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Qiagen lysis buffer
Lysis Buffer, supplied by Qiagen, used in various techniques. Bioz Stars score: 95/100, based on 2142 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/lysis buffer/product/Qiagen
Average 95 stars, based on 2142 article reviews
Price from $9.99 to $1999.99
lysis buffer - by Bioz Stars, 2020-09
95/100 stars

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DNA Extraction:

Article Title: Differential Responses of Bovine Monocyte-Derived Macrophages to Infection by Neospora caninum Isolates of High and Low Virulence
Article Snippet: .. Samples were collected by adding 200 μl of PBS, 180 μl of lysis buffer, and 20 μl of proteinase K (Qiagen, Germany) to each well and were stored at −80°C until DNA extraction. ..

Article Title: Rapid antibiotic susceptibility testing from blood culture bottles with species agnostic real-time polymerase chain reaction
Article Snippet: .. Pellets were resuspended in 200 μL lysis buffer (10 mM Tris-HCl, 1 mM disodium EDTA, pH 8.0, 20 mg/mL lysozyme, 300 U/mL mutanolysin, 1.2% Triton X-100) and incubated at 37°C with shaking at 1,200 rpm in a ThermoMixer for 20 min. Glass beads (0.1 mm PowerBeads; Qiagen, Hilden, Germany) were then added to the samples to an approximate volume of 20 μL and the samples were vortexed on the fastest setting for 5 min. Buffers and spin columns from a QIaAMP DNA Mini Kit (Qiagen) were then used to complete the DNA extraction. ..

RNA Extraction:

Article Title: Multi‐omics identify xanthine as a pro‐survival metabolite for nematodes with mitochondrial dysfunction
Article Snippet: .. Pellets were then pestle‐homogenized in the lysis buffer of the RNA Extraction Kit (RNeasy, Qiagen). .. RNA was isolated as per the manufacturer's instructions.

RNA Sequencing Assay:

Article Title: The CREB coactivator CRTC2 promotes oncogenesis in LKB1-mutant non–small cell lung cancer
Article Snippet: .. Real-time qRT-PCR and RNA-seq Cultured cells were disrupted in lysis buffer from the RNeasy Mini Kit (Qiagen), and mRNA was purified following the manufacturer’s instructions. cDNA was synthesized using the Transcriptor First Strand cDNA Synthesis Kit (Roche). .. Quantitative polymerase chain reaction real-time qRT-PCR was carried out with diluted cDNA, appropriate primers, and LightCycler 480 SYBR Green I Master (Roche) on a Lightcycler 480 instrument (Roche).

Synthesized:

Article Title: The CREB coactivator CRTC2 promotes oncogenesis in LKB1-mutant non–small cell lung cancer
Article Snippet: .. Real-time qRT-PCR and RNA-seq Cultured cells were disrupted in lysis buffer from the RNeasy Mini Kit (Qiagen), and mRNA was purified following the manufacturer’s instructions. cDNA was synthesized using the Transcriptor First Strand cDNA Synthesis Kit (Roche). .. Quantitative polymerase chain reaction real-time qRT-PCR was carried out with diluted cDNA, appropriate primers, and LightCycler 480 SYBR Green I Master (Roche) on a Lightcycler 480 instrument (Roche).

Cell Culture:

Article Title: The CREB coactivator CRTC2 promotes oncogenesis in LKB1-mutant non–small cell lung cancer
Article Snippet: .. Real-time qRT-PCR and RNA-seq Cultured cells were disrupted in lysis buffer from the RNeasy Mini Kit (Qiagen), and mRNA was purified following the manufacturer’s instructions. cDNA was synthesized using the Transcriptor First Strand cDNA Synthesis Kit (Roche). .. Quantitative polymerase chain reaction real-time qRT-PCR was carried out with diluted cDNA, appropriate primers, and LightCycler 480 SYBR Green I Master (Roche) on a Lightcycler 480 instrument (Roche).

Purification:

Article Title: The CREB coactivator CRTC2 promotes oncogenesis in LKB1-mutant non–small cell lung cancer
Article Snippet: .. Real-time qRT-PCR and RNA-seq Cultured cells were disrupted in lysis buffer from the RNeasy Mini Kit (Qiagen), and mRNA was purified following the manufacturer’s instructions. cDNA was synthesized using the Transcriptor First Strand cDNA Synthesis Kit (Roche). .. Quantitative polymerase chain reaction real-time qRT-PCR was carried out with diluted cDNA, appropriate primers, and LightCycler 480 SYBR Green I Master (Roche) on a Lightcycler 480 instrument (Roche).

Incubation:

Article Title: Rapid antibiotic susceptibility testing from blood culture bottles with species agnostic real-time polymerase chain reaction
Article Snippet: .. Pellets were resuspended in 200 μL lysis buffer (10 mM Tris-HCl, 1 mM disodium EDTA, pH 8.0, 20 mg/mL lysozyme, 300 U/mL mutanolysin, 1.2% Triton X-100) and incubated at 37°C with shaking at 1,200 rpm in a ThermoMixer for 20 min. Glass beads (0.1 mm PowerBeads; Qiagen, Hilden, Germany) were then added to the samples to an approximate volume of 20 μL and the samples were vortexed on the fastest setting for 5 min. Buffers and spin columns from a QIaAMP DNA Mini Kit (Qiagen) were then used to complete the DNA extraction. ..

Article Title: 1,25-Dihydroxyvitamin D3 affects gastric cancer progression by repressing BMP3 promoter methylation
Article Snippet: .. Then, gastric tissues were dissolved in 200 µL of lysis buffer from the DNA Micro Kit (catalog no. 56304; Qiagen NV, Venlo, the Netherlands) and incubated with proteinase K overnight at 56°C for two nights. .. DNA was extracted according to the manufacturer’s protocol (QIAamp DNA Micro Kit; Qiagen NV) and stored at −80°C to prevent degradation.

