λ exonuclease reaction buffer  (New England Biolabs)


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    Name:
    Exonuclease I Reaction Buffer
    Description:
    Exonuclease I Reaction Buffer 6 0 ml
    Catalog Number:
    b0293s
    Price:
    24
    Size:
    6 0 ml
    Category:
    Buffers
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    Structured Review

    New England Biolabs λ exonuclease reaction buffer
    Exonuclease I Reaction Buffer
    Exonuclease I Reaction Buffer 6 0 ml
    https://www.bioz.com/result/λ exonuclease reaction buffer/product/New England Biolabs
    Average 95 stars, based on 38 article reviews
    Price from $9.99 to $1999.99
    λ exonuclease reaction buffer - by Bioz Stars, 2020-02
    95/100 stars

    Images

    1) Product Images from "Simplified ChIP-exo assays"

    Article Title: Simplified ChIP-exo assays

    Journal: Nature Communications

    doi: 10.1038/s41467-018-05265-7

    ChIP-exo 5.0 increases library yield. a Schematic of ChIP-exo 5.0. The purple triangle indicates the location of the Read_1 start site, which is also the λ exonuclease stop site. b 2% agarose gel of the electrophoresed library following 18 cycles of PCR for various S. cerevisiae transcription factors assayed by ChIP-exo 1.1 or 5.0. Following ChIP, the sample was split and libraries prepared using the indicated protocols. After splitting the sample, each reaction contained a 50 ml cell equivalent (OD 600 = 0.8) of yeast chromatin, which is five-fold less than the amount optimized for ChIP-exo 1.1. ChIP-exo 5.0 produced greater library yield for all samples. c Heatmaps comparing ChIP-exo 1.1 and 5.0 at the 975 Reb1 primary motifs in a 200 bp window. d Composite plot of data from panel ( c )
    Figure Legend Snippet: ChIP-exo 5.0 increases library yield. a Schematic of ChIP-exo 5.0. The purple triangle indicates the location of the Read_1 start site, which is also the λ exonuclease stop site. b 2% agarose gel of the electrophoresed library following 18 cycles of PCR for various S. cerevisiae transcription factors assayed by ChIP-exo 1.1 or 5.0. Following ChIP, the sample was split and libraries prepared using the indicated protocols. After splitting the sample, each reaction contained a 50 ml cell equivalent (OD 600 = 0.8) of yeast chromatin, which is five-fold less than the amount optimized for ChIP-exo 1.1. ChIP-exo 5.0 produced greater library yield for all samples. c Heatmaps comparing ChIP-exo 1.1 and 5.0 at the 975 Reb1 primary motifs in a 200 bp window. d Composite plot of data from panel ( c )

    Techniques Used: Chromatin Immunoprecipitation, Agarose Gel Electrophoresis, Polymerase Chain Reaction, Produced

    2) Product Images from "Simplified ChIP-exo assays"

    Article Title: Simplified ChIP-exo assays

    Journal: Nature Communications

    doi: 10.1038/s41467-018-05265-7

    ChIP-exo 5.0 increases library yield. a Schematic of ChIP-exo 5.0. The purple triangle indicates the location of the Read_1 start site, which is also the λ exonuclease stop site. b 2% agarose gel of the electrophoresed library following 18 cycles of PCR for various S. cerevisiae transcription factors assayed by ChIP-exo 1.1 or 5.0. Following ChIP, the sample was split and libraries prepared using the indicated protocols. After splitting the sample, each reaction contained a 50 ml cell equivalent (OD 600 = 0.8) of yeast chromatin, which is five-fold less than the amount optimized for ChIP-exo 1.1. ChIP-exo 5.0 produced greater library yield for all samples. c Heatmaps comparing ChIP-exo 1.1 and 5.0 at the 975 Reb1 primary motifs in a 200 bp window. d Composite plot of data from panel ( c )
    Figure Legend Snippet: ChIP-exo 5.0 increases library yield. a Schematic of ChIP-exo 5.0. The purple triangle indicates the location of the Read_1 start site, which is also the λ exonuclease stop site. b 2% agarose gel of the electrophoresed library following 18 cycles of PCR for various S. cerevisiae transcription factors assayed by ChIP-exo 1.1 or 5.0. Following ChIP, the sample was split and libraries prepared using the indicated protocols. After splitting the sample, each reaction contained a 50 ml cell equivalent (OD 600 = 0.8) of yeast chromatin, which is five-fold less than the amount optimized for ChIP-exo 1.1. ChIP-exo 5.0 produced greater library yield for all samples. c Heatmaps comparing ChIP-exo 1.1 and 5.0 at the 975 Reb1 primary motifs in a 200 bp window. d Composite plot of data from panel ( c )

    Techniques Used: Chromatin Immunoprecipitation, Agarose Gel Electrophoresis, Polymerase Chain Reaction, Produced

    Related Articles

    Methylation Sequencing:

    Article Title: Novel DNA methylation biomarkers show high sensitivity and specificity for blood-based detection of colorectal cancer—a clinical biomarker discovery and validation study
    Article Snippet: Paragraph title: Bisulfite sequencing ... To remove excess primers and dNTPs, 2 μl PCR products were treated with 1 μl FastAP Thermosensitive Alkaline Phosphatase (1 μmol/μl) (Life Technologies) and 1 μl EXO1 (20,000 U/ml) (New England Biolabs) diluted 1:4 in 10x Exonuclease I Reaction buffer (New England Biolabs).

    Clone Assay:

    Article Title: Light-activated cell identification and sorting (LACIS) for selection of edited clones on a nanofluidic device
    Article Snippet: Sample processing for next-generation sequencing Genomic DNA was extracted from exported clones by incubating in Proteinase K buffer (0.1 M NaCl, 10 mM Tris-HCl pH 8.0, 1 mM EDTA) for 30 min at 55 °C, then for 20 min at 80 °C to inactivate Proteinase K. The genomic region around the CRISPR/Cas9 target site for CXCR4 gene was amplified by PCR with primers positioned outside of the HDR repair template sequence (positioned to avoid amplification of exogenous template) for ten cycles using KAPA HiFi Hotstart ReadyMix (Kapa Biosystems, KR0370) according to the manufacturer’s protocol (PCR primers listed in Supplementary Data ). .. Excess PCR primers were removed by incubating with Exonuclease I (NEB, M0293S) in 1 × exonuclease reaction buffer (NEB, B0293S) for 1 h at 37 °C, followed by enzyme inactivation for 20 min at 80 °C.

