mirneasy micro kit  (Qiagen)


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    Name:
    miRNeasy Micro Kit
    Description:
    For purification of miRNA and total RNA from small amounts of cells and tissues. Kit contents: Qiagen miRNeasy Micro Kit, 50 preps, Easy, Robust Procedures, Automatable Protocols, For Purification of miRNA and Total RNA from Small Amounts of Cells and Tissues, High-purity RNA Suitable, Ideal for Northern Blot Analysis, Quantitative, Real-time RT-PCR, Microarray Analysis, Ideal for Northern Blot Analysis, Quantitative, Real-time RT-PCR, Microarray Analysis, Includes 50 RNeasy minElute Spin Columns, Collection Tubes (1.5mL and 2mL), QIAzol Lysis Reagent, RNase-free Reagents and Buffers. Benefits: Effective purification of miRNA and total RNA from small samples. High-purity RNA suitable for all downstream applications. Easy, robust procedures. Automatable protocol
    Catalog Number:
    217084
    Price:
    None
    Category:
    miRNeasy Micro Kit
    Score:
    85
    Buy from Supplier


    Structured Review

    Qiagen mirneasy micro kit
    miRNeasy Micro Kit
    For purification of miRNA and total RNA from small amounts of cells and tissues. Kit contents: Qiagen miRNeasy Micro Kit, 50 preps, Easy, Robust Procedures, Automatable Protocols, For Purification of miRNA and Total RNA from Small Amounts of Cells and Tissues, High-purity RNA Suitable, Ideal for Northern Blot Analysis, Quantitative, Real-time RT-PCR, Microarray Analysis, Ideal for Northern Blot Analysis, Quantitative, Real-time RT-PCR, Microarray Analysis, Includes 50 RNeasy minElute Spin Columns, Collection Tubes (1.5mL and 2mL), QIAzol Lysis Reagent, RNase-free Reagents and Buffers. Benefits: Effective purification of miRNA and total RNA from small samples. High-purity RNA suitable for all downstream applications. Easy, robust procedures. Automatable protocol
    https://www.bioz.com/result/mirneasy micro kit/product/Qiagen
    Average 99 stars, based on 480 article reviews
    Price from $9.99 to $1999.99
    mirneasy micro kit - by Bioz Stars, 2019-12
    99/100 stars

    Images

    1) Product Images from "Optimisation of laboratory methods for whole transcriptomic RNA analyses in human left ventricular biopsies and blood samples of clinical relevance"

    Article Title: Optimisation of laboratory methods for whole transcriptomic RNA analyses in human left ventricular biopsies and blood samples of clinical relevance

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0213685

    Capillary electrophoresis analysis of pig LV biopsy RNA. ( A) Bioanalyzer nano chip traces of all tested samples. Inspection of electropherograms provides additional information to assess RNA quality and protocol performance. (B) Representative traces from each protocol with respective RIN values. Numbers 1–4 correspond to numbered boxes in A. Note that trace 3 has a high RIN despite a large smear in the gel. (C) Summary of RIN values of all the samples tested (mean±SEM). The RNAqueous protocol had undetectable RINs. The mir Vana and miRCURY tissue protocols gave consistently high RIN values, while despite high average RIN, the samples from the miRNeasy micro protocol are more variable. Numbers in brackets indicate the number of samples in each group. (D) Bioanalyzer small RNA chip traces for each kit. Although the RIN of samples marked * were similar (8.6–8.7), the spectrum of small RNAs recovered is highly heterogeneous.
    Figure Legend Snippet: Capillary electrophoresis analysis of pig LV biopsy RNA. ( A) Bioanalyzer nano chip traces of all tested samples. Inspection of electropherograms provides additional information to assess RNA quality and protocol performance. (B) Representative traces from each protocol with respective RIN values. Numbers 1–4 correspond to numbered boxes in A. Note that trace 3 has a high RIN despite a large smear in the gel. (C) Summary of RIN values of all the samples tested (mean±SEM). The RNAqueous protocol had undetectable RINs. The mir Vana and miRCURY tissue protocols gave consistently high RIN values, while despite high average RIN, the samples from the miRNeasy micro protocol are more variable. Numbers in brackets indicate the number of samples in each group. (D) Bioanalyzer small RNA chip traces for each kit. Although the RIN of samples marked * were similar (8.6–8.7), the spectrum of small RNAs recovered is highly heterogeneous.

    Techniques Used: Electrophoresis, Chromatin Immunoprecipitation

    Evaluating plasma RNA yield using RT-qPCR for miRNAs. As biofluid RNA is below the detection limit of standard quantification tools, all RT-qPCRs were performed using 1.5 μL RNA extracted from 200 μL plasma. Lines indicate paired samples i.e. aliquots of plasma from the same blood draw. (A) RT-qPCR for the spike-in cel-miR-39-3p using the four kits with and without glycogen. (B-D) RT-qPCR for three endogenous miRNAs isolated using the four kits with and without glycogen. (E) The miRNeasy serum/plasma protocol recommends elution in 14 μL, compared with 100 μL for the mir Vana protocol. To control for effects of this, RNA extractions were performed with both protocols, eluting in 30 μL or 50 μL. RT-qPCR for cel-miR-39-3p, hsa-miR-16-5p and hsa-miR-21-5p were performed to evaluate the different elution volumes.
    Figure Legend Snippet: Evaluating plasma RNA yield using RT-qPCR for miRNAs. As biofluid RNA is below the detection limit of standard quantification tools, all RT-qPCRs were performed using 1.5 μL RNA extracted from 200 μL plasma. Lines indicate paired samples i.e. aliquots of plasma from the same blood draw. (A) RT-qPCR for the spike-in cel-miR-39-3p using the four kits with and without glycogen. (B-D) RT-qPCR for three endogenous miRNAs isolated using the four kits with and without glycogen. (E) The miRNeasy serum/plasma protocol recommends elution in 14 μL, compared with 100 μL for the mir Vana protocol. To control for effects of this, RNA extractions were performed with both protocols, eluting in 30 μL or 50 μL. RT-qPCR for cel-miR-39-3p, hsa-miR-16-5p and hsa-miR-21-5p were performed to evaluate the different elution volumes.

    Techniques Used: Quantitative RT-PCR, Isolation

    Spectrophotometry analysis of pig LV biopsy RNA. RNA extracted according to each manufacturer’s protocol, with the addition of a proteinase K digestion in half the samples. n = 3 biopsies/experimental condition, mean±SEM. N . B . Exiqon miRCURY tissue protocol requires proteinase K, therefore no–proteinase K condition was done with this kit. (A) RNA yield, assessed as ng RNA recovered per mg input tissue, was not significantly different between the protocols (Kruskal-Wallis test with Dunn correction), and additional proteinase K digestion in the RNAqueous micro, mir Vana and miRNeasy micro protocols did not significantly improve RNA yield (Kruskal-Wallis test with Dunn correction). (B) 260/280 and 260/230 ratios provide an assessment of the purity of the RNA. Ratio
    Figure Legend Snippet: Spectrophotometry analysis of pig LV biopsy RNA. RNA extracted according to each manufacturer’s protocol, with the addition of a proteinase K digestion in half the samples. n = 3 biopsies/experimental condition, mean±SEM. N . B . Exiqon miRCURY tissue protocol requires proteinase K, therefore no–proteinase K condition was done with this kit. (A) RNA yield, assessed as ng RNA recovered per mg input tissue, was not significantly different between the protocols (Kruskal-Wallis test with Dunn correction), and additional proteinase K digestion in the RNAqueous micro, mir Vana and miRNeasy micro protocols did not significantly improve RNA yield (Kruskal-Wallis test with Dunn correction). (B) 260/280 and 260/230 ratios provide an assessment of the purity of the RNA. Ratio