Quantitative RT-PCR:

Article Title: The CREB coactivator CRTC2 promotes oncogenesis in LKB1-mutant non–small cell lung cancer
Article Snippet: .. Real-time qRT-PCR and RNA-seq Cultured cells were disrupted in lysis buffer from the RNeasy Mini Kit (Qiagen), and mRNA was purified following the manufacturer’s instructions. cDNA was synthesized using the Transcriptor First Strand cDNA Synthesis Kit (Roche). .. Quantitative polymerase chain reaction real-time qRT-PCR was carried out with diluted cDNA, appropriate primers, and LightCycler 480 SYBR Green I Master (Roche) on a Lightcycler 480 instrument (Roche).

Reverse Transcription Polymerase Chain Reaction:

Article Title: AntagomiR-103 and -107 Treatment Affects Cardiac Function and Metabolism
Article Snippet: .. RNA and RT-PCR Cardiac tissue was lysed with stainless steel beads (QIAGEN, Venlo, the Netherlands) in lysis buffer with the TissueLyser LT (QIAGEN, Venlo, the Netherlands). .. RNA was isolated from the lysate with mirVANA kit (Thermo Fisher Scientific, Waltham, MA), according to the manufacturer’s instructions.

Lysis:

Article Title: Differential Responses of Bovine Monocyte-Derived Macrophages to Infection by Neospora caninum Isolates of High and Low Virulence
Article Snippet: .. Samples were collected by adding 200 μl of PBS, 180 μl of lysis buffer, and 20 μl of proteinase K (Qiagen, Germany) to each well and were stored at −80°C until DNA extraction. ..

Article Title: Nanocomplexes of Graphene Oxide and Platinum Nanoparticles against Colorectal Cancer Colo205, HT-29, HTC-116, SW480, Liver Cancer HepG2, Human Breast Cancer MCF-7, and Adenocarcinoma LNCaP and Human Cervical Hela B Cell Lines
Article Snippet: .. The resulting cell pellet was resuspended in lysis buffer containing 1% 2-mercaptoethanol, and subsequently, the frozen metal balls were added to the probe and homogenized in a TissueLyser ball mill (Qiagen, Germantown, MD, USA) for 5 min at 50 Hz. ..

Article Title: Rapid antibiotic susceptibility testing from blood culture bottles with species agnostic real-time polymerase chain reaction
Article Snippet: .. Pellets were resuspended in 200 μL lysis buffer (10 mM Tris-HCl, 1 mM disodium EDTA, pH 8.0, 20 mg/mL lysozyme, 300 U/mL mutanolysin, 1.2% Triton X-100) and incubated at 37°C with shaking at 1,200 rpm in a ThermoMixer for 20 min. Glass beads (0.1 mm PowerBeads; Qiagen, Hilden, Germany) were then added to the samples to an approximate volume of 20 μL and the samples were vortexed on the fastest setting for 5 min. Buffers and spin columns from a QIaAMP DNA Mini Kit (Qiagen) were then used to complete the DNA extraction. ..

Article Title: 1,25-Dihydroxyvitamin D3 affects gastric cancer progression by repressing BMP3 promoter methylation
Article Snippet: .. Then, gastric tissues were dissolved in 200 µL of lysis buffer from the DNA Micro Kit (catalog no. 56304; Qiagen NV, Venlo, the Netherlands) and incubated with proteinase K overnight at 56°C for two nights. .. DNA was extracted according to the manufacturer’s protocol (QIAamp DNA Micro Kit; Qiagen NV) and stored at −80°C to prevent degradation.

Article Title: The CREB coactivator CRTC2 promotes oncogenesis in LKB1-mutant non–small cell lung cancer
Article Snippet: .. Real-time qRT-PCR and RNA-seq Cultured cells were disrupted in lysis buffer from the RNeasy Mini Kit (Qiagen), and mRNA was purified following the manufacturer’s instructions. cDNA was synthesized using the Transcriptor First Strand cDNA Synthesis Kit (Roche). .. Quantitative polymerase chain reaction real-time qRT-PCR was carried out with diluted cDNA, appropriate primers, and LightCycler 480 SYBR Green I Master (Roche) on a Lightcycler 480 instrument (Roche).

Article Title: Multi‐omics identify xanthine as a pro‐survival metabolite for nematodes with mitochondrial dysfunction
Article Snippet: .. Pellets were then pestle‐homogenized in the lysis buffer of the RNA Extraction Kit (RNeasy, Qiagen). .. RNA was isolated as per the manufacturer's instructions.

Article Title: Identification of biomarkers that distinguish chemical contaminants based on gene expression profiles
Article Snippet: .. Cells were homogenized in the lysis buffer with FAST Prep-24 from MP at speed 6.0/s twice, each for 30s before using RNeasy kits (Qiagen). .. Total RNA concentrations were measured using NanoDrop® ND-1000 Spectrophotometer (NanoDrop technologies, Wilmington, DE, USA).

Article Title: AntagomiR-103 and -107 Treatment Affects Cardiac Function and Metabolism
Article Snippet: .. RNA and RT-PCR Cardiac tissue was lysed with stainless steel beads (QIAGEN, Venlo, the Netherlands) in lysis buffer with the TissueLyser LT (QIAGEN, Venlo, the Netherlands). .. RNA was isolated from the lysate with mirVANA kit (Thermo Fisher Scientific, Waltham, MA), according to the manufacturer’s instructions.