    Amplification:

    Article Title: Spatial distribution and function of T follicular regulatory cells in human lymph nodes
    Article Snippet: Then, each sample was reverse-transcribed using Reverse Transcription Mastermix (Fluidigm). cDNA was amplified with 15 cycles of specific target preamplification using the Fluidigm Pre-Amp Master Mix and the full complement of primers. .. Preamplified samples were subjected to an exonuclease reaction, using Exonuclease I and Exonuclease I Reaction Buffer (New England Biolabs).

    Article Title: Light-activated cell identification and sorting (LACIS) for selection of edited clones on a nanofluidic device
    Article Snippet: Sample processing for next-generation sequencing Genomic DNA was extracted from exported clones by incubating in Proteinase K buffer (0.1 M NaCl, 10 mM Tris-HCl pH 8.0, 1 mM EDTA) for 30 min at 55 °C, then for 20 min at 80 °C to inactivate Proteinase K. The genomic region around the CRISPR/Cas9 target site for CXCR4 gene was amplified by PCR with primers positioned outside of the HDR repair template sequence (positioned to avoid amplification of exogenous template) for ten cycles using KAPA HiFi Hotstart ReadyMix (Kapa Biosystems, KR0370) according to the manufacturer’s protocol (PCR primers listed in Supplementary Data ). .. Excess PCR primers were removed by incubating with Exonuclease I (NEB, M0293S) in 1 × exonuclease reaction buffer (NEB, B0293S) for 1 h at 37 °C, followed by enzyme inactivation for 20 min at 80 °C.

    Article Title: Novel DNA methylation biomarkers show high sensitivity and specificity for blood-based detection of colorectal cancer—a clinical biomarker discovery and validation study
    Article Snippet: Following PCR amplification, 5 μl of each amplicon were visualized by agarose gel electrophoresis in Tris-acetate-EDTA (TAE) buffer (Fagron) with ethidium bromide (EtBr) to test for correct band size. .. To remove excess primers and dNTPs, 2 μl PCR products were treated with 1 μl FastAP Thermosensitive Alkaline Phosphatase (1 μmol/μl) (Life Technologies) and 1 μl EXO1 (20,000 U/ml) (New England Biolabs) diluted 1:4 in 10x Exonuclease I Reaction buffer (New England Biolabs).

    Article Title: Infantile Hemangioma Originates From A Dysregulated But Not Fully Transformed Multipotent Stem Cell
    Article Snippet: The 2-step process included STA (specific target amplification) reaction (RT-STA cycling conditions and an Exonuclease I treatment method to remove unincorporated primers). .. The Exo I treatment method (total volume 3.5 μl [per 9 μl RT-STA solution]) consisted of: 2.52 μl Nuclease Free H2O, 0.36 μl Exonuclease I Reaction Buffer (10X), and 0.72 μl Exonuclease I (20units/μl) (New England BioLabs).

    Article Title: Functional Gut Microbiota Remodeling Contributes to the Caloric Restriction-Induced Metabolic Improvements
    Article Snippet: The V3-4 region of the bacterial 16S rRNA genes (E. coli positions 341-805) was amplified using 1 μL of DNA extract adjusted to 1 ng/μL for samples (undiluted for negative extraction controls) in a 25 μL volume that contained 12.5 μL KAPA2G Robust HotStart ReadyMix (Kapa Biosystems, Boston, MA, USA) and 0.5 μL each of 10 μM forward primer 341F 5′-CCTACGGGNGGCWGCAG-3′ and reverse primer 805R 5′-GACTACHVGGGTATCTAAKCC-3′ ( ). .. The primers from the first round of PCR were removed by digesting 5 μL samples (containing 9.5-13.9 ng/μL DNA except in negative extraction controls for which DNA concentration was under the detection level) with 1 unit Exonuclease I (New England Biolabs, Ipswich, MA, USA) in a total volume of 10 μL Exonuclease I Reaction Buffer (New England Biolabs) at 37°C for 30 min.

    Article Title: Decontamination of 16S rRNA gene amplicon sequence datasets based on bacterial load assessment by qPCR
    Article Snippet: PCR and sequencing The V3–4 region of the bacterial 16S rRNA genes (E. coli positions 341–805) was amplified using template DNA from E. coli and S. aureus cultures, and from NECs. .. The primers from the first round of PCR were removed by digesting 5-μl samples with 1 unit Exonuclease I (New England Biolabs, Ipswich, MA, USA) in a total volume of 10 μl Exonuclease I Reaction Buffer (New England Biolabs) at 37 °C for 30 min.

    Article Title: Direct elicitation of template concentration from quantification cycle (Cq) distributions in digital PCR
    Article Snippet: Pre-amplification was performed on an iQ5 thermocycler (BioRad) with the following program: denaturation at 95°C for 10 min and 18 cycles of amplification (15 s at 95°C, 4 min at 60°C). .. Unincorporated primers were digested by adding a 6 μl solution containing 4 units Exonuclease I (New England BioLabs M0293L) in 1× Exonuclease I Reaction Buffer (New England BioLabs B0293S) and using the following thermal protocol: 30 min at 37°C, 15 min at 80°C, hold at 4°C.

    Article Title: Single-cell full-length total RNA sequencing uncovers dynamics of recursive splicing and enhancer RNAs
    Article Snippet: Next, 0.5 μL of genomic DNA digestion mix (0.1 U of DNase I Amplification Grade (Thermo Fisher) and 2× DNase I Reaction Buffer (Thermo Fisher) in RNase-free water) was added to 1 μL of the single-cell lysate in a 96-well PCR plate and incubated at 25 °C for 5 min. After genomic DNA digestion, we added 0.5 µL of denaturing mix (8 mM EDTA and 0.02% NP40 in RNase-free water) to the digested sample, followed by incubation at 70 °C for 5 min to inactivate DNase I and desaturate the RNAs. .. The PCR conditions were as follows: activation at 95 °C for 2 min; 14 cycles of denaturation at 95 °C for 30 s; annealing at 60 °C for 90 s; and extension at 72 °C for 60 s. After PCR preamplification, the PCR products were added 8 µL of primer digestion mix (32 U of Exonuclease I (NEB) and 1× Exonuclease I Reaction Buffer (NEB) in nuclease-free water) and incubated at 37 °C for 30 min and 80 °C for 15 min.

    Article Title: Endovascular biopsy: Strategy for analyzing gene expression profiles of individual endothelial cells obtained from human vessels✩
    Article Snippet: The samples were incubated at 50 °C for 15 min for the reverse transcription, 95 °C for 2 min for inactivating reverse transcriptase and activating Taq polymerase, then subjected to 18 PCR cycles (95 °C 15 sec then 60 °C for 4 min for each cycle) for specific targets amplification (STA). .. To remove the unincorporated primers for best results, each sample was then mixed with 3.6 μL exonuclease treatment buffer composed of 2.52 μL water, 0.36 μL 10× Exonuclease I reaction buffer and 0.72 μL 20 units/μL Exonuclease I (New England BioLabs, Ipswich, MA), incubated at 37 °C for 30 min for digestion and 80 °C for 15 min to inactivate the exonuclease.