    Techniques Used: Spectrophotometry

    Summary of experimental design for Figs 2 , 3 and 4 . (A) Left ventricular biopsies were taken from three pigs undergoing cardio-pulmonary bypass (CPB). 7 biopsies were taken per pig. RNA was isolated from the biopsies using four commercially available kits; RNAqueous micro, mir Vana, miRCURY tissue, miRNeasy micro. Three kits (RNAqueous micro, mir Vana, miRNeasy micro) were tested with and without an additional proteinase K digestion step, giving 7 protocols in total. The miRCURY tissue protocol includes a proteinase K digestion, therefore this kit was not tested without this. RNA samples were characterised by spectrophotometry and capillary electrophoresis. (B) Blood samples were taken from three pigs undergoing CPB and processed to plasma. RNA was isolated from the plasma using four commercially-available kits; miRNeasy serum/plasma, mir Vana, miRCURY biofluids, Norgen Biotek plasma/serum. Each kit was tested with and without glycogen as a co-precipitant, giving 8 protocols in total. RNA samples were evaluated by RT-qPCR for the exogenous spike-in cel-miR-39-3p, and endogenous miRNAs miR-16-5p, miR-21-5p, miR-92a-3p.
    Figure Legend Snippet: Summary of experimental design for Figs 2 , 3 and 4 . (A) Left ventricular biopsies were taken from three pigs undergoing cardio-pulmonary bypass (CPB). 7 biopsies were taken per pig. RNA was isolated from the biopsies using four commercially available kits; RNAqueous micro, mir Vana, miRCURY tissue, miRNeasy micro. Three kits (RNAqueous micro, mir Vana, miRNeasy micro) were tested with and without an additional proteinase K digestion step, giving 7 protocols in total. The miRCURY tissue protocol includes a proteinase K digestion, therefore this kit was not tested without this. RNA samples were characterised by spectrophotometry and capillary electrophoresis. (B) Blood samples were taken from three pigs undergoing CPB and processed to plasma. RNA was isolated from the plasma using four commercially-available kits; miRNeasy serum/plasma, mir Vana, miRCURY biofluids, Norgen Biotek plasma/serum. Each kit was tested with and without glycogen as a co-precipitant, giving 8 protocols in total. RNA samples were evaluated by RT-qPCR for the exogenous spike-in cel-miR-39-3p, and endogenous miRNAs miR-16-5p, miR-21-5p, miR-92a-3p.

    Techniques Used: Isolation, Spectrophotometry, Electrophoresis, Quantitative RT-PCR

    Related Articles

    Amplification:

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    Synthesized:

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    Construct:

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    Real-time Polymerase Chain Reaction:

    Article Title: CD4+ T cells with an activated and exhausted phenotype distinguish immunodeficiency during aviremic HIV-2 infection
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    Article Title: Unique long non-coding RNA expression signature in ETV6/RUNX1-driven B-cell precursor acute lymphoblastic leukemia
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    Microarray:

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    Incubation:

    Article Title: Mesenchymal stem cells release exosomes that transfer miRNAs to endothelial cells and promote angiogenesis
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    Expressing:

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    Hybridization:

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    Transfection:

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    One-tailed Test:

    Article Title: Modulation of Hematopoietic Lineage Specification Impacts TREM2 Expression in Microglia-Like Cells Derived From Human Stem Cells
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    Dissection:

    Article Title: Cis-regulatory architecture of a brain-signaling center predates the origin of chordates
    Article Snippet: The 429M20eGFP BAC reporter line was used to guide the dissection of Shh expressing cells from the ventral midbrain, ventroposterior diencephalon and zli of E10.5 embryos under a fluorescent stereomicroscope. .. Total RNA was extracted from GFP+ brain tissue isolated from approximately 30 embryos using the miRNeasy Micro Kit (Qiagen, Valencia, CA).

    Cell Culture:

    Article Title: Genetic deficiency of Wnt5a diminishes disease severity in a murine model of rheumatoid arthritis
    Article Snippet: BMDM were isolated from female C57BL6J mice (Jackson Laboratories) and cultured in alpha-MEM with 10% FBS and pen/strep/Lglut. .. RNA was isolated at d2 and d5 using the miRNeasy Micro Kit (Qiagen) and 1 μg of RNA was transcribed using the Quantitect kit (Qiagen).

    Reverse Transcription Polymerase Chain Reaction:

    Article Title: Assembly and Validation of Versatile Transcription Activator-Like Effector Libraries
    Article Snippet: Paragraph title: Quantitative reverse transcription-PCR (rtPCR) ... For measurement of miRNAs in mammalian cells, total RNA was extracted using the miRNeasy Micro Kit (Qiagen, #217084).

    Generated:

    Article Title: Extracellular RNAs: development as biomarkers of human disease
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    Article Title: Modulation of nonsense mediated decay by rapamycin
    Article Snippet: RNA was extracted from cytoplasmic extracts employing TRIzol LS following manufacturer's instructions, and re-purified and DNAse treated employing miRNeasy Micro Kit (Qiagen) following manufacturer's instructions. .. RNA was extracted from cytoplasmic extracts employing TRIzol LS following manufacturer's instructions, and re-purified and DNAse treated employing miRNeasy Micro Kit (Qiagen) following manufacturer's instructions.

    Sequencing:

    Article Title: Absence of Functional Leptin Receptor Isoforms in the POUND (Leprdb/lb) Mouse Is Associated with Muscle Atrophy and Altered Myoblast Proliferation and Differentiation
    Article Snippet: For miRNA isolation, miRNeasy Micro kit (Qiagen, CA) was utilized. .. The microarray analysis was performed at the Georgia Health Science University Microarray Core facility.

    Article Title: Cis-regulatory architecture of a brain-signaling center predates the origin of chordates
    Article Snippet: Total RNA was extracted from GFP+ brain tissue isolated from approximately 30 embryos using the miRNeasy Micro Kit (Qiagen, Valencia, CA). .. The RNA-seq library was prepared from 1μg of total RNA according to the manufacturer’s protocol for TruSeq RNA Sample Prep Kits (Illumina).

    Article Title: Modulation of nonsense mediated decay by rapamycin
    Article Snippet: RNA was extracted from cytoplasmic extracts employing TRIzol LS following manufacturer's instructions, and re-purified and DNAse treated employing miRNeasy Micro Kit (Qiagen) following manufacturer's instructions. .. RNA was extracted from cytoplasmic extracts employing TRIzol LS following manufacturer's instructions, and re-purified and DNAse treated employing miRNeasy Micro Kit (Qiagen) following manufacturer's instructions.

    Article Title: High content image analysis reveals function of miR-124 upstream of Vimentin in regulating motor neuron mitochondria
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    Article Title: The 3'-5' exoribonuclease Dis3 regulates the expression of specific microRNAs in Drosophila wing imaginal discs
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    Software:

    Article Title: High content image analysis reveals function of miR-124 upstream of Vimentin in regulating motor neuron mitochondria
    Article Snippet: Total RNA was isolated with miRNeasy micro kit (Qiagen), assessed with Nano Drop ND-1000 Spectrophotometer (Peqlab) and reverse transcribed to cDNA. qPCR, performed with SYBR Green (Thermo Fisher Scientific or Qiagen). miRNA/mRNA levels were normalized to U6/hypoxanthine phosphoribosyltransferase 1 (Hprt), respectively. .. Library prep for NGS, following [46] and sequencing performed on Illumina 2500 at 50 bp single read.

    Article Title: Unique long non-coding RNA expression signature in ETV6/RUNX1-driven B-cell precursor acute lymphoblastic leukemia
    Article Snippet: Total RNA was isolated using the miRNeasy mini or micro kit (Qiagen) according to the manufacturer's instructions. .. After cDNA preparation, reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) was carried out using custom 2X SsoAdvanced SYBR Green Supermix (Bio-Rad) in two technical replicates.

    RNA Sequencing Assay:

    Article Title: Extracellular RNAs: development as biomarkers of human disease
    Article Snippet: RNA isolation techniques were optimized for salivary RNA isolation efficiency by systematically comparing 6 commercially available kits with optimized protocols: (a) organic extraction method (Trizol LS); (b) spin filter based method [QIAamp Viral (Qiagen), NucleoSpin (Clontech) and miRVana (Life Technologies)]; and (c) combined method of organic extraction and spin filter clean up (miRNeasy Micro Kit (Qiagen), Quick-RNA MicroPrep (Zymo Research). .. RNA isolation techniques were optimized for salivary RNA isolation efficiency by systematically comparing 6 commercially available kits with optimized protocols: (a) organic extraction method (Trizol LS); (b) spin filter based method [QIAamp Viral (Qiagen), NucleoSpin (Clontech) and miRVana (Life Technologies)]; and (c) combined method of organic extraction and spin filter clean up (miRNeasy Micro Kit (Qiagen), Quick-RNA MicroPrep (Zymo Research).