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    Qiagen ii plant dna lysis buffer pa1 c ispc
    Boxplots of the <t>DNA</t> concentrations (ng/μL) of five victorian plant pathology herbarium (VPRI) apple powdery mildew specimens produced by 13 extraction protocols as measured by Invitrogen Qubit ™ fluorometer. Median line __ ; Mean □; Outlier ◆ Extraction method abbreviations: Chelex ® 100 (CheX), innuPrep Plant DNA (InuP), sodium dodecyl sulphate (SDS), E.Z.N.A. ® SP Plant (EznS), DNAzol ™ (DnaZ), E.Z.N.A. ® Forensic DNA (EznF), Qiagen DNeasy ® Plant (DneP), Isolate II Plant DNA Lysis buffer PA1 C (IspC), Isolate II Plant DNA Lysis buffer PA2 S (IspS), Wizard ® Genomic DNA Purification (WizG), E.Z.N.A. ® Plant (EznP), Cetyl trimethyl ammonium bromide (CTAB) and Qiagen DNeasy ® Plant plus PTB (DneP+).
    Ii Plant Dna Lysis Buffer Pa1 C Ispc, supplied by Qiagen, used in various techniques. Bioz Stars score: 84/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/ii plant dna lysis buffer pa1 c ispc/product/Qiagen
    Average 84 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    ii plant dna lysis buffer pa1 c ispc - by Bioz Stars, 2020-09
    84/100 stars
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    99
    Qiagen rlt lysis buffer
    Asymmetric competition is dependent on direct interactions between resident and challenger pneumococci. a, Day 4–5 old pups were inoculated with a heat-killed serotype 23F resident strain (10 5 CFU) for 6h before introduction of the live isogenic challenger strain with increasing CFU (10 1 -10 3 ). Nasal lavages were cultured 3 days later to determine challenger colonization densities, n=3–4. Median values are shown. b, Pups were inoculated with live (10 3 CFU) or heat-killed (10 5 CFU) resident strain for 6h and nasal lavages with <t>RLT</t> lysis buffer were obtained to isolate host <t>RNA.</t> Early inflammatory cytokine expression levels were assessed by SYBR green real-time quantitative RT-PCR. Cytokine expression induced by heat-killed S. pneumoniae were shown as fold change relative to that elicited by live bacteria, n=5–6. Data are shown as mean ± SEM. c-d, Pups were inoculated with either a streptomycin sensitive ( c ) or resistant ( d ) resident strain for 3 days before intranasal administration of streptomycin. An isogenic streptomycin resistant challenger was inoculated 6h later and its colonization density determined after 3 days. Median values are shown. Colonization levels by 10 1 CFU challenger alone, and in resident-colonized pups treated with either phosphate buffered saline (PBS) or streptomycin, were compared by Kruskal–Wallis one-way analysis of variance in ( d ), n=4–8, * p
    Rlt Lysis Buffer, supplied by Qiagen, used in various techniques. Bioz Stars score: 99/100, based on 1054 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rlt lysis buffer/product/Qiagen
    Average 99 stars, based on 1054 article reviews
    Price from $9.99 to $1999.99
    rlt lysis buffer - by Bioz Stars, 2020-09
    99/100 stars
      Buy from Supplier

    99
    Qiagen rna lysis buffer
    Conventional GFP derivatives are not compatible with laser-directed microdissection. The cellular signal of a nuclear-targeted EYFP (EYFPnuc) is preserved in freeze-dried cryosections obtained from transgenic mice. <t>RNA</t> isolated from cryosections is intact. A , The cellular resolution of EYFP expressed in transgenic mice is lost after cryosectioning. Sagittal sections of mouse brains from mice expressing cytoplasmic localized EYFP under the control of the Thy-1 promoter were either fixed with 4% buffered PFA or immediately frozen on dry ice. Vibratome sections (PFA, left) obtained from PFA fixed tissue (30 μm) and cryosections (15 μm; Cryo, middle) from frozen tissue, were analyzed for EYFP fluorescence. After cryosectioning, the cellular resolution of the EYFP fluorescent signal is lost. The EYFP signal is retained but appears to be diffuse and faded. Staining of the cryosection with thionin (Cryo, right) reveals that the cellular integrity of the tissue specimen is intact. B , Nuclear targeted fluorescent proteins appear to be promising candidates for a combined in vivo labeling and microdissection approach. COS7 or HeLa cells were transiently transfected with expression constructs coding for GFP derivatives localized to different cellular compartments: EGFP (cytoplasmic localization), EGFP carrying a farnesylation signal (EGFP-F, attached to membranes), EGFP fused to the N terminus of a synthetic transmembrane protein (TM-EGFP), and EYFP fused to three nuclear localization signals (EYFPnuc). Forty-eight hours after transfection, COS7 cells were either fixed with a PBS-buffered 4% PFA or fixed with 70% ethanol (EtOH) and subsequently dry mounted. HeLa cells transfected with EYFPnuc were either dry mounted (DRY) or fixed with 70% ethanol (EtOH) and subsequently frozen at −80°C before analysis. C , Transgenic mice were generated that express a nuclear-targeted EYFP (EYFPnuc) under control of the Thy-1 Promoter (TYNC mice). The Thy-1 minigene was modified by replacing the Thy-1 ORF with sequences coding for EYFPnuc. D , E , Vibratome sections (30 μm) from brains of PFA-fixed TYNC mice were analyzed for YFP fluorescence and for cellular marker gene expression with immunohistochemistry. D , Within the hippocampus, the YFP expression is most prominent in pyramidal neurons of the CA1 field (CA1) and in the granular cell layer of the dentate gyrus (DG). In the CA3 field (CA3), fewer cells are YFP positive. The YFP fluorescence appears to be strictly overlapping with the indirect GFP immune fluorescence analysis (α-GFP), although it is less sensitive. The pan-neuronal marker NeuN (α-NeuN) colocalizes with the nuclear YFP signal. Staining with GFAP (α-GFAP) and parvalbumin (α-Parv) does not reveal YFP overlapping signals. E , Higher-magnification photographs obtained from vibratome sections at the level of the primary motor and sensory cortex. YFP-positive nuclei (YFP) do not colocalize with the interneuronal marker parvalbumin (α-Parv) and GAD67 (α-GAD67) and also not with the astrocyte marker GFAP (α-GFAP). Arrows mark parvalbumin, GAD67, or GFAP-positive cells stained in red in the left panel. No overlap with YFP fluorescence can be detected in the right panel with arrows pointing to identical positions as in the left panel. Scale bar, 50 μm. F–H , Cryosections (8 μm; Cryo) were mounted on PEN foils (Leica) and analyzed for YFP fluorescence. F , In the hippocampus, YFP-positive nuclei can be detected in the granular cell layer (DG) and in the CA1 field of the pyramidal cell layer (CA1). In the CA3 field (CA3), fewer cells are YFP positive. The inset shows the morphology of the hippocampus in bright field. G , Higher magnification of YFP-positive nuclei in the somatosensory cortex. Scale bar, 50 μm. H , In the cortex, cells located in deeper layers show a nuclear YFP signal. The inset shows the morphology of the section at the level of the somatosensory cortex in bright field. I , Total RNA analyzed with a Bioanalyzer (Agilent); each lane represents half of the RNA isolated from one coronal. brain section. Adjacent 8-μ-thick sections were cryomounted on PEN foil slides, dried, and treated as follows: (1) stored for 2 h at room temperature, (2) stored for 8 h at room temperature, (3) stored for 24 h at room temperature, (4) stored for 48 h at room temperature, (5) frozen at −80°C and then kept at room temperature for 2 h, and (6) <t>microdissected</t> regions pooled from eight sections (total area size isolated comparable with one section). The ratio of the 28S versus the 18S rRNA bands was determined with the Bioanalyzer software and for all samples was 1.3± 0.1. The amount of RNA isolated from one coronal 8 μ brain cryosection was ∼50 ng.
    Rna Lysis Buffer, supplied by Qiagen, used in various techniques. Bioz Stars score: 99/100, based on 214 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rna lysis buffer/product/Qiagen
    Average 99 stars, based on 214 article reviews
    Price from $9.99 to $1999.99
    rna lysis buffer - by Bioz Stars, 2020-09
    99/100 stars
      Buy from Supplier