    Article Title: Single-cell full-length total RNA sequencing uncovers dynamics of recursive splicing and enhancer RNAs
    Article Snippet: Next, 0.5 μL of genomic DNA digestion mix (0.1 U of DNase I Amplification Grade (Thermo Fisher) and 2× DNase I Reaction Buffer (Thermo Fisher) in RNase-free water) was added to 1 μL of the single-cell lysate in a 96-well PCR plate and incubated at 25 °C for 5 min. After genomic DNA digestion, we added 0.5 µL of denaturing mix (8 mM EDTA and 0.02% NP40 in RNase-free water) to the digested sample, followed by incubation at 70 °C for 5 min to inactivate DNase I and desaturate the RNAs. .. The PCR conditions were as follows: activation at 95 °C for 2 mi n; 14 cycles of denaturation at 95 °C for 30 s; annealing at 60 °C for 90 s; and extension at 72 °C for 60 s. After PCR preamplification, the PCR products were added 8 µL of primer digestion mix (32 U of Exonuclease I (NEB) and 1× Exonuclease I Reaction Buffer (NEB) in nuclease-free water) and incubated at 37 °C for 30 min and 80 °C for 15 min.

    Quantitative RT-PCR:

    Article Title: Spatial distribution and function of T follicular regulatory cells in human lymph nodes
    Article Snippet: Paragraph title: Real-time quantitative RT-PCR ... Preamplified samples were subjected to an exonuclease reaction, using Exonuclease I and Exonuclease I Reaction Buffer (New England Biolabs).

    Article Title: Single-cell full-length total RNA sequencing uncovers dynamics of recursive splicing and enhancer RNAs
    Article Snippet: Paragraph title: Single-cell RT-qPCR ... The PCR conditions were as follows: activation at 95 °C for 2 min; 14 cycles of denaturation at 95 °C for 30 s; annealing at 60 °C for 90 s; and extension at 72 °C for 60 s. After PCR preamplification, the PCR products were added 8 µL of primer digestion mix (32 U of Exonuclease I (NEB) and 1× Exonuclease I Reaction Buffer (NEB) in nuclease-free water) and incubated at 37 °C for 30 min and 80 °C for 15 min.

    Real-time Polymerase Chain Reaction:

    Article Title: Single-cell full-length total RNA sequencing uncovers dynamics of recursive splicing and enhancer RNAs
    Article Snippet: One microliter of the RT mix (3× VILO Reaction Mix (Thermo Fisher) and 3× SuperScript Enzyme Mix (Thermo Fisher) in RNase-free water) was added to the sample plate and incubated at 25 °C for 10 min, 42 °C for 60 min, and 85 °C for 5 min. For single-cell RT-qPCR in Supplementary Fig. , the RT product was diluted 1:5 in RNase-free water for qPCR analysis. .. The PCR conditions were as follows: activation at 95 °C for 2 min; 14 cycles of denaturation at 95 °C for 30 s; annealing at 60 °C for 90 s; and extension at 72 °C for 60 s. After PCR preamplification, the PCR products were added 8 µL of primer digestion mix (32 U of Exonuclease I (NEB) and 1× Exonuclease I Reaction Buffer (NEB) in nuclease-free water) and incubated at 37 °C for 30 min and 80 °C for 15 min.

    Article Title: Single-cell full-length total RNA sequencing uncovers dynamics of recursive splicing and enhancer RNAs
    Article Snippet: One microliter of the RT mix (3× VILO Reaction Mix (Thermo Fisher) and 3× SuperScript Enzyme Mix (Thermo Fisher) in RNase-free water) was added to the sample plate and incubated at 25 °C for 10 min, 42 °C for 60 min, and 85 °C for 5 min. For single-cell RT-qPCR in Supplementary Fig. , the RT product was diluted 1:5 in RNase-free water for qPCR analysis. .. The PCR conditions were as follows: activation at 95 °C for 2 mi n; 14 cycles of denaturation at 95 °C for 30 s; annealing at 60 °C for 90 s; and extension at 72 °C for 60 s. After PCR preamplification, the PCR products were added 8 µL of primer digestion mix (32 U of Exonuclease I (NEB) and 1× Exonuclease I Reaction Buffer (NEB) in nuclease-free water) and incubated at 37 °C for 30 min and 80 °C for 15 min.

    Incubation:

    Article Title: Novel DNA methylation biomarkers show high sensitivity and specificity for blood-based detection of colorectal cancer—a clinical biomarker discovery and validation study
    Article Snippet: To remove excess primers and dNTPs, 2 μl PCR products were treated with 1 μl FastAP Thermosensitive Alkaline Phosphatase (1 μmol/μl) (Life Technologies) and 1 μl EXO1 (20,000 U/ml) (New England Biolabs) diluted 1:4 in 10x Exonuclease I Reaction buffer (New England Biolabs). .. The mix was incubated on a C1000 thermal cycler (Bio-Rad) at 37 °C for 15 min and 85 °C for 15 min. Sanger sequencing was performed using 0.5 μl Ready Reaction mix (Life Technologies), 1.5 μl Big-Dye sequencing buffer (Life Technologies), 1 μl primer (2 pmol/μl), 2 μl purified PCR product, and RNase-free water to a total volume of 10 μl.

    Article Title: Single-cell full-length total RNA sequencing uncovers dynamics of recursive splicing and enhancer RNAs
    Article Snippet: .. The PCR conditions were as follows: activation at 95 °C for 2 min; 14 cycles of denaturation at 95 °C for 30 s; annealing at 60 °C for 90 s; and extension at 72 °C for 60 s. After PCR preamplification, the PCR products were added 8 µL of primer digestion mix (32 U of Exonuclease I (NEB) and 1× Exonuclease I Reaction Buffer (NEB) in nuclease-free water) and incubated at 37 °C for 30 min and 80 °C for 15 min. .. The final products were diluted 1:87.6 and 1.5 µL of the diluted products were used for qPCR analysis.

    Article Title: Endovascular biopsy: Strategy for analyzing gene expression profiles of individual endothelial cells obtained from human vessels✩
    Article Snippet: .. To remove the unincorporated primers for best results, each sample was then mixed with 3.6 μL exonuclease treatment buffer composed of 2.52 μL water, 0.36 μL 10× Exonuclease I reaction buffer and 0.72 μL 20 units/μL Exonuclease I (New England BioLabs, Ipswich, MA), incubated at 37 °C for 30 min for digestion and 80 °C for 15 min to inactivate the exonuclease. .. Quantitative RT-PCR 48.48 nanofluidic chips and a BioMark HD system (Fluidigm, South San Francisco, CA) were used.