    Article Title: Cis-regulatory architecture of a brain-signaling center predates the origin of chordates
    Article Snippet: Paragraph title: RNA-seq ... Total RNA was extracted from GFP+ brain tissue isolated from approximately 30 embryos using the miRNeasy Micro Kit (Qiagen, Valencia, CA).

    Article Title: Modulation of nonsense mediated decay by rapamycin
    Article Snippet: Paragraph title: RNA-seq ... RNA was extracted from cytoplasmic extracts employing TRIzol LS following manufacturer's instructions, and re-purified and DNAse treated employing miRNeasy Micro Kit (Qiagen) following manufacturer's instructions.

    Isolation:

    Article Title: Absence of Functional Leptin Receptor Isoforms in the POUND (Leprdb/lb) Mouse Is Associated with Muscle Atrophy and Altered Myoblast Proliferation and Differentiation
    Article Snippet: Total RNA was extracted using a standard chloroform protocol. .. For miRNA isolation, miRNeasy Micro kit (Qiagen, CA) was utilized. .. The microarray analysis was performed at the Georgia Health Science University Microarray Core facility.

    Article Title: Extracellular RNAs: development as biomarkers of human disease
    Article Snippet: Finally, the newly configured salivary exRNA biomarker panel for gastric cancer detection will be validated in an independent set of 250 gastric cancer and 250 non-gastric cancer matched control subjects. .. RNA isolation techniques were optimized for salivary RNA isolation efficiency by systematically comparing 6 commercially available kits with optimized protocols: (a) organic extraction method (Trizol LS); (b) spin filter based method [QIAamp Viral (Qiagen), NucleoSpin (Clontech) and miRVana (Life Technologies)]; and (c) combined method of organic extraction and spin filter clean up (miRNeasy Micro Kit (Qiagen), Quick-RNA MicroPrep (Zymo Research). .. The quantity and size distributions of the resulting RNA samples were assessed using RiboGreen reagent and Bioanalyzer, respectively, with the best yields from NucleoSpin and miRNeasy Micro kits. qPCR and ddPCR were used to determine the efficiency of long and small RNA isolation from each kit; the studies revealed that the miRNeasy micro Kit and NucleoSpin are the best kits in yielding small RNAs at the same time as long RNAs.

    Article Title: miR-31 is distinctively overexpressed in primary male extramammary Paget's disease
    Article Snippet: Paragraph title: RNA isolation ... All the collected tissue of these three groups underwent lysis and extraction of total RNA including miRNA by Qiagen miRNeasy Micro Kit with the protocol provided by Qiagen.

    Article Title: Cis-regulatory architecture of a brain-signaling center predates the origin of chordates
    Article Snippet: The 429M20eGFP BAC reporter line was used to guide the dissection of Shh expressing cells from the ventral midbrain, ventroposterior diencephalon and zli of E10.5 embryos under a fluorescent stereomicroscope. .. Total RNA was extracted from GFP+ brain tissue isolated from approximately 30 embryos using the miRNeasy Micro Kit (Qiagen, Valencia, CA). .. The RNA-seq library was prepared from 1μg of total RNA according to the manufacturer’s protocol for TruSeq RNA Sample Prep Kits (Illumina).

    Article Title: Splicing Factor 1 Modulates Dietary Restriction and TORC1 Pathway Longevity in C. elegans
    Article Snippet: Paragraph title: RNA isolation and cDNA synthesis ... Total RNA was extracted using Qiazol reagent (QIAGEN), column purified by RNeasy mini or miRNeasy micro kit (QIAGEN) according to manufacturer’s instructions. cDNA was synthesized using SuperScript® VILO Master mix (Invitrogen).

    Article Title: Mesenchymal stem cells release exosomes that transfer miRNAs to endothelial cells and promote angiogenesis
    Article Snippet: The plate was placed into the IncuCyte ZOOM Live Content Imaging System (ESSEN BIOSCIENCE) installed inside an incubator and recorded every 2 h for 12 h. .. Total RNA was isolated from concentrated CdM, exosomes, MSCs, and HUVECs using miRNeasy Micro Kit (Qiagen). .. Briefly, 200 μl CdM, exosome pellet or cell pellet was lysed by 1 ml QIAzol Lysis Reagent.

    Article Title: Genetic deficiency of Wnt5a diminishes disease severity in a murine model of rheumatoid arthritis
    Article Snippet: BMDM were induced to fuse with the stimulation cocktail (MCSF (50 ng/mL) and RANKL (50 ng/mL)), which was replenished at d2 and d4. .. RNA was isolated at d2 and d5 using the miRNeasy Micro Kit (Qiagen) and 1 μg of RNA was transcribed using the Quantitect kit (Qiagen). .. SYBR chemistry was used to detect transcripts and these were analyzed on a ViiA7 using the primers listed in Table .

    Article Title: High content image analysis reveals function of miR-124 upstream of Vimentin in regulating motor neuron mitochondria
    Article Snippet: For transmission electron microscopy, motor neuron were prepared, following and electron micrographs were captured with a FEI Tecnai SPIRIT transmission electron microscope (FEI, Eidhoven, Netherlands), operated at 120 kV and equipped with an EAGLE CCD Camera. .. Total RNA was isolated with miRNeasy micro kit (Qiagen), assessed with Nano Drop ND-1000 Spectrophotometer (Peqlab) and reverse transcribed to cDNA. qPCR, performed with SYBR Green (Thermo Fisher Scientific or Qiagen). miRNA/mRNA levels were normalized to U6/hypoxanthine phosphoribosyltransferase 1 (Hprt), respectively. .. Primer sequences are described in Table .

    Article Title: Effects of cell adhesion motif, fiber stiffness, and cyclic strain on tenocyte gene expression in a tendon mimetic fiber composite hydrogel
    Article Snippet: Composites were placed into individual wells in 6-well plates (free-swelling) or a sterile custom bioreactor (loaded) [35] in an incubator at 37 °C and 5% CO2 , and left to stabilize for 24 h. The loaded sample was subjected to a 5% amplitude strain applied in a sinusoidal waveform at 1 Hz continuously for 24 h. For each substrate stiffness and peptide motif, there were two loaded samples and two free swelling samples per biological replicate. .. 2.8 RNA extraction and RT-qPCR RNA was extracted using a miRNeasy Micro Kit (QIAgen, USA) from freshly isolated tenocytes of each donor (referred to as donor tenocytes) and from tenocyte-seeded fiber composites at experiment's end. .. Samples were snapped frozen in liquid nitrogen and homogenized (TissueLyser II).

    Article Title: miR-494-3p overexpression promotes megakaryocytopoiesis in primary myelofibrosis hematopoietic stem/progenitor cells by targeting SOCS6
    Article Snippet: SOCS6 or miR-494-3p expression were analyzed 24 hours upon the last nucleofection. .. Total cellular RNA was harvested from 5×104 cells from each sample using the miRNeasy Micro RNA isolation kit (QIAGEN), according to the manufacturer's instructions and as previously described [ ]. .. Relative quantification (RQ) of mRNA and miRNA expression levels was performed as previously described [ ].

    Article Title: Deregulated expression of miR-29a-3p, miR-494-3p and miR-660-5p affects sensitivity to tyrosine kinase inhibitors in CML leukemic stem cells
    Article Snippet: As previously reported [ ], FISH analysis and RQ-PCR for BCR-ABL transcript demonstrated that Ph+ cells were 70.5% (± 5%), and 32% (± 2%) of total Lin-CD34+CD38− and Lin-CD34-CD38- CML cells at diagnosis, respectively. .. Total cellular RNA was harvested from 1 × 105 cells from each sample using the miRNeasy Micro RNA isolation kit (QIAGEN), according to the manufacturer's instructions. .. RNA samples concentration and purity (assessed as 260/280 nm and 260/230 nm ratios) were evaluated by NanoDrop ND-1000 spectrophotometer (NanoDrop Technologies; Wilmington, DE), while RNA integrity was assessed by using the Agilent 2100 Bioanalyzer (Agilent Technologies; Waldbrunn, Germany).