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    Boxplots of the DNA concentrations (ng/μL) of five victorian plant pathology herbarium (VPRI) apple powdery mildew specimens produced by 13 extraction protocols as measured by Invitrogen Qubit ™ fluorometer. Median line __ ; Mean □; Outlier ◆ Extraction method abbreviations: Chelex ® 100 (CheX), innuPrep Plant DNA (InuP), sodium dodecyl sulphate (SDS), E.Z.N.A. ® SP Plant (EznS), DNAzol ™ (DnaZ), E.Z.N.A. ® Forensic DNA (EznF), Qiagen DNeasy ® Plant (DneP), Isolate II Plant DNA Lysis buffer PA1 C (IspC), Isolate II Plant DNA Lysis buffer PA2 S (IspS), Wizard ® Genomic DNA Purification (WizG), E.Z.N.A. ® Plant (EznP), Cetyl trimethyl ammonium bromide (CTAB) and Qiagen DNeasy ® Plant plus PTB (DneP+).

    Journal: PLoS ONE

    Article Title: Rediscovering an old foe: Optimised molecular methods for DNA extraction and sequencing applications for fungarium specimens of powdery mildew (Erysiphales)

    doi: 10.1371/journal.pone.0232535

    Figure Lengend Snippet: Boxplots of the DNA concentrations (ng/μL) of five victorian plant pathology herbarium (VPRI) apple powdery mildew specimens produced by 13 extraction protocols as measured by Invitrogen Qubit ™ fluorometer. Median line __ ; Mean □; Outlier ◆ Extraction method abbreviations: Chelex ® 100 (CheX), innuPrep Plant DNA (InuP), sodium dodecyl sulphate (SDS), E.Z.N.A. ® SP Plant (EznS), DNAzol ™ (DnaZ), E.Z.N.A. ® Forensic DNA (EznF), Qiagen DNeasy ® Plant (DneP), Isolate II Plant DNA Lysis buffer PA1 C (IspC), Isolate II Plant DNA Lysis buffer PA2 S (IspS), Wizard ® Genomic DNA Purification (WizG), E.Z.N.A. ® Plant (EznP), Cetyl trimethyl ammonium bromide (CTAB) and Qiagen DNeasy ® Plant plus PTB (DneP+).

    Article Snippet: The 13 protocols tested were Chelex® 100 (CheX), innuPrep Plant DNA (InuP), sodium dodecyl sulphate (SDS), E.Z.N.A.® SP Plant (EznS), DNAzol™ (DnaZ), E.Z.N.A.® Forensic DNA (EznF), Qiagen DNeasy® Plant (DneP), Isolate II Plant DNA Lysis buffer PA1 C (IspC), Isolate II Plant DNA Lysis buffer PA2 S (IspS), Wizard® Genomic DNA Purification (WizG), E.Z.N.A.® Plant (EznP), Cetyl trimethyl ammonium bromide (CTAB) and Qiagen DNeasy® Plant plus PTB (DneP+).

    Techniques: Produced, Lysis, DNA Purification

    Boxplots of the DNA quality measured by thermo scientific nanodrop 2000 ™ absorbency measurement 260 nm / 280 nm ratios of five victorian plant pathology herbarium (VPRI) apple powdery mildew specimens produced by 13 DNA extraction methods. The red line indicates the desired target absorbency ratio 1.8–1.9. Median line __ ; Mean □; Outlier ◆ Extraction method abbreviations: Chelex ® 100 (CheX), innuPrep Plant DNA (InuP), sodium dodecyl sulphate (SDS), E.Z.N.A. ® SP Plant (EznS), DNAzol ™ (DnaZ), E.Z.N.A. ® Forensic DNA (EznF), Qiagen DNeasy ® Plant (DneP), Isolate II Plant DNA Lysis buffer PA1 C (IspC), Isolate II Plant DNA Lysis buffer PA2 S (IspS), Wizard ® Genomic DNA Purification (WizG), E.Z.N.A. ® Plant (EznP), Cetyl trimethyl ammonium bromide (CTAB) and Qiagen DNeasy ® Plant plus PTB (DneP+).