    Article Title: Single-cell full-length total RNA sequencing uncovers dynamics of recursive splicing and enhancer RNAs
    Article Snippet: .. The PCR conditions were as follows: activation at 95 °C for 2 mi n; 14 cycles of denaturation at 95 °C for 30 s; annealing at 60 °C for 90 s; and extension at 72 °C for 60 s. After PCR preamplification, the PCR products were added 8 µL of primer digestion mix (32 U of Exonuclease I (NEB) and 1× Exonuclease I Reaction Buffer (NEB) in nuclease-free water) and incubated at 37 °C for 30 min and 80 °C for 15 min. .. The final products were diluted 1:87.6 and 1.5 µL of the diluted products were used for qPCR analysis.

    Expressing:

    Article Title: Limited immune surveillance in lymphoid tissue by cytolytic CD4+ T cells during health and HIV disease
    Article Snippet: Paragraph title: Single-cell gene expression analysis ... Exonuclease mixture, containing of Exonuclease I (New England BioLabs), 10× Exonuclease I Reaction Buffer (New England BioLabs) and H2 O was added to each well to remove excessive primers and the plate was run on a thermocycler (37°C for 30 min, 80°C for 15 min).

    Article Title: Infantile Hemangioma Originates From A Dysregulated But Not Fully Transformed Multipotent Stem Cell
    Article Snippet: Paragraph title: Single-Cell Gene Expression ... The Exo I treatment method (total volume 3.5 μl [per 9 μl RT-STA solution]) consisted of: 2.52 μl Nuclease Free H2O, 0.36 μl Exonuclease I Reaction Buffer (10X), and 0.72 μl Exonuclease I (20units/μl) (New England BioLabs).

    Article Title: Single-cell full-length total RNA sequencing uncovers dynamics of recursive splicing and enhancer RNAs
    Article Snippet: The PCR conditions were as follows: activation at 95 °C for 2 min; 14 cycles of denaturation at 95 °C for 30 s; annealing at 60 °C for 90 s; and extension at 72 °C for 60 s. After PCR preamplification, the PCR products were added 8 µL of primer digestion mix (32 U of Exonuclease I (NEB) and 1× Exonuclease I Reaction Buffer (NEB) in nuclease-free water) and incubated at 37 °C for 30 min and 80 °C for 15 min. .. The copy number of gene expression was adjusted and estimated using the input copy number of the spike RNAs (Lys: 1000 copies; Thr: 5 copies).

    Article Title: Single-cell full-length total RNA sequencing uncovers dynamics of recursive splicing and enhancer RNAs
    Article Snippet: The PCR conditions were as follows: activation at 95 °C for 2 mi n; 14 cycles of denaturation at 95 °C for 30 s; annealing at 60 °C for 90 s; and extension at 72 °C for 60 s. After PCR preamplification, the PCR products were added 8 µL of primer digestion mix (32 U of Exonuclease I (NEB) and 1× Exonuclease I Reaction Buffer (NEB) in nuclease-free water) and incubated at 37 °C for 30 min and 80 °C for 15 min. .. The copy number of gene expression was adjusted and estimated using the input copy number of the spike RNAs (Lys: 1000 copies; Thr: 5 copies).

    Flow Cytometry:

    Article Title: Infantile Hemangioma Originates From A Dysregulated But Not Fully Transformed Multipotent Stem Cell
    Article Snippet: Single-Cell Gene Expression To survey gene expression profiles by a HT approach, fluorescence-activated cell sorting was performed using BD Aria II, 100 cells per well and for single-cell samples 1 cell directly flow sorted into the wells of 96-well plates containing Platinum Taq reverse transcriptase, polymerase master mix (Invitrogen) and primers for each gene target (SABiosciences) as per Fluidigm manufacturers Protocol 41 instructions and as described in the Nature Methods workflow for single-cell profiling . .. The Exo I treatment method (total volume 3.5 μl [per 9 μl RT-STA solution]) consisted of: 2.52 μl Nuclease Free H2O, 0.36 μl Exonuclease I Reaction Buffer (10X), and 0.72 μl Exonuclease I (20units/μl) (New England BioLabs).

    Concentration Assay:

    Article Title: Spatial distribution and function of T follicular regulatory cells in human lymph nodes
    Article Snippet: Preamplified samples were subjected to an exonuclease reaction, using Exonuclease I and Exonuclease I Reaction Buffer (New England Biolabs). .. Assays were prepared using 100-µM primers combined with 2× Assay Loading Reagent (Fluidigm) and DNA suspension buffer (TEKnova), for a final primer concentration of 5 µM.

    Article Title: Infantile Hemangioma Originates From A Dysregulated But Not Fully Transformed Multipotent Stem Cell
    Article Snippet: The Exo I treatment method (total volume 3.5 μl [per 9 μl RT-STA solution]) consisted of: 2.52 μl Nuclease Free H2O, 0.36 μl Exonuclease I Reaction Buffer (10X), and 0.72 μl Exonuclease I (20units/μl) (New England BioLabs). .. The final concentration of STAreaction + Exonuclease I was diluted 5-fold in this experiment.

    Article Title: Functional Gut Microbiota Remodeling Contributes to the Caloric Restriction-Induced Metabolic Improvements
    Article Snippet: .. The primers from the first round of PCR were removed by digesting 5 μL samples (containing 9.5-13.9 ng/μL DNA except in negative extraction controls for which DNA concentration was under the detection level) with 1 unit Exonuclease I (New England Biolabs, Ipswich, MA, USA) in a total volume of 10 μL Exonuclease I Reaction Buffer (New England Biolabs) at 37°C for 30 min. .. The enzyme was inactivated at 95°C for 15 min. Amplicon barcoding was performed by re-amplification using 1 μL of Exonuclease I-treated first-round PCR, 15 pmol each of forward primer 5′-NNNNNNNNNNTCCTACGGGNGGCWGCAG-3′ and reverse primer 5′- NNNNNNNNNNTGACTACHVGGGTATCTAAKCC-3′ in a 20-μL volume of MyTaq buffer that contained 1.5 units MyTaq DNA polymerase (Bioline, London, UK) and 2 μL of BioStab PCR optimizer (II) (Sigma-Aldrich, Munich, Germany).