    Article Title: Unique long non-coding RNA expression signature in ETV6/RUNX1-driven B-cell precursor acute lymphoblastic leukemia
    Article Snippet: Cells were cultured in RPMI 1640 media (Life Technologies Europe) supplemented with 10% fetal bovine serum (Biochrom AG), 1% penicillin/streptomycin (Life Technologies Europe), 1% kanamycin (Life Technologies Europe), 1% glutamine (Life Technologies Europe) at 37°C in 5% CO2. .. Total RNA was isolated using the miRNeasy mini or micro kit (Qiagen) according to the manufacturer's instructions. .. For each sample, RNA quality and purity were assessed by Experion analysis (Bio-Rad, Nazareth Eke, Belgium) and concentration was measured using the NanoDrop ND-1000 (NanoDrop Technologies, Wilmington, DE, USA). cDNA was generated using the iScript cDNA synthesis kit (Bio-Rad, Nazareth Eke, Belgium) according to the instructions of the manufacturer.

    Microscopy:

    Article Title: Genetic deficiency of Wnt5a diminishes disease severity in a murine model of rheumatoid arthritis
    Article Snippet: RNA was isolated at d2 and d5 using the miRNeasy Micro Kit (Qiagen) and 1 μg of RNA was transcribed using the Quantitect kit (Qiagen). .. To enumerate osteoclasts, d5 cultures were stained for TRAP (Sigma) and co-stained with 4',6-diamidino-2-phenylindole (DAPI) (Thermo Fisher Scientific).

    Purification:

    Article Title: Absence of Functional Leptin Receptor Isoforms in the POUND (Leprdb/lb) Mouse Is Associated with Muscle Atrophy and Altered Myoblast Proliferation and Differentiation
    Article Snippet: For miRNA isolation, miRNeasy Micro kit (Qiagen, CA) was utilized. .. The microarray analysis was performed at the Georgia Health Science University Microarray Core facility.

    Article Title: Splicing Factor 1 Modulates Dietary Restriction and TORC1 Pathway Longevity in C. elegans
    Article Snippet: These lines are named SFA-1 OE line 3 and SFA-1 OE line 4. .. Total RNA was extracted using Qiazol reagent (QIAGEN), column purified by RNeasy mini or miRNeasy micro kit (QIAGEN) according to manufacturer’s instructions. cDNA was synthesized using SuperScript® VILO Master mix (Invitrogen). .. WT (N2) and eat-2(ad1116) (DR) worms were synchronized on HT115 empty vector (ev) or sfa-1 RNAi bacteria by egg lay.

    Article Title: Assembly and Validation of Versatile Transcription Activator-Like Effector Libraries
    Article Snippet: For measurement of mRNAs in yeast cells, the same protocol was followed, except that total RNA was extracted using the MasterPure Yeast Purification Kit (Epicentre, #MPY80200), and yeast ACT1 was used for normalization. .. For measurement of miRNAs in mammalian cells, total RNA was extracted using the miRNeasy Micro Kit (Qiagen, #217084).

    Polymerase Chain Reaction:

    Article Title: CD4+ T cells with an activated and exhausted phenotype distinguish immunodeficiency during aviremic HIV-2 infection
    Article Snippet: With minor modifications, HIV-1 and HIV-2 plasma viral loads were determined by in-house quantitiative PCR (qPCR) protocols as described [ ]. .. Briefly, viral RNA was extracted using miRNeasy micro Kit (Qiagen, Hilden, Germany), and TaqMan qRT-PCR was performed using the Superscript III Platinum One Step qRT-PCR kit (Life Technologies, Carlsbad, California, USA).

    Article Title: Mesenchymal stem cells release exosomes that transfer miRNAs to endothelial cells and promote angiogenesis
    Article Snippet: Total RNA was isolated from concentrated CdM, exosomes, MSCs, and HUVECs using miRNeasy Micro Kit (Qiagen). .. Total RNA was isolated from concentrated CdM, exosomes, MSCs, and HUVECs using miRNeasy Micro Kit (Qiagen).

    Article Title: The 3'-5' exoribonuclease Dis3 regulates the expression of specific microRNAs in Drosophila wing imaginal discs
    Article Snippet: RNA extractions were performed using a miRNeasy RNA extraction kit (Qiagen, cat. no. 217084), with an on-column DNase digestion (Qiagen, cat. no. 79254). .. For qRT-PCR, RNA samples were diluted to a consistent concentration then cDNA was prepared in duplicate using a High Capacity cDNA Reverse Transcription Kit (Life Technologies, cat. no. 4368814) with random primers.

    Article Title: A novel role for the 3′-5′ exoribonuclease Dis3L2 in controlling cell proliferation and tissue growth
    Article Snippet: RNA extractions were performed using a miRNeasy RNA extraction kit (Qiagen, cat. no. 217084), with on-column DNAse digestion (Qiagen, cat. no. 79254). .. For qRT-PCR, 500ng of total RNA was converted to cDNA in duplicate using a High Capacity cDNA Reverse Transcription Kit (Life Technologies, cat. no. 4368814) with random primers or oligo(dT) primers.

    Article Title: Unique long non-coding RNA expression signature in ETV6/RUNX1-driven B-cell precursor acute lymphoblastic leukemia
    Article Snippet: Total RNA was isolated using the miRNeasy mini or micro kit (Qiagen) according to the manufacturer's instructions. .. After cDNA preparation, reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) was carried out using custom 2X SsoAdvanced SYBR Green Supermix (Bio-Rad) in two technical replicates.

    FACS:

    Article Title: Modulation of Hematopoietic Lineage Specification Impacts TREM2 Expression in Microglia-Like Cells Derived From Human Stem Cells
    Article Snippet: Differentiated ScMglia cells expressing TREM2 were sorted into PLO/Fn-coated 96-well plates by FACS and allowed to rest for 24 hr in serum-free specification medium. .. Wells were then washed with PBS and lysed with QIAzol reagent (QIAgen). mRNA was extracted using a miRNeasy Micro Kit (QIAgen) then amplified and transcribed to cDNA (Ovation Pico WTA System V2 kit, NuGEN) before being analyzed for expression of interleukin-1 beta (IL-1β, Applied Biosystems, #Hs00174097_m1), monocyte chemoattractant factor-1 (MCP-1 or CCL2, Applied Biosystems, #Hs00234140_m1), tumor necrosis factor (TNFα, Applied Biosystems, #Hs01113624_g1), interleukin-6 (IL-6, Applied Biosystems, #Hs00985639_m1), tumor necrosis factor alpha-induced protein 3 (TNFAIP3, Applied Biosystems, #Hs00234713_m1), and cluster of differentiation 68 (CD68, Applied Biosystems, #Hs02836816_g1).

    Quantitative RT-PCR:

    Article Title: CD4+ T cells with an activated and exhausted phenotype distinguish immunodeficiency during aviremic HIV-2 infection
    Article Snippet: With minor modifications, HIV-1 and HIV-2 plasma viral loads were determined by in-house quantitiative PCR (qPCR) protocols as described [ ]. .. Briefly, viral RNA was extracted using miRNeasy micro Kit (Qiagen, Hilden, Germany), and TaqMan qRT-PCR was performed using the Superscript III Platinum One Step qRT-PCR kit (Life Technologies, Carlsbad, California, USA). .. The detection limit for the viral loads was 75 RNA copies/ml plasma for HIV-1 or HIV-2 singly-infected, and 135 RNA copies/ml plasma for HIV-D-infected.

    Article Title: Effects of cell adhesion motif, fiber stiffness, and cyclic strain on tenocyte gene expression in a tendon mimetic fiber composite hydrogel
    Article Snippet: Composites were placed into individual wells in 6-well plates (free-swelling) or a sterile custom bioreactor (loaded) [35] in an incubator at 37 °C and 5% CO2 , and left to stabilize for 24 h. The loaded sample was subjected to a 5% amplitude strain applied in a sinusoidal waveform at 1 Hz continuously for 24 h. For each substrate stiffness and peptide motif, there were two loaded samples and two free swelling samples per biological replicate. .. 2.8 RNA extraction and RT-qPCR RNA was extracted using a miRNeasy Micro Kit (QIAgen, USA) from freshly isolated tenocytes of each donor (referred to as donor tenocytes) and from tenocyte-seeded fiber composites at experiment's end. .. Samples were snapped frozen in liquid nitrogen and homogenized (TissueLyser II).

    Article Title: The 3'-5' exoribonuclease Dis3 regulates the expression of specific microRNAs in Drosophila wing imaginal discs
    Article Snippet: Paragraph title: RNA extraction and qRT-PCR ... RNA extractions were performed using a miRNeasy RNA extraction kit (Qiagen, cat. no. 217084), with an on-column DNase digestion (Qiagen, cat. no. 79254).