    Journal: PLoS ONE

    Article Title: Rediscovering an old foe: Optimised molecular methods for DNA extraction and sequencing applications for fungarium specimens of powdery mildew (Erysiphales)

    doi: 10.1371/journal.pone.0232535

    Figure Lengend Snippet: Boxplots of the DNA quality measured by thermo scientific nanodrop 2000 ™ absorbency measurement 260 nm / 280 nm ratios of five victorian plant pathology herbarium (VPRI) apple powdery mildew specimens produced by 13 DNA extraction methods. The red line indicates the desired target absorbency ratio 1.8–1.9. Median line __ ; Mean □; Outlier ◆ Extraction method abbreviations: Chelex ® 100 (CheX), innuPrep Plant DNA (InuP), sodium dodecyl sulphate (SDS), E.Z.N.A. ® SP Plant (EznS), DNAzol ™ (DnaZ), E.Z.N.A. ® Forensic DNA (EznF), Qiagen DNeasy ® Plant (DneP), Isolate II Plant DNA Lysis buffer PA1 C (IspC), Isolate II Plant DNA Lysis buffer PA2 S (IspS), Wizard ® Genomic DNA Purification (WizG), E.Z.N.A. ® Plant (EznP), Cetyl trimethyl ammonium bromide (CTAB) and Qiagen DNeasy ® Plant plus PTB (DneP+).

    Article Snippet: The 13 protocols tested were Chelex® 100 (CheX), innuPrep Plant DNA (InuP), sodium dodecyl sulphate (SDS), E.Z.N.A.® SP Plant (EznS), DNAzol™ (DnaZ), E.Z.N.A.® Forensic DNA (EznF), Qiagen DNeasy® Plant (DneP), Isolate II Plant DNA Lysis buffer PA1 C (IspC), Isolate II Plant DNA Lysis buffer PA2 S (IspS), Wizard® Genomic DNA Purification (WizG), E.Z.N.A.® Plant (EznP), Cetyl trimethyl ammonium bromide (CTAB) and Qiagen DNeasy® Plant plus PTB (DneP+).

    Techniques: Produced, DNA Extraction, Lysis, DNA Purification

    Box plots of the DNA concentrations (ng/μL) of five victorian plant pathology herbarium (VPRI) apple powdery mildew specimens produced by 13 extraction protocols, as measured by agilent 2200 Tapestation ® DNA. Median line __ ; Mean □; Outlier ◆ Extraction method abbreviations: Chelex ® 100 (CheX), innuPrep Plant DNA (InuP), sodium dodecyl sulphate (SDS), E.Z.N.A. ® SP Plant (EznS), DNAzol ™ (DnaZ), E.Z.N.A. ® Forensic DNA (EznF), Qiagen DNeasy ® Plant (DneP), Isolate II Plant DNA Lysis buffer PA1 C (IspC), Isolate II Plant DNA Lysis buffer PA2 S (IspS), Wizard ® Genomic DNA Purification (WizG), E.Z.N.A. ® Plant (EznP), Cetyl trimethyl ammonium bromide (CTAB) and Qiagen DNeasy ® Plant plus PTB (DneP+).

    Journal: PLoS ONE

    Article Title: Rediscovering an old foe: Optimised molecular methods for DNA extraction and sequencing applications for fungarium specimens of powdery mildew (Erysiphales)

    doi: 10.1371/journal.pone.0232535

    Figure Lengend Snippet: Box plots of the DNA concentrations (ng/μL) of five victorian plant pathology herbarium (VPRI) apple powdery mildew specimens produced by 13 extraction protocols, as measured by agilent 2200 Tapestation ® DNA. Median line __ ; Mean □; Outlier ◆ Extraction method abbreviations: Chelex ® 100 (CheX), innuPrep Plant DNA (InuP), sodium dodecyl sulphate (SDS), E.Z.N.A. ® SP Plant (EznS), DNAzol ™ (DnaZ), E.Z.N.A. ® Forensic DNA (EznF), Qiagen DNeasy ® Plant (DneP), Isolate II Plant DNA Lysis buffer PA1 C (IspC), Isolate II Plant DNA Lysis buffer PA2 S (IspS), Wizard ® Genomic DNA Purification (WizG), E.Z.N.A. ® Plant (EznP), Cetyl trimethyl ammonium bromide (CTAB) and Qiagen DNeasy ® Plant plus PTB (DneP+).

    Article Snippet: The 13 protocols tested were Chelex® 100 (CheX), innuPrep Plant DNA (InuP), sodium dodecyl sulphate (SDS), E.Z.N.A.® SP Plant (EznS), DNAzol™ (DnaZ), E.Z.N.A.® Forensic DNA (EznF), Qiagen DNeasy® Plant (DneP), Isolate II Plant DNA Lysis buffer PA1 C (IspC), Isolate II Plant DNA Lysis buffer PA2 S (IspS), Wizard® Genomic DNA Purification (WizG), E.Z.N.A.® Plant (EznP), Cetyl trimethyl ammonium bromide (CTAB) and Qiagen DNeasy® Plant plus PTB (DneP+).