    Article Title: Decontamination of 16S rRNA gene amplicon sequence datasets based on bacterial load assessment by qPCR
    Article Snippet: The primers from the first round of PCR were removed by digesting 5-μl samples with 1 unit Exonuclease I (New England Biolabs, Ipswich, MA, USA) in a total volume of 10 μl Exonuclease I Reaction Buffer (New England Biolabs) at 37 °C for 30 min. .. PCRs were carried out using the following parameters: pre-denaturation for 2 min at 96 °C, followed by eight cycles of 96 °C for 15 s, 50 °C for 30 s, and 70 °C for 90 s. DNA concentration of amplicons of interest was determined by gel electrophoresis.

    Article Title: Direct elicitation of template concentration from quantification cycle (Cq) distributions in digital PCR
    Article Snippet: Final primer concentration in pre-amplification reactions was 45 nM. .. Unincorporated primers were digested by adding a 6 μl solution containing 4 units Exonuclease I (New England BioLabs M0293L) in 1× Exonuclease I Reaction Buffer (New England BioLabs B0293S) and using the following thermal protocol: 30 min at 37°C, 15 min at 80°C, hold at 4°C.

    Digital PCR:

    Article Title: Direct elicitation of template concentration from quantification cycle (Cq) distributions in digital PCR
    Article Snippet: Unincorporated primers were digested by adding a 6 μl solution containing 4 units Exonuclease I (New England BioLabs M0293L) in 1× Exonuclease I Reaction Buffer (New England BioLabs B0293S) and using the following thermal protocol: 30 min at 37°C, 15 min at 80°C, hold at 4°C. .. Ten-fold diluted pre-amplified samples were analyzed on the dPCR platform (cf. dPCR analysis ) and reported \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{upgreek} \usepackage{mathrsfs} \setlength{\oddsidemargin}{-69pt} \begin{document} }{}$C_{\rm q}$\end{document} -values were used for the inference of transcript numbers.

    Generated:

    Article Title: Limited immune surveillance in lymphoid tissue by cytolytic CD4+ T cells during health and HIV disease
    Article Snippet: Exonuclease mixture, containing of Exonuclease I (New England BioLabs), 10× Exonuclease I Reaction Buffer (New England BioLabs) and H2 O was added to each well to remove excessive primers and the plate was run on a thermocycler (37°C for 30 min, 80°C for 15 min). .. Distinct primer assays were generated by adding individual primer pairs (5μM) together with a mix of 2x Assay Loading Reagent (Fluidigm) and 1x DNA suspension buffer to each well of a new plate.

    Article Title: T-bet+ B cells are induced by human viral infections and dominate the HIV gp140 response
    Article Snippet: To remove excess primers, 4 μl of an exonuclease mixture, containing 0.8 μl 20 units/μl of Exonuclease I (New England BioLabs), 0.4 μl 10× Exonuclease I Reaction Buffer (New England BioLabs), and 2.8 μl H2 O was added to each well. .. In a new PCR plate, distinct primer assays were generated by adding individual primer pairs (5 μM) together with a mix of 2.6 μl 2× Assay Loading Reagent (Fluidigm) and 2.4 μl 1× DNA Suspension Buffer to each well.

    Polymerase Chain Reaction:

    Article Title: Light-activated cell identification and sorting (LACIS) for selection of edited clones on a nanofluidic device
    Article Snippet: .. Excess PCR primers were removed by incubating with Exonuclease I (NEB, M0293S) in 1 × exonuclease reaction buffer (NEB, B0293S) for 1 h at 37 °C, followed by enzyme inactivation for 20 min at 80 °C. .. Amplicon pools were re-amplified by PCR for 15 cycles using a universal primer to add the sequencing adaptor and secondary barcodes to allow parallel sequencing of multiple amplicon pools (Primer sequences are listed in Supplementary Data ).

    Article Title: Limited immune surveillance in lymphoid tissue by cytolytic CD4+ T cells during health and HIV disease
    Article Snippet: Similar to before [ ], PCR plates were thawed and pre-heated for 90 seconds at 65°C. .. Exonuclease mixture, containing of Exonuclease I (New England BioLabs), 10× Exonuclease I Reaction Buffer (New England BioLabs) and H2 O was added to each well to remove excessive primers and the plate was run on a thermocycler (37°C for 30 min, 80°C for 15 min).

    Article Title: Novel DNA methylation biomarkers show high sensitivity and specificity for blood-based detection of colorectal cancer—a clinical biomarker discovery and validation study
    Article Snippet: .. To remove excess primers and dNTPs, 2 μl PCR products were treated with 1 μl FastAP Thermosensitive Alkaline Phosphatase (1 μmol/μl) (Life Technologies) and 1 μl EXO1 (20,000 U/ml) (New England Biolabs) diluted 1:4 in 10x Exonuclease I Reaction buffer (New England Biolabs). .. The mix was incubated on a C1000 thermal cycler (Bio-Rad) at 37 °C for 15 min and 85 °C for 15 min. Sanger sequencing was performed using 0.5 μl Ready Reaction mix (Life Technologies), 1.5 μl Big-Dye sequencing buffer (Life Technologies), 1 μl primer (2 pmol/μl), 2 μl purified PCR product, and RNase-free water to a total volume of 10 μl.

    Article Title: Functional Gut Microbiota Remodeling Contributes to the Caloric Restriction-Induced Metabolic Improvements
    Article Snippet: .. The primers from the first round of PCR were removed by digesting 5 μL samples (containing 9.5-13.9 ng/μL DNA except in negative extraction controls for which DNA concentration was under the detection level) with 1 unit Exonuclease I (New England Biolabs, Ipswich, MA, USA) in a total volume of 10 μL Exonuclease I Reaction Buffer (New England Biolabs) at 37°C for 30 min. .. The enzyme was inactivated at 95°C for 15 min. Amplicon barcoding was performed by re-amplification using 1 μL of Exonuclease I-treated first-round PCR, 15 pmol each of forward primer 5′-NNNNNNNNNNTCCTACGGGNGGCWGCAG-3′ and reverse primer 5′- NNNNNNNNNNTGACTACHVGGGTATCTAAKCC-3′ in a 20-μL volume of MyTaq buffer that contained 1.5 units MyTaq DNA polymerase (Bioline, London, UK) and 2 μL of BioStab PCR optimizer (II) (Sigma-Aldrich, Munich, Germany).