    Article Title: A novel role for the 3′-5′ exoribonuclease Dis3L2 in controlling cell proliferation and tissue growth
    Article Snippet: Paragraph title: RNA extraction and qRT-PCR ... RNA extractions were performed using a miRNeasy RNA extraction kit (Qiagen, cat. no. 217084), with on-column DNAse digestion (Qiagen, cat. no. 79254).

    Article Title: miR-494-3p overexpression promotes megakaryocytopoiesis in primary myelofibrosis hematopoietic stem/progenitor cells by targeting SOCS6
    Article Snippet: Paragraph title: RNA extraction and qRT-PCR ... Total cellular RNA was harvested from 5×104 cells from each sample using the miRNeasy Micro RNA isolation kit (QIAGEN), according to the manufacturer's instructions and as previously described [ ].

    Article Title: Unique long non-coding RNA expression signature in ETV6/RUNX1-driven B-cell precursor acute lymphoblastic leukemia
    Article Snippet: Paragraph title: RNA isolation, cDNA synthesis and RT-qPCR ... Total RNA was isolated using the miRNeasy mini or micro kit (Qiagen) according to the manufacturer's instructions.

    Lysis:

    Article Title: miR-31 is distinctively overexpressed in primary male extramammary Paget's disease
    Article Snippet: By laser capture micro-dissection (MMI cellcut), EMPD tumor cells (ET), normal epidermal cells (NE) and normal apocrine gland cells (NA) were precisely isolated, respectively ( , available online) .. All the collected tissue of these three groups underwent lysis and extraction of total RNA including miRNA by Qiagen miRNeasy Micro Kit with the protocol provided by Qiagen. .. Due to limited RNA could be obtained using microdissection technique, a preamplification step was added per manufacture's protocol when miRNA array was performed in two paired samples of ET and NE from same patients for gross screening.

    Mouse Assay:

    Article Title: Genetic deficiency of Wnt5a diminishes disease severity in a murine model of rheumatoid arthritis
    Article Snippet: BMDM were isolated from female C57BL6J mice (Jackson Laboratories) and cultured in alpha-MEM with 10% FBS and pen/strep/Lglut. .. RNA was isolated at d2 and d5 using the miRNeasy Micro Kit (Qiagen) and 1 μg of RNA was transcribed using the Quantitect kit (Qiagen).

    TaqMan Assay:

    Article Title: The 3'-5' exoribonuclease Dis3 regulates the expression of specific microRNAs in Drosophila wing imaginal discs
    Article Snippet: RNA extractions were performed using a miRNeasy RNA extraction kit (Qiagen, cat. no. 217084), with an on-column DNase digestion (Qiagen, cat. no. 79254). .. A control “no RT” reaction was performed in parallel to confirm that all genomic DNA had been degraded. qRT-PCR was performed on each cDNA replicate in duplicate (i.e. 4 technical replicates in total), using TaqMan Universal PCR Master Mix, No AmpErase UNG (Life Technologies, cat. no. 4324018) and an appropriate custom designed TaqMan pre-miRNA assay (Life Technologies).

    Article Title: A novel role for the 3′-5′ exoribonuclease Dis3L2 in controlling cell proliferation and tissue growth
    Article Snippet: RNA extractions were performed using a miRNeasy RNA extraction kit (Qiagen, cat. no. 217084), with on-column DNAse digestion (Qiagen, cat. no. 79254). .. RNA extractions were performed using a miRNeasy RNA extraction kit (Qiagen, cat. no. 217084), with on-column DNAse digestion (Qiagen, cat. no. 79254).

    SYBR Green Assay:

    Article Title: High content image analysis reveals function of miR-124 upstream of Vimentin in regulating motor neuron mitochondria
    Article Snippet: For transmission electron microscopy, motor neuron were prepared, following and electron micrographs were captured with a FEI Tecnai SPIRIT transmission electron microscope (FEI, Eidhoven, Netherlands), operated at 120 kV and equipped with an EAGLE CCD Camera. .. Total RNA was isolated with miRNeasy micro kit (Qiagen), assessed with Nano Drop ND-1000 Spectrophotometer (Peqlab) and reverse transcribed to cDNA. qPCR, performed with SYBR Green (Thermo Fisher Scientific or Qiagen). miRNA/mRNA levels were normalized to U6/hypoxanthine phosphoribosyltransferase 1 (Hprt), respectively. .. Primer sequences are described in Table .

    Article Title: Effects of cell adhesion motif, fiber stiffness, and cyclic strain on tenocyte gene expression in a tendon mimetic fiber composite hydrogel
    Article Snippet: 2.8 RNA extraction and RT-qPCR RNA was extracted using a miRNeasy Micro Kit (QIAgen, USA) from freshly isolated tenocytes of each donor (referred to as donor tenocytes) and from tenocyte-seeded fiber composites at experiment's end. .. 2.8 RNA extraction and RT-qPCR RNA was extracted using a miRNeasy Micro Kit (QIAgen, USA) from freshly isolated tenocytes of each donor (referred to as donor tenocytes) and from tenocyte-seeded fiber composites at experiment's end.

    Article Title: Assembly and Validation of Versatile Transcription Activator-Like Effector Libraries
    Article Snippet: For measurement of miRNAs in mammalian cells, total RNA was extracted using the miRNeasy Micro Kit (Qiagen, #217084). .. For measurement of miRNAs in mammalian cells, total RNA was extracted using the miRNeasy Micro Kit (Qiagen, #217084).

    Article Title: Unique long non-coding RNA expression signature in ETV6/RUNX1-driven B-cell precursor acute lymphoblastic leukemia
    Article Snippet: Total RNA was isolated using the miRNeasy mini or micro kit (Qiagen) according to the manufacturer's instructions. .. For each sample, RNA quality and purity were assessed by Experion analysis (Bio-Rad, Nazareth Eke, Belgium) and concentration was measured using the NanoDrop ND-1000 (NanoDrop Technologies, Wilmington, DE, USA). cDNA was generated using the iScript cDNA synthesis kit (Bio-Rad, Nazareth Eke, Belgium) according to the instructions of the manufacturer.

    RNA Extraction:

    Article Title: Gene expression profiling in pbMEC – in search of molecular biomarkers to predict immunoglobulin production in bovine milk
    Article Snippet: Paragraph title: RNA extraction and reverse transcription ... RNA was extracted using the miRNeasy Micro Kit (Qiagen), according to the manufacturer’s protocol with slight modifications.

    Article Title: Effects of cell adhesion motif, fiber stiffness, and cyclic strain on tenocyte gene expression in a tendon mimetic fiber composite hydrogel
    Article Snippet: Composites were placed into individual wells in 6-well plates (free-swelling) or a sterile custom bioreactor (loaded) [35] in an incubator at 37 °C and 5% CO2 , and left to stabilize for 24 h. The loaded sample was subjected to a 5% amplitude strain applied in a sinusoidal waveform at 1 Hz continuously for 24 h. For each substrate stiffness and peptide motif, there were two loaded samples and two free swelling samples per biological replicate. .. 2.8 RNA extraction and RT-qPCR RNA was extracted using a miRNeasy Micro Kit (QIAgen, USA) from freshly isolated tenocytes of each donor (referred to as donor tenocytes) and from tenocyte-seeded fiber composites at experiment's end. .. Samples were snapped frozen in liquid nitrogen and homogenized (TissueLyser II).

    Article Title: The 3'-5' exoribonuclease Dis3 regulates the expression of specific microRNAs in Drosophila wing imaginal discs
    Article Snippet: Cy3-conjugated monoclonal donkey anti-mouse IgG secondary antibody was used at 1:400 (Jackson ImmunoResearch, cat. no.715–165–151). .. RNA extractions were performed using a miRNeasy RNA extraction kit (Qiagen, cat. no. 217084), with an on-column DNase digestion (Qiagen, cat. no. 79254). .. RNA concentrations were measured on a NanoDrop 1000 spectrophotometer (Thermo Scientific).

    Article Title: A novel role for the 3′-5′ exoribonuclease Dis3L2 in controlling cell proliferation and tissue growth
    Article Snippet: The mitotic index was then calculated for each disc by dividing the number of cells in M phase by the area of the disc. .. RNA extractions were performed using a miRNeasy RNA extraction kit (Qiagen, cat. no. 217084), with on-column DNAse digestion (Qiagen, cat. no. 79254). .. RNA concentrations were measured on a NanoDrop1000 spectrophotometer (Thermo Scientific).