    Techniques: Produced, Lysis, DNA Purification

    Asymmetric competition is dependent on direct interactions between resident and challenger pneumococci. a, Day 4–5 old pups were inoculated with a heat-killed serotype 23F resident strain (10 5 CFU) for 6h before introduction of the live isogenic challenger strain with increasing CFU (10 1 -10 3 ). Nasal lavages were cultured 3 days later to determine challenger colonization densities, n=3–4. Median values are shown. b, Pups were inoculated with live (10 3 CFU) or heat-killed (10 5 CFU) resident strain for 6h and nasal lavages with RLT lysis buffer were obtained to isolate host RNA. Early inflammatory cytokine expression levels were assessed by SYBR green real-time quantitative RT-PCR. Cytokine expression induced by heat-killed S. pneumoniae were shown as fold change relative to that elicited by live bacteria, n=5–6. Data are shown as mean ± SEM. c-d, Pups were inoculated with either a streptomycin sensitive ( c ) or resistant ( d ) resident strain for 3 days before intranasal administration of streptomycin. An isogenic streptomycin resistant challenger was inoculated 6h later and its colonization density determined after 3 days. Median values are shown. Colonization levels by 10 1 CFU challenger alone, and in resident-colonized pups treated with either phosphate buffered saline (PBS) or streptomycin, were compared by Kruskal–Wallis one-way analysis of variance in ( d ), n=4–8, * p

    Journal: Nature microbiology

    Article Title: Pneumococcal quorum sensing drives an asymmetric owner-intruder competitive strategy during carriage via the competence regulon

    doi: 10.1038/s41564-018-0314-4

    Figure Lengend Snippet: Asymmetric competition is dependent on direct interactions between resident and challenger pneumococci. a, Day 4–5 old pups were inoculated with a heat-killed serotype 23F resident strain (10 5 CFU) for 6h before introduction of the live isogenic challenger strain with increasing CFU (10 1 -10 3 ). Nasal lavages were cultured 3 days later to determine challenger colonization densities, n=3–4. Median values are shown. b, Pups were inoculated with live (10 3 CFU) or heat-killed (10 5 CFU) resident strain for 6h and nasal lavages with RLT lysis buffer were obtained to isolate host RNA. Early inflammatory cytokine expression levels were assessed by SYBR green real-time quantitative RT-PCR. Cytokine expression induced by heat-killed S. pneumoniae were shown as fold change relative to that elicited by live bacteria, n=5–6. Data are shown as mean ± SEM. c-d, Pups were inoculated with either a streptomycin sensitive ( c ) or resistant ( d ) resident strain for 3 days before intranasal administration of streptomycin. An isogenic streptomycin resistant challenger was inoculated 6h later and its colonization density determined after 3 days. Median values are shown. Colonization levels by 10 1 CFU challenger alone, and in resident-colonized pups treated with either phosphate buffered saline (PBS) or streptomycin, were compared by Kruskal–Wallis one-way analysis of variance in ( d ), n=4–8, * p

    Article Snippet: To obtain host RNA, nasal lavages with RLT lysis buffer (Qiagen) were performed and RNA isolated with the RNeasy minikit (Qiagen) as previously described . cDNA was reverse transcribed using a high-capacity cDNA kit (Applied Biosystems) following the manufacturer’s instructions.

    Techniques: Cell Culture, Lysis, Expressing, SYBR Green Assay, Quantitative RT-PCR

    Interleukin 17 (IL-17) blockade and Streptococcus pneumoniae pathogenesis in an S. pneumoniae– influenza coinfection model. A, C57BL/6 mice 6–8 weeks old (wild-type [WT] and RoRγt −/− ) were intranasally inoculated (10 µL) with ST 6A (1 × 10 6 colony-forming units [CFUs]). Twenty-four hours later, they were inoculated intranasally (10 µL) with influenza A H1N1 A/Puerto Rico/8/1934 (PR8) virus (100× median tissue culture infective dose). Interleukin 17A (IL-17A) was neutralized in WT mice (WT IL-17A − ) by injecting 200 µg of anti-IL-17A antibody. Control coinfected WT mice received an equal amount of isotype control antibody. Five days after coinfection, the mice were euthanized, and the first retrograde nasopharyngeal (NP) lavage fluid sample was collected (with phosphate-buffered saline). Cytokine levels in NP lavage fluid from coinfected mice (WT, WT IL-17A − , RoRγt −/− ) were determined by means of cytometric multiplex bead array. Data represent pooled NP lavage fluid from 2 independent experiments (5 per group). B, Total protein in pooled NP lavage fluid from WT, WT IL-17A − and RoRγt −/− coinfected mice. C, Neutrophil levels in NP lavage fluid from coinfected mice (WT, WT IL-17A − , and RoRγt −/− ). Data are expressed as representative contour plots and bar graphs with the percentage of neutrophils. Bar graphs represent data from pooled NP lavage fluid of 2 independent experiments (4–7 mice per group). D, The second retrograde NP lavage fluid sample from coinfected mice was collected in cell lysing RLT buffer (Qiagen). The messenger RNA (mRNA) levels of epithelial tight junction proteins ZO-1 and occludin were determined with quantitative reverse-transcription polymerase chain reaction. Data were analyzed using 1-way analysis of variance, followed by pair-based comparisons with the Tukey test. * P

    Journal: The Journal of Infectious Diseases

    Article Title: Double-Edged Role of Interleukin 17A in Streptococcus pneumoniae Pathogenesis During Influenza Virus Coinfection