    Article Title: Decontamination of 16S rRNA gene amplicon sequence datasets based on bacterial load assessment by qPCR
    Article Snippet: .. The primers from the first round of PCR were removed by digesting 5-μl samples with 1 unit Exonuclease I (New England Biolabs, Ipswich, MA, USA) in a total volume of 10 μl Exonuclease I Reaction Buffer (New England Biolabs) at 37 °C for 30 min. .. The enzyme was inactivated at 95 °C for 15 min. Amplicon barcoding was performed by re-amplification using 1 μl of Exonuclease I-treated first-round PCR, 15 pmol each of forward primer 5’-NNNNNNNNNNTCCTACGGGNGGCWGCAG-3’ and reverse primer 5’- NNNNNNNNNNTGACTACHVGGGTATCTAAKCC-3’ in a 20-μL volume of MyTaq buffer that contained 1.5 units MyTaq DNA polymerase (Bioline, London, UK) and 2 μl of BioStab PCR optimizer (II) (Sigma-Aldrich, Munich, Germany).

    Article Title: Single-cell full-length total RNA sequencing uncovers dynamics of recursive splicing and enhancer RNAs
    Article Snippet: .. The PCR conditions were as follows: activation at 95 °C for 2 min; 14 cycles of denaturation at 95 °C for 30 s; annealing at 60 °C for 90 s; and extension at 72 °C for 60 s. After PCR preamplification, the PCR products were added 8 µL of primer digestion mix (32 U of Exonuclease I (NEB) and 1× Exonuclease I Reaction Buffer (NEB) in nuclease-free water) and incubated at 37 °C for 30 min and 80 °C for 15 min. .. The final products were diluted 1:87.6 and 1.5 µL of the diluted products were used for qPCR analysis.

    Article Title: Endovascular biopsy: Strategy for analyzing gene expression profiles of individual endothelial cells obtained from human vessels✩
    Article Snippet: The samples were incubated at 50 °C for 15 min for the reverse transcription, 95 °C for 2 min for inactivating reverse transcriptase and activating Taq polymerase, then subjected to 18 PCR cycles (95 °C 15 sec then 60 °C for 4 min for each cycle) for specific targets amplification (STA). .. To remove the unincorporated primers for best results, each sample was then mixed with 3.6 μL exonuclease treatment buffer composed of 2.52 μL water, 0.36 μL 10× Exonuclease I reaction buffer and 0.72 μL 20 units/μL Exonuclease I (New England BioLabs, Ipswich, MA), incubated at 37 °C for 30 min for digestion and 80 °C for 15 min to inactivate the exonuclease.

    Article Title: Single-cell full-length total RNA sequencing uncovers dynamics of recursive splicing and enhancer RNAs
    Article Snippet: .. The PCR conditions were as follows: activation at 95 °C for 2 mi n; 14 cycles of denaturation at 95 °C for 30 s; annealing at 60 °C for 90 s; and extension at 72 °C for 60 s. After PCR preamplification, the PCR products were added 8 µL of primer digestion mix (32 U of Exonuclease I (NEB) and 1× Exonuclease I Reaction Buffer (NEB) in nuclease-free water) and incubated at 37 °C for 30 min and 80 °C for 15 min. .. The final products were diluted 1:87.6 and 1.5 µL of the diluted products were used for qPCR analysis.

    Article Title: T-bet+ B cells are induced by human viral infections and dominate the HIV gp140 response
    Article Snippet: PCR plates were thawed on ice and preheated for 90 seconds at 65°C. .. To remove excess primers, 4 μl of an exonuclease mixture, containing 0.8 μl 20 units/μl of Exonuclease I (New England BioLabs), 0.4 μl 10× Exonuclease I Reaction Buffer (New England BioLabs), and 2.8 μl H2 O was added to each well.

    Binding Assay:

    Article Title: Spatial distribution and function of T follicular regulatory cells in human lymph nodes
    Article Snippet: Preamplified samples were subjected to an exonuclease reaction, using Exonuclease I and Exonuclease I Reaction Buffer (New England Biolabs). .. An aliquot of each of the diluted, preamplified samples was then mixed with SsoFast EvaGreen Supermix with Low Rox (Bio-Rad) and DNA Binding Dye Sample Loading Reagent (Fluidigm).

    Article Title: Limited immune surveillance in lymphoid tissue by cytolytic CD4+ T cells during health and HIV disease
    Article Snippet: Exonuclease mixture, containing of Exonuclease I (New England BioLabs), 10× Exonuclease I Reaction Buffer (New England BioLabs) and H2 O was added to each well to remove excessive primers and the plate was run on a thermocycler (37°C for 30 min, 80°C for 15 min). .. A “sample PCR plate” was created by dispensing a sample master mix, containing 2x Sso Fast EvaGreen Supermix with Low ROX (Bio-Rad), 20x DNA Binding Dye Sample Loading Reagent (Fluidigm) and H2O to each well.

    Nucleic Acid Electrophoresis:

    Article Title: Functional Gut Microbiota Remodeling Contributes to the Caloric Restriction-Induced Metabolic Improvements
    Article Snippet: The primers from the first round of PCR were removed by digesting 5 μL samples (containing 9.5-13.9 ng/μL DNA except in negative extraction controls for which DNA concentration was under the detection level) with 1 unit Exonuclease I (New England Biolabs, Ipswich, MA, USA) in a total volume of 10 μL Exonuclease I Reaction Buffer (New England Biolabs) at 37°C for 30 min. .. PCRs were carried out using the following parameters: pre-denaturation for 2 min at 96°C, followed by eight cycles of 96°C for 15 s, 50°C for 30 s, and 70°C for 90 s. DNA concentration of amplicons of interest was determined by gel electrophoresis.

    Article Title: Decontamination of 16S rRNA gene amplicon sequence datasets based on bacterial load assessment by qPCR
    Article Snippet: The primers from the first round of PCR were removed by digesting 5-μl samples with 1 unit Exonuclease I (New England Biolabs, Ipswich, MA, USA) in a total volume of 10 μl Exonuclease I Reaction Buffer (New England Biolabs) at 37 °C for 30 min. .. PCRs were carried out using the following parameters: pre-denaturation for 2 min at 96 °C, followed by eight cycles of 96 °C for 15 s, 50 °C for 30 s, and 70 °C for 90 s. DNA concentration of amplicons of interest was determined by gel electrophoresis.

    Fluorescence:

    Article Title: Infantile Hemangioma Originates From A Dysregulated But Not Fully Transformed Multipotent Stem Cell
    Article Snippet: Single-Cell Gene Expression To survey gene expression profiles by a HT approach, fluorescence-activated cell sorting was performed using BD Aria II, 100 cells per well and for single-cell samples 1 cell directly flow sorted into the wells of 96-well plates containing Platinum Taq reverse transcriptase, polymerase master mix (Invitrogen) and primers for each gene target (SABiosciences) as per Fluidigm manufacturers Protocol 41 instructions and as described in the Nature Methods workflow for single-cell profiling . .. The Exo I treatment method (total volume 3.5 μl [per 9 μl RT-STA solution]) consisted of: 2.52 μl Nuclease Free H2O, 0.36 μl Exonuclease I Reaction Buffer (10X), and 0.72 μl Exonuclease I (20units/μl) (New England BioLabs).

