

    Journal: Scientific Reports

    Article Title: A Plasma Biomarker Panel of Four MicroRNAs for the Diagnosis of Prostate Cancer

    doi: 10.1038/s41598-018-24424-w

    Figure Lengend Snippet: Screening of plasma miRNA markers in pooled patient and control samples and validation in the discovery cohort. ( a) Scatter plot of 372 cancer-associated miRNAs in a screening cohort of pooled plasma samples from prostate cancer patients and healthy controls screened using a miScript miRNA PCR array. miRNAs found to be differentially expressed between the patient and control groups are shown in black, and unchanged miRNAs are shown in grey. A fold regulation cut-off of 2.5 was selected for the analysis. (b) Scatter plot showing the 11 differentially regulated miRNAs re-analysed by qRT-PCR in patient samples in the discovery cohort (N = 61). The mean fold regulation of each miRNA across the patient and control samples was taken into account and those that were below the 2 fold regulation cut-off were excluded from further analysis. The selected miRNAs are shown in colour. (c) Relative levels of miR-4289, miR-326, miR-152-3p and miR-98-5p analysed by qRT-PCR as in ( b ) in patients vs healthy controls. Statistically significant differences were assessed using a Mann-Whitney U test; p values are shown after Bonferroni correction for multiple testing. Each data point represents a plasma sample, the horizontal middle line in each data set represents the mean, and the limits of the vertical lines represent the standard deviation.

    Article Snippet: RNA was extracted from cell lysates using the miRNeasy Micro kit (Qiagen) according to the manufacturer’s instructions followed by cDNA synthesis using the miScript II RT kit with HiSpec Buffer (Qiagen). qRT-PCR using the miScript primer assays (Qiagen) and QuantiTect SYBR Green PCR Master Mix (Qiagen) was performed as described previously.

    Techniques: Polymerase Chain Reaction, Quantitative RT-PCR, MANN-WHITNEY, Standard Deviation

    O-GlcNAc regulates DI splicing to control both O-GlcNAc homeostasis and cell state transitions. (A) Volcano plot of fold changes (log 2 ) in DIs of polyadenylated transcripts isolated from cells treated with OGT inhibitor for 6 h (10 μM OSMI-2 relative to DMSO) showing a global decrease in DI abundance; ( B ) A similar volcano plot as in (A), but generated from cells treated with OGA inhibitor for 2 h (5 uM TMG), showing a global shift toward increased intron detention. (C) RT-Qpcr analysis comparing the abundance of DI-containing transcripts in the nuclear and the cytoplasmic fractions for a panel of genes. mRNA was isolated from subcellular fractions obtained from HEK293T cells. The transcript for actin acts as a cytoplasmic localization control, and the transcript for MALAT1 acts as a nuclear localization control. (D) RT-qPCR analysis of the abundance of a panel of DI-containing transcripts from HEK293T cells that were treated with either DMSO or OSMI-2 for 6 h. Transcript abundances are normalized to DMSO control. (E) Changes in alternative splicing events after 6 h treatment with OGT inhibitor (10 uM OSMI-2, left) or OGA inhibitor (5 uM TMG, right) represented as boxplots. Color coded dots represent splicing events that meet the statistical cutoff. The black dotted line denotes no change compared to the DMSO control.

    Journal: bioRxiv

    Article Title: O-GlcNAc regulates gene expression by controlling detained intron splicing

    doi: 10.1101/2020.03.27.012781

    Figure Lengend Snippet: O-GlcNAc regulates DI splicing to control both O-GlcNAc homeostasis and cell state transitions. (A) Volcano plot of fold changes (log 2 ) in DIs of polyadenylated transcripts isolated from cells treated with OGT inhibitor for 6 h (10 μM OSMI-2 relative to DMSO) showing a global decrease in DI abundance; ( B ) A similar volcano plot as in (A), but generated from cells treated with OGA inhibitor for 2 h (5 uM TMG), showing a global shift toward increased intron detention. (C) RT-Qpcr analysis comparing the abundance of DI-containing transcripts in the nuclear and the cytoplasmic fractions for a panel of genes. mRNA was isolated from subcellular fractions obtained from HEK293T cells. The transcript for actin acts as a cytoplasmic localization control, and the transcript for MALAT1 acts as a nuclear localization control. (D) RT-qPCR analysis of the abundance of a panel of DI-containing transcripts from HEK293T cells that were treated with either DMSO or OSMI-2 for 6 h. Transcript abundances are normalized to DMSO control. (E) Changes in alternative splicing events after 6 h treatment with OGT inhibitor (10 uM OSMI-2, left) or OGA inhibitor (5 uM TMG, right) represented as boxplots. Color coded dots represent splicing events that meet the statistical cutoff. The black dotted line denotes no change compared to the DMSO control.

    Article Snippet: Total RNA extracted per fraction was quantified and used to calculate the percentage of mRNA localization for the RT–qPCR quantification.

    Techniques: Isolation, Generated, Quantitative RT-PCR

    O-GlcNAc levels regulate OGT and OGA DI splicing so that exon inclusion/skipping results in opposite effects on productive message. (A) Splice junctions derived from RNA-seq for OGT and OGA show two predominant fully-spliced isoforms, a functional one for OGT (A) and a non-functional one for OGA (B) . HEK-293T cells were treated with DMSO (white bars), OGT inhibitor (10 μM OSMI-2, dark grey) or OGA inhibitor (5 μM TMG, light grey) for 2 h and RT-qPCR was used to measure levels of the indicated products for OGT (C) or OGA (D) , which report on the presence of DIs as well as skipped or included exons. Red arrows depict location of primers with respect to detected products. Products levels were determined relative to a housekeeping gene (actin) and were normalized to the DMSO control (n ≥ 3 biological replicates; mean ± s.d, * P ≤ 0.05, *** P ≤ 0.001, two-tailed Student’s t -test). (E) Low O-GlcNAc favors the non-DI pathway to increase functional OGT mRNA and non-functional OGA mRNA. High O-GlcNAc favors formation of DIs, the splicing of which, increases non-functional OGT and functional OGA .

    Journal: bioRxiv

    Article Title: O-GlcNAc regulates gene expression by controlling detained intron splicing

    doi: 10.1101/2020.03.27.012781

    Figure Lengend Snippet: O-GlcNAc levels regulate OGT and OGA DI splicing so that exon inclusion/skipping results in opposite effects on productive message. (A) Splice junctions derived from RNA-seq for OGT and OGA show two predominant fully-spliced isoforms, a functional one for OGT (A) and a non-functional one for OGA (B) . HEK-293T cells were treated with DMSO (white bars), OGT inhibitor (10 μM OSMI-2, dark grey) or OGA inhibitor (5 μM TMG, light grey) for 2 h and RT-qPCR was used to measure levels of the indicated products for OGT (C) or OGA (D) , which report on the presence of DIs as well as skipped or included exons. Red arrows depict location of primers with respect to detected products. Products levels were determined relative to a housekeeping gene (actin) and were normalized to the DMSO control (n ≥ 3 biological replicates; mean ± s.d, * P ≤ 0.05, *** P ≤ 0.001, two-tailed Student’s t -test). (E) Low O-GlcNAc favors the non-DI pathway to increase functional OGT mRNA and non-functional OGA mRNA. High O-GlcNAc favors formation of DIs, the splicing of which, increases non-functional OGT and functional OGA .

    Article Snippet: Total RNA extracted per fraction was quantified and used to calculate the percentage of mRNA localization for the RT–qPCR quantification.

    Techniques: Derivative Assay, RNA Sequencing Assay, Functional Assay, Quantitative RT-PCR, Two Tailed Test