    doi: 10.1093/infdis/jiz193

    Figure Lengend Snippet: Interleukin 17 (IL-17) blockade and Streptococcus pneumoniae pathogenesis in an S. pneumoniae– influenza coinfection model. A, C57BL/6 mice 6–8 weeks old (wild-type [WT] and RoRγt −/− ) were intranasally inoculated (10 µL) with ST 6A (1 × 10 6 colony-forming units [CFUs]). Twenty-four hours later, they were inoculated intranasally (10 µL) with influenza A H1N1 A/Puerto Rico/8/1934 (PR8) virus (100× median tissue culture infective dose). Interleukin 17A (IL-17A) was neutralized in WT mice (WT IL-17A − ) by injecting 200 µg of anti-IL-17A antibody. Control coinfected WT mice received an equal amount of isotype control antibody. Five days after coinfection, the mice were euthanized, and the first retrograde nasopharyngeal (NP) lavage fluid sample was collected (with phosphate-buffered saline). Cytokine levels in NP lavage fluid from coinfected mice (WT, WT IL-17A − , RoRγt −/− ) were determined by means of cytometric multiplex bead array. Data represent pooled NP lavage fluid from 2 independent experiments (5 per group). B, Total protein in pooled NP lavage fluid from WT, WT IL-17A − and RoRγt −/− coinfected mice. C, Neutrophil levels in NP lavage fluid from coinfected mice (WT, WT IL-17A − , and RoRγt −/− ). Data are expressed as representative contour plots and bar graphs with the percentage of neutrophils. Bar graphs represent data from pooled NP lavage fluid of 2 independent experiments (4–7 mice per group). D, The second retrograde NP lavage fluid sample from coinfected mice was collected in cell lysing RLT buffer (Qiagen). The messenger RNA (mRNA) levels of epithelial tight junction proteins ZO-1 and occludin were determined with quantitative reverse-transcription polymerase chain reaction. Data were analyzed using 1-way analysis of variance, followed by pair-based comparisons with the Tukey test. * P

    Article Snippet: Quantitative Reverse-Transcription Polymerase Chain Reaction Total RNA was extracted from NP lavage fluid collected with RLT lysis buffer (Qiagen) containing 0.001% 2-mercaptoethanol (Qiagen).

    Techniques: Mouse Assay, Multiplex Assay, Reverse Transcription Polymerase Chain Reaction

    Conventional GFP derivatives are not compatible with laser-directed microdissection. The cellular signal of a nuclear-targeted EYFP (EYFPnuc) is preserved in freeze-dried cryosections obtained from transgenic mice. RNA isolated from cryosections is intact. A , The cellular resolution of EYFP expressed in transgenic mice is lost after cryosectioning. Sagittal sections of mouse brains from mice expressing cytoplasmic localized EYFP under the control of the Thy-1 promoter were either fixed with 4% buffered PFA or immediately frozen on dry ice. Vibratome sections (PFA, left) obtained from PFA fixed tissue (30 μm) and cryosections (15 μm; Cryo, middle) from frozen tissue, were analyzed for EYFP fluorescence. After cryosectioning, the cellular resolution of the EYFP fluorescent signal is lost. The EYFP signal is retained but appears to be diffuse and faded. Staining of the cryosection with thionin (Cryo, right) reveals that the cellular integrity of the tissue specimen is intact. B , Nuclear targeted fluorescent proteins appear to be promising candidates for a combined in vivo labeling and microdissection approach. COS7 or HeLa cells were transiently transfected with expression constructs coding for GFP derivatives localized to different cellular compartments: EGFP (cytoplasmic localization), EGFP carrying a farnesylation signal (EGFP-F, attached to membranes), EGFP fused to the N terminus of a synthetic transmembrane protein (TM-EGFP), and EYFP fused to three nuclear localization signals (EYFPnuc). Forty-eight hours after transfection, COS7 cells were either fixed with a PBS-buffered 4% PFA or fixed with 70% ethanol (EtOH) and subsequently dry mounted. HeLa cells transfected with EYFPnuc were either dry mounted (DRY) or fixed with 70% ethanol (EtOH) and subsequently frozen at −80°C before analysis. C , Transgenic mice were generated that express a nuclear-targeted EYFP (EYFPnuc) under control of the Thy-1 Promoter (TYNC mice). The Thy-1 minigene was modified by replacing the Thy-1 ORF with sequences coding for EYFPnuc. D , E , Vibratome sections (30 μm) from brains of PFA-fixed TYNC mice were analyzed for YFP fluorescence and for cellular marker gene expression with immunohistochemistry. D , Within the hippocampus, the YFP expression is most prominent in pyramidal neurons of the CA1 field (CA1) and in the granular cell layer of the dentate gyrus (DG). In the CA3 field (CA3), fewer cells are YFP positive. The YFP fluorescence appears to be strictly overlapping with the indirect GFP immune fluorescence analysis (α-GFP), although it is less sensitive. The pan-neuronal marker NeuN (α-NeuN) colocalizes with the nuclear YFP signal. Staining with GFAP (α-GFAP) and parvalbumin (α-Parv) does not reveal YFP overlapping signals. E , Higher-magnification photographs obtained from vibratome sections at the level of the primary motor and sensory cortex. YFP-positive nuclei (YFP) do not colocalize with the interneuronal marker parvalbumin (α-Parv) and GAD67 (α-GAD67) and also not with the astrocyte marker GFAP (α-GFAP). Arrows mark parvalbumin, GAD67, or GFAP-positive cells stained in red in the left panel. No overlap with YFP fluorescence can be detected in the right panel with arrows pointing to identical positions as in the left panel. Scale bar, 50 μm. F–H , Cryosections (8 μm; Cryo) were mounted on PEN foils (Leica) and analyzed for YFP fluorescence. F , In the hippocampus, YFP-positive nuclei can be detected in the granular cell layer (DG) and in the CA1 field of the pyramidal cell layer (CA1). In the CA3 field (CA3), fewer cells are YFP positive. The inset shows the morphology of the hippocampus in bright field. G , Higher magnification of YFP-positive nuclei in the somatosensory cortex. Scale bar, 50 μm. H , In the cortex, cells located in deeper layers show a nuclear YFP signal. The inset shows the morphology of the section at the level of the somatosensory cortex in bright field. I , Total RNA analyzed with a Bioanalyzer (Agilent); each lane represents half of the RNA isolated from one coronal. brain section. Adjacent 8-μ-thick sections were cryomounted on PEN foil slides, dried, and treated as follows: (1) stored for 2 h at room temperature, (2) stored for 8 h at room temperature, (3) stored for 24 h at room temperature, (4) stored for 48 h at room temperature, (5) frozen at −80°C and then kept at room temperature for 2 h, and (6) microdissected regions pooled from eight sections (total area size isolated comparable with one section). The ratio of the 28S versus the 18S rRNA bands was determined with the Bioanalyzer software and for all samples was 1.3± 0.1. The amount of RNA isolated from one coronal 8 μ brain cryosection was ∼50 ng.