    Isolation:

    Article Title: Spatial distribution and function of T follicular regulatory cells in human lymph nodes
    Article Snippet: Real-time quantitative RT-PCR RNA was first isolated from sorted cell populations using the RNeasy Micro Plus kit (Qiagen). .. Preamplified samples were subjected to an exonuclease reaction, using Exonuclease I and Exonuclease I Reaction Buffer (New England Biolabs).

    Article Title: Light-activated cell identification and sorting (LACIS) for selection of edited clones on a nanofluidic device
    Article Snippet: Excess PCR primers were removed by incubating with Exonuclease I (NEB, M0293S) in 1 × exonuclease reaction buffer (NEB, B0293S) for 1 h at 37 °C, followed by enzyme inactivation for 20 min at 80 °C. .. PCR products of the expected size were isolated with Select-A-Size DNA Clean and Concentrator (Zymo research, D4080) as sequencing libraries.

    Size-exclusion Chromatography:

    Article Title: Limited immune surveillance in lymphoid tissue by cytolytic CD4+ T cells during health and HIV disease
    Article Snippet: Next, pre-amplification mix, consisting of pooled mixture of all primer assays (500nM), 5× PreAmp Master Mix (Fluidigm) and H2 O, was added to each well and run on a thermocycler (95°C for 5 min followed by 18 cycles: 96°C for 5 sec 60°C for 6 min). .. Exonuclease mixture, containing of Exonuclease I (New England BioLabs), 10× Exonuclease I Reaction Buffer (New England BioLabs) and H2 O was added to each well to remove excessive primers and the plate was run on a thermocycler (37°C for 30 min, 80°C for 15 min).

    Article Title: Infantile Hemangioma Originates From A Dysregulated But Not Fully Transformed Multipotent Stem Cell
    Article Snippet: The thermal cycling profile protocol was as follows: Stage 1: 50 °C, 15 min; Stage 2: 95 °C, 2 min; Stage 3: 20 cycles of (95 °C, 15 sec; and 60 °C, 4 min). .. The Exo I treatment method (total volume 3.5 μl [per 9 μl RT-STA solution]) consisted of: 2.52 μl Nuclease Free H2O, 0.36 μl Exonuclease I Reaction Buffer (10X), and 0.72 μl Exonuclease I (20units/μl) (New England BioLabs).

    Article Title: Endovascular biopsy: Strategy for analyzing gene expression profiles of individual endothelial cells obtained from human vessels✩
    Article Snippet: The samples were incubated at 50 °C for 15 min for the reverse transcription, 95 °C for 2 min for inactivating reverse transcriptase and activating Taq polymerase, then subjected to 18 PCR cycles (95 °C 15 sec then 60 °C for 4 min for each cycle) for specific targets amplification (STA). .. To remove the unincorporated primers for best results, each sample was then mixed with 3.6 μL exonuclease treatment buffer composed of 2.52 μL water, 0.36 μL 10× Exonuclease I reaction buffer and 0.72 μL 20 units/μL Exonuclease I (New England BioLabs, Ipswich, MA), incubated at 37 °C for 30 min for digestion and 80 °C for 15 min to inactivate the exonuclease.

    Purification:

    Article Title: Novel DNA methylation biomarkers show high sensitivity and specificity for blood-based detection of colorectal cancer—a clinical biomarker discovery and validation study
    Article Snippet: To remove excess primers and dNTPs, 2 μl PCR products were treated with 1 μl FastAP Thermosensitive Alkaline Phosphatase (1 μmol/μl) (Life Technologies) and 1 μl EXO1 (20,000 U/ml) (New England Biolabs) diluted 1:4 in 10x Exonuclease I Reaction buffer (New England Biolabs). .. The mix was incubated on a C1000 thermal cycler (Bio-Rad) at 37 °C for 15 min and 85 °C for 15 min. Sanger sequencing was performed using 0.5 μl Ready Reaction mix (Life Technologies), 1.5 μl Big-Dye sequencing buffer (Life Technologies), 1 μl primer (2 pmol/μl), 2 μl purified PCR product, and RNase-free water to a total volume of 10 μl.

    Sequencing:

    Article Title: Light-activated cell identification and sorting (LACIS) for selection of edited clones on a nanofluidic device
    Article Snippet: Sample processing for next-generation sequencing Genomic DNA was extracted from exported clones by incubating in Proteinase K buffer (0.1 M NaCl, 10 mM Tris-HCl pH 8.0, 1 mM EDTA) for 30 min at 55 °C, then for 20 min at 80 °C to inactivate Proteinase K. The genomic region around the CRISPR/Cas9 target site for CXCR4 gene was amplified by PCR with primers positioned outside of the HDR repair template sequence (positioned to avoid amplification of exogenous template) for ten cycles using KAPA HiFi Hotstart ReadyMix (Kapa Biosystems, KR0370) according to the manufacturer’s protocol (PCR primers listed in Supplementary Data ). .. Excess PCR primers were removed by incubating with Exonuclease I (NEB, M0293S) in 1 × exonuclease reaction buffer (NEB, B0293S) for 1 h at 37 °C, followed by enzyme inactivation for 20 min at 80 °C.

    Article Title: Novel DNA methylation biomarkers show high sensitivity and specificity for blood-based detection of colorectal cancer—a clinical biomarker discovery and validation study
    Article Snippet: To remove excess primers and dNTPs, 2 μl PCR products were treated with 1 μl FastAP Thermosensitive Alkaline Phosphatase (1 μmol/μl) (Life Technologies) and 1 μl EXO1 (20,000 U/ml) (New England Biolabs) diluted 1:4 in 10x Exonuclease I Reaction buffer (New England Biolabs). .. The mix was incubated on a C1000 thermal cycler (Bio-Rad) at 37 °C for 15 min and 85 °C for 15 min. Sanger sequencing was performed using 0.5 μl Ready Reaction mix (Life Technologies), 1.5 μl Big-Dye sequencing buffer (Life Technologies), 1 μl primer (2 pmol/μl), 2 μl purified PCR product, and RNase-free water to a total volume of 10 μl.