    Article Title: miR-494-3p overexpression promotes megakaryocytopoiesis in primary myelofibrosis hematopoietic stem/progenitor cells by targeting SOCS6
    Article Snippet: Paragraph title: RNA extraction and qRT-PCR ... Total cellular RNA was harvested from 5×104 cells from each sample using the miRNeasy Micro RNA isolation kit (QIAGEN), according to the manufacturer's instructions and as previously described [ ].

    Article Title: Deregulated expression of miR-29a-3p, miR-494-3p and miR-660-5p affects sensitivity to tyrosine kinase inhibitors in CML leukemic stem cells
    Article Snippet: Paragraph title: RNA extraction ... Total cellular RNA was harvested from 1 × 105 cells from each sample using the miRNeasy Micro RNA isolation kit (QIAGEN), according to the manufacturer's instructions.

    In Vitro:

    Article Title: Absence of Functional Leptin Receptor Isoforms in the POUND (Leprdb/lb) Mouse Is Associated with Muscle Atrophy and Altered Myoblast Proliferation and Differentiation
    Article Snippet: For miRNA isolation, miRNeasy Micro kit (Qiagen, CA) was utilized. .. The microarray analysis was performed at the Georgia Health Science University Microarray Core facility.

    Article Title: Genetic deficiency of Wnt5a diminishes disease severity in a murine model of rheumatoid arthritis
    Article Snippet: Paragraph title: In vitro analysis of fusion ... RNA was isolated at d2 and d5 using the miRNeasy Micro Kit (Qiagen) and 1 μg of RNA was transcribed using the Quantitect kit (Qiagen).

    Next-Generation Sequencing:

    Article Title: Cis-regulatory architecture of a brain-signaling center predates the origin of chordates
    Article Snippet: Total RNA was extracted from GFP+ brain tissue isolated from approximately 30 embryos using the miRNeasy Micro Kit (Qiagen, Valencia, CA). .. The RNA-seq library was prepared from 1μg of total RNA according to the manufacturer’s protocol for TruSeq RNA Sample Prep Kits (Illumina).

    Article Title: High content image analysis reveals function of miR-124 upstream of Vimentin in regulating motor neuron mitochondria
    Article Snippet: Paragraph title: RNA analysis and Next Generation Sequencing ... Total RNA was isolated with miRNeasy micro kit (Qiagen), assessed with Nano Drop ND-1000 Spectrophotometer (Peqlab) and reverse transcribed to cDNA. qPCR, performed with SYBR Green (Thermo Fisher Scientific or Qiagen). miRNA/mRNA levels were normalized to U6/hypoxanthine phosphoribosyltransferase 1 (Hprt), respectively.

    Quantitation Assay:

    Article Title: Cis-regulatory architecture of a brain-signaling center predates the origin of chordates
    Article Snippet: Total RNA was extracted from GFP+ brain tissue isolated from approximately 30 embryos using the miRNeasy Micro Kit (Qiagen, Valencia, CA). .. Total RNA was extracted from GFP+ brain tissue isolated from approximately 30 embryos using the miRNeasy Micro Kit (Qiagen, Valencia, CA).

    Spectrophotometry:

    Article Title: Gene expression profiling in pbMEC – in search of molecular biomarkers to predict immunoglobulin production in bovine milk
    Article Snippet: RNA was extracted using the miRNeasy Micro Kit (Qiagen), according to the manufacturer’s protocol with slight modifications. .. The miRNeasy Micro spin column was incubated for 5 min with buffer RPE after the second addition of buffer RPE to reduce contamination of the RNA with guanidine thiocyanate.

    Article Title: High content image analysis reveals function of miR-124 upstream of Vimentin in regulating motor neuron mitochondria
    Article Snippet: For transmission electron microscopy, motor neuron were prepared, following and electron micrographs were captured with a FEI Tecnai SPIRIT transmission electron microscope (FEI, Eidhoven, Netherlands), operated at 120 kV and equipped with an EAGLE CCD Camera. .. Total RNA was isolated with miRNeasy micro kit (Qiagen), assessed with Nano Drop ND-1000 Spectrophotometer (Peqlab) and reverse transcribed to cDNA. qPCR, performed with SYBR Green (Thermo Fisher Scientific or Qiagen). miRNA/mRNA levels were normalized to U6/hypoxanthine phosphoribosyltransferase 1 (Hprt), respectively. .. Primer sequences are described in Table .

    Concentration Assay:

    Article Title: Gene expression profiling in pbMEC – in search of molecular biomarkers to predict immunoglobulin production in bovine milk
    Article Snippet: RNA was extracted using the miRNeasy Micro Kit (Qiagen), according to the manufacturer’s protocol with slight modifications. .. The miRNeasy Micro spin column was incubated for 5 min with buffer RPE after the second addition of buffer RPE to reduce contamination of the RNA with guanidine thiocyanate.

    Article Title: The 3'-5' exoribonuclease Dis3 regulates the expression of specific microRNAs in Drosophila wing imaginal discs
    Article Snippet: RNA extractions were performed using a miRNeasy RNA extraction kit (Qiagen, cat. no. 217084), with an on-column DNase digestion (Qiagen, cat. no. 79254). .. RNA concentrations were measured on a NanoDrop 1000 spectrophotometer (Thermo Scientific).

    BAC Assay:

    Article Title: Cis-regulatory architecture of a brain-signaling center predates the origin of chordates
    Article Snippet: The 429M20eGFP BAC reporter line was used to guide the dissection of Shh expressing cells from the ventral midbrain, ventroposterior diencephalon and zli of E10.5 embryos under a fluorescent stereomicroscope. .. Total RNA was extracted from GFP+ brain tissue isolated from approximately 30 embryos using the miRNeasy Micro Kit (Qiagen, Valencia, CA).

    Staining:

    Article Title: Genetic deficiency of Wnt5a diminishes disease severity in a murine model of rheumatoid arthritis
    Article Snippet: RNA was isolated at d2 and d5 using the miRNeasy Micro Kit (Qiagen) and 1 μg of RNA was transcribed using the Quantitect kit (Qiagen). .. RNA was isolated at d2 and d5 using the miRNeasy Micro Kit (Qiagen) and 1 μg of RNA was transcribed using the Quantitect kit (Qiagen).

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    Qiagen mirneasy mini kit
    Correlations between mean “fold recovery” (FR) values of miRNAs and base 10 logarithm of miRNA relative abundance in sample (SRA), using yE protocol (A) and yQ protocol (B). yE, miRCURY RNA isolation kit Biofluids modified protocol using yeast RNA carrier; yQ, <t>miRNeasy</t> Mini kit modified protocol using yeast RNA carrier; FR, the fold recovery of each miRNA vs the same protocol without carrier; SRA, the mean relative abundance of each miRNA on samples isolated without carrier.
    Mirneasy Mini Kit, supplied by Qiagen, used in various techniques. Bioz Stars score: 99/100, based on 478 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Correlations between mean “fold recovery” (FR) values of miRNAs and base 10 logarithm of miRNA relative abundance in sample (SRA), using yE protocol (A) and yQ protocol (B). yE, miRCURY RNA isolation kit Biofluids modified protocol using yeast RNA carrier; yQ, miRNeasy Mini kit modified protocol using yeast RNA carrier; FR, the fold recovery of each miRNA vs the same protocol without carrier; SRA, the mean relative abundance of each miRNA on samples isolated without carrier.

    Journal: PLoS ONE

    Article Title: Comparison of protocols and RNA carriers for plasma miRNA isolation. Unraveling RNA carrier influence on miRNA isolation

    doi: 10.1371/journal.pone.0187005

    Figure Lengend Snippet: Correlations between mean “fold recovery” (FR) values of miRNAs and base 10 logarithm of miRNA relative abundance in sample (SRA), using yE protocol (A) and yQ protocol (B). yE, miRCURY RNA isolation kit Biofluids modified protocol using yeast RNA carrier; yQ, miRNeasy Mini kit modified protocol using yeast RNA carrier; FR, the fold recovery of each miRNA vs the same protocol without carrier; SRA, the mean relative abundance of each miRNA on samples isolated without carrier.

    Article Snippet: The quality and isolation efficiency of miRNAs were analyzed in four plasma samples using two protocols: phenol and column based isolation procedures (miRNeasy Mini kit, Qiagen, Q protocol) and column based procedures (miRCURY RNA Isolation kit Biofluids, Exiqon, E protocol).