    Journal: The Journal of Neuroscience

    Article Title: Global Transcriptome Analysis of Genetically Identified Neurons in the Adult Cortex

    doi: 10.1523/JNEUROSCI.0468-06.2006

    Figure Lengend Snippet: Conventional GFP derivatives are not compatible with laser-directed microdissection. The cellular signal of a nuclear-targeted EYFP (EYFPnuc) is preserved in freeze-dried cryosections obtained from transgenic mice. RNA isolated from cryosections is intact. A , The cellular resolution of EYFP expressed in transgenic mice is lost after cryosectioning. Sagittal sections of mouse brains from mice expressing cytoplasmic localized EYFP under the control of the Thy-1 promoter were either fixed with 4% buffered PFA or immediately frozen on dry ice. Vibratome sections (PFA, left) obtained from PFA fixed tissue (30 μm) and cryosections (15 μm; Cryo, middle) from frozen tissue, were analyzed for EYFP fluorescence. After cryosectioning, the cellular resolution of the EYFP fluorescent signal is lost. The EYFP signal is retained but appears to be diffuse and faded. Staining of the cryosection with thionin (Cryo, right) reveals that the cellular integrity of the tissue specimen is intact. B , Nuclear targeted fluorescent proteins appear to be promising candidates for a combined in vivo labeling and microdissection approach. COS7 or HeLa cells were transiently transfected with expression constructs coding for GFP derivatives localized to different cellular compartments: EGFP (cytoplasmic localization), EGFP carrying a farnesylation signal (EGFP-F, attached to membranes), EGFP fused to the N terminus of a synthetic transmembrane protein (TM-EGFP), and EYFP fused to three nuclear localization signals (EYFPnuc). Forty-eight hours after transfection, COS7 cells were either fixed with a PBS-buffered 4% PFA or fixed with 70% ethanol (EtOH) and subsequently dry mounted. HeLa cells transfected with EYFPnuc were either dry mounted (DRY) or fixed with 70% ethanol (EtOH) and subsequently frozen at −80°C before analysis. C , Transgenic mice were generated that express a nuclear-targeted EYFP (EYFPnuc) under control of the Thy-1 Promoter (TYNC mice). The Thy-1 minigene was modified by replacing the Thy-1 ORF with sequences coding for EYFPnuc. D , E , Vibratome sections (30 μm) from brains of PFA-fixed TYNC mice were analyzed for YFP fluorescence and for cellular marker gene expression with immunohistochemistry. D , Within the hippocampus, the YFP expression is most prominent in pyramidal neurons of the CA1 field (CA1) and in the granular cell layer of the dentate gyrus (DG). In the CA3 field (CA3), fewer cells are YFP positive. The YFP fluorescence appears to be strictly overlapping with the indirect GFP immune fluorescence analysis (α-GFP), although it is less sensitive. The pan-neuronal marker NeuN (α-NeuN) colocalizes with the nuclear YFP signal. Staining with GFAP (α-GFAP) and parvalbumin (α-Parv) does not reveal YFP overlapping signals. E , Higher-magnification photographs obtained from vibratome sections at the level of the primary motor and sensory cortex. YFP-positive nuclei (YFP) do not colocalize with the interneuronal marker parvalbumin (α-Parv) and GAD67 (α-GAD67) and also not with the astrocyte marker GFAP (α-GFAP). Arrows mark parvalbumin, GAD67, or GFAP-positive cells stained in red in the left panel. No overlap with YFP fluorescence can be detected in the right panel with arrows pointing to identical positions as in the left panel. Scale bar, 50 μm. F–H , Cryosections (8 μm; Cryo) were mounted on PEN foils (Leica) and analyzed for YFP fluorescence. F , In the hippocampus, YFP-positive nuclei can be detected in the granular cell layer (DG) and in the CA1 field of the pyramidal cell layer (CA1). In the CA3 field (CA3), fewer cells are YFP positive. The inset shows the morphology of the hippocampus in bright field. G , Higher magnification of YFP-positive nuclei in the somatosensory cortex. Scale bar, 50 μm. H , In the cortex, cells located in deeper layers show a nuclear YFP signal. The inset shows the morphology of the section at the level of the somatosensory cortex in bright field. I , Total RNA analyzed with a Bioanalyzer (Agilent); each lane represents half of the RNA isolated from one coronal. brain section. Adjacent 8-μ-thick sections were cryomounted on PEN foil slides, dried, and treated as follows: (1) stored for 2 h at room temperature, (2) stored for 8 h at room temperature, (3) stored for 24 h at room temperature, (4) stored for 48 h at room temperature, (5) frozen at −80°C and then kept at room temperature for 2 h, and (6) microdissected regions pooled from eight sections (total area size isolated comparable with one section). The ratio of the 28S versus the 18S rRNA bands was determined with the Bioanalyzer software and for all samples was 1.3± 0.1. The amount of RNA isolated from one coronal 8 μ brain cryosection was ∼50 ng.

    Article Snippet: Microdissected samples were collected in 100 μl of RNA lysis buffer (Qiagen, Hilden, Germany) and stored at room temperature until further processing.

    Techniques: Laser Capture Microdissection, Transgenic Assay, Mouse Assay, Isolation, Expressing, Fluorescence, Staining, In Vivo, Labeling, Transfection, Construct, Generated, Modification, Marker, Immunohistochemistry, Software