    Techniques: Isolation, Modification

    Raw Cq values. Evaluation of Cq values of UniSp2, miR-103a-3p and miR-451a in four samples using different isolation protocols and RNA carriers. y, yeast RNA carrier; m, MS2 RNA carrier; w, without carrier; Q, miRNeasy Mini kit modified protocol; E, miRCURY RNA isolation kit Biofluids modified protocol. Mean and standard deviation values are indicated under each box plot diagram.

    Journal: PLoS ONE

    Article Title: Comparison of protocols and RNA carriers for plasma miRNA isolation. Unraveling RNA carrier influence on miRNA isolation

    doi: 10.1371/journal.pone.0187005

    Figure Lengend Snippet: Raw Cq values. Evaluation of Cq values of UniSp2, miR-103a-3p and miR-451a in four samples using different isolation protocols and RNA carriers. y, yeast RNA carrier; m, MS2 RNA carrier; w, without carrier; Q, miRNeasy Mini kit modified protocol; E, miRCURY RNA isolation kit Biofluids modified protocol. Mean and standard deviation values are indicated under each box plot diagram.

    Article Snippet: The quality and isolation efficiency of miRNAs were analyzed in four plasma samples using two protocols: phenol and column based isolation procedures (miRNeasy Mini kit, Qiagen, Q protocol) and column based procedures (miRCURY RNA Isolation kit Biofluids, Exiqon, E protocol).

    Techniques: Isolation, Modification, Standard Deviation

    Correlations between mean “fold recovery” (FR) values of miRNAs and ΔG of each miRNA, using yE protocol (A) and yQ protocol (B). To avoid the influence of GC content in the analysis of ΔG, for these correlations we used only miRNAs with a GC content between 25% and 75% percentile. yE, miRCURY RNA isolation kit Biofluids modified protocol using yeast RNA carrier; yQ, miRNeasy Mini kit modified protocol using yeast RNA carrier; FR, the fold recovery of each miRNA vs the same protocol without carrier; ΔG, the free energy of the most stable secondary structure of each miRNA.

    Journal: PLoS ONE

    Article Title: Comparison of protocols and RNA carriers for plasma miRNA isolation. Unraveling RNA carrier influence on miRNA isolation

    doi: 10.1371/journal.pone.0187005

    Figure Lengend Snippet: Correlations between mean “fold recovery” (FR) values of miRNAs and ΔG of each miRNA, using yE protocol (A) and yQ protocol (B). To avoid the influence of GC content in the analysis of ΔG, for these correlations we used only miRNAs with a GC content between 25% and 75% percentile. yE, miRCURY RNA isolation kit Biofluids modified protocol using yeast RNA carrier; yQ, miRNeasy Mini kit modified protocol using yeast RNA carrier; FR, the fold recovery of each miRNA vs the same protocol without carrier; ΔG, the free energy of the most stable secondary structure of each miRNA.

    Article Snippet: The quality and isolation efficiency of miRNAs were analyzed in four plasma samples using two protocols: phenol and column based isolation procedures (miRNeasy Mini kit, Qiagen, Q protocol) and column based procedures (miRCURY RNA Isolation kit Biofluids, Exiqon, E protocol).

    Techniques: Gas Chromatography, Isolation, Modification

    Correlations between mean “fold recovery” (FR) values of miRNAs and GC content of each miRNA, using yE protocol (A) and yQ protocol (B). yE, miRCURY RNA isolation kit Biofluids modified protocol using yeast RNA carrier; yQ, miRNeasy Mini kit modified protocol using yeast RNA carrier; FR, the fold recovery of each miRNA vs the same protocol without carrier.

    Journal: PLoS ONE

    Article Title: Comparison of protocols and RNA carriers for plasma miRNA isolation. Unraveling RNA carrier influence on miRNA isolation

    doi: 10.1371/journal.pone.0187005

    Figure Lengend Snippet: Correlations between mean “fold recovery” (FR) values of miRNAs and GC content of each miRNA, using yE protocol (A) and yQ protocol (B). yE, miRCURY RNA isolation kit Biofluids modified protocol using yeast RNA carrier; yQ, miRNeasy Mini kit modified protocol using yeast RNA carrier; FR, the fold recovery of each miRNA vs the same protocol without carrier.

    Article Snippet: The quality and isolation efficiency of miRNAs were analyzed in four plasma samples using two protocols: phenol and column based isolation procedures (miRNeasy Mini kit, Qiagen, Q protocol) and column based procedures (miRCURY RNA Isolation kit Biofluids, Exiqon, E protocol).

    Techniques: Gas Chromatography, Isolation, Modification

    miRNA levels normalized by plasma volume and by an endogenous miRNA. Fold abundance of miRNAs normalized by plasma volume (FR of miRNAs, left column) and normalized by hsa-miR-103a-3p (apFC of miRNAs, right column) and referred to control plasma samples isolated with the E protocol without carrier (in gray). Data were determined in plasma RNA-derived samples isolated with Q and E modified protocols with yeast RNA as carrier (y) or without RNA carrier (w). E, miRCURY RNA isolation kit Biofluids modified protocol; Q, miRNeasy Mini kit modified protocol; FR, fold recovery of miRNAs vs wE protocol; apFC, apparent fold change of miRNAs vs wE protocol. P -values were calculated using Wilcoxon Signed Rank Test, Exact Signification two-tailed, with the IBM SPSS Statistics 20 software. ϕ P

    Journal: PLoS ONE

    Article Title: Comparison of protocols and RNA carriers for plasma miRNA isolation. Unraveling RNA carrier influence on miRNA isolation

    doi: 10.1371/journal.pone.0187005

    Figure Lengend Snippet: miRNA levels normalized by plasma volume and by an endogenous miRNA. Fold abundance of miRNAs normalized by plasma volume (FR of miRNAs, left column) and normalized by hsa-miR-103a-3p (apFC of miRNAs, right column) and referred to control plasma samples isolated with the E protocol without carrier (in gray). Data were determined in plasma RNA-derived samples isolated with Q and E modified protocols with yeast RNA as carrier (y) or without RNA carrier (w). E, miRCURY RNA isolation kit Biofluids modified protocol; Q, miRNeasy Mini kit modified protocol; FR, fold recovery of miRNAs vs wE protocol; apFC, apparent fold change of miRNAs vs wE protocol. P -values were calculated using Wilcoxon Signed Rank Test, Exact Signification two-tailed, with the IBM SPSS Statistics 20 software. ϕ P

    Article Snippet: The quality and isolation efficiency of miRNAs were analyzed in four plasma samples using two protocols: phenol and column based isolation procedures (miRNeasy Mini kit, Qiagen, Q protocol) and column based procedures (miRCURY RNA Isolation kit Biofluids, Exiqon, E protocol).

    Techniques: Isolation, Derivative Assay, Modification, Two Tailed Test, Software

    Capillary electrophoresis analysis of pig LV biopsy RNA. ( A) Bioanalyzer nano chip traces of all tested samples. Inspection of electropherograms provides additional information to assess RNA quality and protocol performance. (B) Representative traces from each protocol with respective RIN values. Numbers 1–4 correspond to numbered boxes in A. Note that trace 3 has a high RIN despite a large smear in the gel. (C) Summary of RIN values of all the samples tested (mean±SEM). The RNAqueous protocol had undetectable RINs. The mir Vana and miRCURY tissue protocols gave consistently high RIN values, while despite high average RIN, the samples from the miRNeasy micro protocol are more variable. Numbers in brackets indicate the number of samples in each group. (D) Bioanalyzer small RNA chip traces for each kit. Although the RIN of samples marked * were similar (8.6–8.7), the spectrum of small RNAs recovered is highly heterogeneous.

    Journal: PLoS ONE

    Article Title: Optimisation of laboratory methods for whole transcriptomic RNA analyses in human left ventricular biopsies and blood samples of clinical relevance

    doi: 10.1371/journal.pone.0213685

    Figure Lengend Snippet: Capillary electrophoresis analysis of pig LV biopsy RNA. ( A) Bioanalyzer nano chip traces of all tested samples. Inspection of electropherograms provides additional information to assess RNA quality and protocol performance. (B) Representative traces from each protocol with respective RIN values. Numbers 1–4 correspond to numbered boxes in A. Note that trace 3 has a high RIN despite a large smear in the gel. (C) Summary of RIN values of all the samples tested (mean±SEM). The RNAqueous protocol had undetectable RINs. The mir Vana and miRCURY tissue protocols gave consistently high RIN values, while despite high average RIN, the samples from the miRNeasy micro protocol are more variable. Numbers in brackets indicate the number of samples in each group. (D) Bioanalyzer small RNA chip traces for each kit. Although the RIN of samples marked * were similar (8.6–8.7), the spectrum of small RNAs recovered is highly heterogeneous.

    Article Snippet: RNA isolation from pig LV biopsies was performed using four commercially available kits: A) RNAqueous-micro (Ambion, now Invitrogen, AM1931); B) mir Vana (Ambion, now Invitrogen, AM1560); C) miRCURY tissue (Exiqon, now Qiagen, 300115); D) miRNeasy micro (Qiagen 217084).

    Techniques: Electrophoresis, Chromatin Immunoprecipitation

    Evaluating plasma RNA yield using RT-qPCR for miRNAs. As biofluid RNA is below the detection limit of standard quantification tools, all RT-qPCRs were performed using 1.5 μL RNA extracted from 200 μL plasma. Lines indicate paired samples i.e. aliquots of plasma from the same blood draw. (A) RT-qPCR for the spike-in cel-miR-39-3p using the four kits with and without glycogen. (B-D) RT-qPCR for three endogenous miRNAs isolated using the four kits with and without glycogen. (E) The miRNeasy serum/plasma protocol recommends elution in 14 μL, compared with 100 μL for the mir Vana protocol. To control for effects of this, RNA extractions were performed with both protocols, eluting in 30 μL or 50 μL. RT-qPCR for cel-miR-39-3p, hsa-miR-16-5p and hsa-miR-21-5p were performed to evaluate the different elution volumes.

    Journal: PLoS ONE

    Article Title: Optimisation of laboratory methods for whole transcriptomic RNA analyses in human left ventricular biopsies and blood samples of clinical relevance

    doi: 10.1371/journal.pone.0213685

    Figure Lengend Snippet: Evaluating plasma RNA yield using RT-qPCR for miRNAs. As biofluid RNA is below the detection limit of standard quantification tools, all RT-qPCRs were performed using 1.5 μL RNA extracted from 200 μL plasma. Lines indicate paired samples i.e. aliquots of plasma from the same blood draw. (A) RT-qPCR for the spike-in cel-miR-39-3p using the four kits with and without glycogen. (B-D) RT-qPCR for three endogenous miRNAs isolated using the four kits with and without glycogen. (E) The miRNeasy serum/plasma protocol recommends elution in 14 μL, compared with 100 μL for the mir Vana protocol. To control for effects of this, RNA extractions were performed with both protocols, eluting in 30 μL or 50 μL. RT-qPCR for cel-miR-39-3p, hsa-miR-16-5p and hsa-miR-21-5p were performed to evaluate the different elution volumes.

    Article Snippet: RNA isolation from pig LV biopsies was performed using four commercially available kits: A) RNAqueous-micro (Ambion, now Invitrogen, AM1931); B) mir Vana (Ambion, now Invitrogen, AM1560); C) miRCURY tissue (Exiqon, now Qiagen, 300115); D) miRNeasy micro (Qiagen 217084).

    Techniques: Quantitative RT-PCR, Isolation

    Spectrophotometry analysis of pig LV biopsy RNA. RNA extracted according to each manufacturer’s protocol, with the addition of a proteinase K digestion in half the samples. n = 3 biopsies/experimental condition, mean±SEM. N . B . Exiqon miRCURY tissue protocol requires proteinase K, therefore no–proteinase K condition was done with this kit. (A) RNA yield, assessed as ng RNA recovered per mg input tissue, was not significantly different between the protocols (Kruskal-Wallis test with Dunn correction), and additional proteinase K digestion in the RNAqueous micro, mir Vana and miRNeasy micro protocols did not significantly improve RNA yield (Kruskal-Wallis test with Dunn correction). (B) 260/280 and 260/230 ratios provide an assessment of the purity of the RNA. Ratio

    Journal: PLoS ONE

    Article Title: Optimisation of laboratory methods for whole transcriptomic RNA analyses in human left ventricular biopsies and blood samples of clinical relevance

    doi: 10.1371/journal.pone.0213685

    Figure Lengend Snippet: Spectrophotometry analysis of pig LV biopsy RNA. RNA extracted according to each manufacturer’s protocol, with the addition of a proteinase K digestion in half the samples. n = 3 biopsies/experimental condition, mean±SEM. N . B . Exiqon miRCURY tissue protocol requires proteinase K, therefore no–proteinase K condition was done with this kit. (A) RNA yield, assessed as ng RNA recovered per mg input tissue, was not significantly different between the protocols (Kruskal-Wallis test with Dunn correction), and additional proteinase K digestion in the RNAqueous micro, mir Vana and miRNeasy micro protocols did not significantly improve RNA yield (Kruskal-Wallis test with Dunn correction). (B) 260/280 and 260/230 ratios provide an assessment of the purity of the RNA. Ratio

    Article Snippet: RNA isolation from pig LV biopsies was performed using four commercially available kits: A) RNAqueous-micro (Ambion, now Invitrogen, AM1931); B) mir Vana (Ambion, now Invitrogen, AM1560); C) miRCURY tissue (Exiqon, now Qiagen, 300115); D) miRNeasy micro (Qiagen 217084).

    Techniques: Spectrophotometry

    Summary of experimental design for Figs 2 , 3 and 4 . (A) Left ventricular biopsies were taken from three pigs undergoing cardio-pulmonary bypass (CPB). 7 biopsies were taken per pig. RNA was isolated from the biopsies using four commercially available kits; RNAqueous micro, mir Vana, miRCURY tissue, miRNeasy micro. Three kits (RNAqueous micro, mir Vana, miRNeasy micro) were tested with and without an additional proteinase K digestion step, giving 7 protocols in total. The miRCURY tissue protocol includes a proteinase K digestion, therefore this kit was not tested without this. RNA samples were characterised by spectrophotometry and capillary electrophoresis. (B) Blood samples were taken from three pigs undergoing CPB and processed to plasma. RNA was isolated from the plasma using four commercially-available kits; miRNeasy serum/plasma, mir Vana, miRCURY biofluids, Norgen Biotek plasma/serum. Each kit was tested with and without glycogen as a co-precipitant, giving 8 protocols in total. RNA samples were evaluated by RT-qPCR for the exogenous spike-in cel-miR-39-3p, and endogenous miRNAs miR-16-5p, miR-21-5p, miR-92a-3p.

    Journal: PLoS ONE

    Article Title: Optimisation of laboratory methods for whole transcriptomic RNA analyses in human left ventricular biopsies and blood samples of clinical relevance

    doi: 10.1371/journal.pone.0213685

    Figure Lengend Snippet: Summary of experimental design for Figs 2 , 3 and 4 . (A) Left ventricular biopsies were taken from three pigs undergoing cardio-pulmonary bypass (CPB). 7 biopsies were taken per pig. RNA was isolated from the biopsies using four commercially available kits; RNAqueous micro, mir Vana, miRCURY tissue, miRNeasy micro. Three kits (RNAqueous micro, mir Vana, miRNeasy micro) were tested with and without an additional proteinase K digestion step, giving 7 protocols in total. The miRCURY tissue protocol includes a proteinase K digestion, therefore this kit was not tested without this. RNA samples were characterised by spectrophotometry and capillary electrophoresis. (B) Blood samples were taken from three pigs undergoing CPB and processed to plasma. RNA was isolated from the plasma using four commercially-available kits; miRNeasy serum/plasma, mir Vana, miRCURY biofluids, Norgen Biotek plasma/serum. Each kit was tested with and without glycogen as a co-precipitant, giving 8 protocols in total. RNA samples were evaluated by RT-qPCR for the exogenous spike-in cel-miR-39-3p, and endogenous miRNAs miR-16-5p, miR-21-5p, miR-92a-3p.

    Article Snippet: RNA isolation from pig LV biopsies was performed using four commercially available kits: A) RNAqueous-micro (Ambion, now Invitrogen, AM1931); B) mir Vana (Ambion, now Invitrogen, AM1560); C) miRCURY tissue (Exiqon, now Qiagen, 300115); D) miRNeasy micro (Qiagen 217084).

    Techniques: Isolation, Spectrophotometry, Electrophoresis, Quantitative RT-PCR