    Article Title: Functional Gut Microbiota Remodeling Contributes to the Caloric Restriction-Induced Metabolic Improvements
    Article Snippet: Paragraph title: Bacterial 16S Sequencing ... The primers from the first round of PCR were removed by digesting 5 μL samples (containing 9.5-13.9 ng/μL DNA except in negative extraction controls for which DNA concentration was under the detection level) with 1 unit Exonuclease I (New England Biolabs, Ipswich, MA, USA) in a total volume of 10 μL Exonuclease I Reaction Buffer (New England Biolabs) at 37°C for 30 min.

    Article Title: Decontamination of 16S rRNA gene amplicon sequence datasets based on bacterial load assessment by qPCR
    Article Snippet: Paragraph title: PCR and sequencing ... The primers from the first round of PCR were removed by digesting 5-μl samples with 1 unit Exonuclease I (New England Biolabs, Ipswich, MA, USA) in a total volume of 10 μl Exonuclease I Reaction Buffer (New England Biolabs) at 37 °C for 30 min.

    CRISPR:

    Article Title: Light-activated cell identification and sorting (LACIS) for selection of edited clones on a nanofluidic device
    Article Snippet: Sample processing for next-generation sequencing Genomic DNA was extracted from exported clones by incubating in Proteinase K buffer (0.1 M NaCl, 10 mM Tris-HCl pH 8.0, 1 mM EDTA) for 30 min at 55 °C, then for 20 min at 80 °C to inactivate Proteinase K. The genomic region around the CRISPR/Cas9 target site for CXCR4 gene was amplified by PCR with primers positioned outside of the HDR repair template sequence (positioned to avoid amplification of exogenous template) for ten cycles using KAPA HiFi Hotstart ReadyMix (Kapa Biosystems, KR0370) according to the manufacturer’s protocol (PCR primers listed in Supplementary Data ). .. Excess PCR primers were removed by incubating with Exonuclease I (NEB, M0293S) in 1 × exonuclease reaction buffer (NEB, B0293S) for 1 h at 37 °C, followed by enzyme inactivation for 20 min at 80 °C.

    cDNA Library Assay:

    Article Title: Infantile Hemangioma Originates From A Dysregulated But Not Fully Transformed Multipotent Stem Cell
    Article Snippet: The Exo I treatment method (total volume 3.5 μl [per 9 μl RT-STA solution]) consisted of: 2.52 μl Nuclease Free H2O, 0.36 μl Exonuclease I Reaction Buffer (10X), and 0.72 μl Exonuclease I (20units/μl) (New England BioLabs). .. The human cDNA library (Biochain) was treated with RTA-STA/ExoI treatment method, diluted 5-fold with additional serial dilutions (1:3).

    Software:

    Article Title: Novel DNA methylation biomarkers show high sensitivity and specificity for blood-based detection of colorectal cancer—a clinical biomarker discovery and validation study
    Article Snippet: To remove excess primers and dNTPs, 2 μl PCR products were treated with 1 μl FastAP Thermosensitive Alkaline Phosphatase (1 μmol/μl) (Life Technologies) and 1 μl EXO1 (20,000 U/ml) (New England Biolabs) diluted 1:4 in 10x Exonuclease I Reaction buffer (New England Biolabs). .. Results were analysed using Sequencer 5.1 software (Gene Codes Cooperation).

    Multiplex Assay:

    Article Title: Single-cell full-length total RNA sequencing uncovers dynamics of recursive splicing and enhancer RNAs
    Article Snippet: Seventeen microliters of the preamplification mix (10 µL of 2× Platinum Multiplex PCR Master Mix (Thermo Fisher), and 2 µL of 10× pooled primer mix in nuclease-free water) was added to 3 µL of the RT products. .. The PCR conditions were as follows: activation at 95 °C for 2 min; 14 cycles of denaturation at 95 °C for 30 s; annealing at 60 °C for 90 s; and extension at 72 °C for 60 s. After PCR preamplification, the PCR products were added 8 µL of primer digestion mix (32 U of Exonuclease I (NEB) and 1× Exonuclease I Reaction Buffer (NEB) in nuclease-free water) and incubated at 37 °C for 30 min and 80 °C for 15 min.

    Article Title: Single-cell full-length total RNA sequencing uncovers dynamics of recursive splicing and enhancer RNAs
    Article Snippet: Seventeen microliters of the preamplification mix (10 µL of 2× Platinum Multiplex PCR Master Mix (Thermo Fisher), and 2 µL of 10× pooled primer mix in nuclease-free water) was added to 3 µL of the RT products. .. The PCR conditions were as follows: activation at 95 °C for 2 mi n; 14 cycles of denaturation at 95 °C for 30 s; annealing at 60 °C for 90 s; and extension at 72 °C for 60 s. After PCR preamplification, the PCR products were added 8 µL of primer digestion mix (32 U of Exonuclease I (NEB) and 1× Exonuclease I Reaction Buffer (NEB) in nuclease-free water) and incubated at 37 °C for 30 min and 80 °C for 15 min.

    Agarose Gel Electrophoresis:

    Article Title: Novel DNA methylation biomarkers show high sensitivity and specificity for blood-based detection of colorectal cancer—a clinical biomarker discovery and validation study
    Article Snippet: Following PCR amplification, 5 μl of each amplicon were visualized by agarose gel electrophoresis in Tris-acetate-EDTA (TAE) buffer (Fagron) with ethidium bromide (EtBr) to test for correct band size. .. To remove excess primers and dNTPs, 2 μl PCR products were treated with 1 μl FastAP Thermosensitive Alkaline Phosphatase (1 μmol/μl) (Life Technologies) and 1 μl EXO1 (20,000 U/ml) (New England Biolabs) diluted 1:4 in 10x Exonuclease I Reaction buffer (New England Biolabs).

    Next-Generation Sequencing:

    Article Title: Light-activated cell identification and sorting (LACIS) for selection of edited clones on a nanofluidic device
    Article Snippet: Paragraph title: Sample processing for next-generation sequencing ... Excess PCR primers were removed by incubating with Exonuclease I (NEB, M0293S) in 1 × exonuclease reaction buffer (NEB, B0293S) for 1 h at 37 °C, followed by enzyme inactivation for 20 min at 80 °C.

    Activation Assay:

    Article Title: Single-cell full-length total RNA sequencing uncovers dynamics of recursive splicing and enhancer RNAs
    Article Snippet: .. The PCR conditions were as follows: activation at 95 °C for 2 min; 14 cycles of denaturation at 95 °C for 30 s; annealing at 60 °C for 90 s; and extension at 72 °C for 60 s. After PCR preamplification, the PCR products were added 8 µL of primer digestion mix (32 U of Exonuclease I (NEB) and 1× Exonuclease I Reaction Buffer (NEB) in nuclease-free water) and incubated at 37 °C for 30 min and 80 °C for 15 min. .. The final products were diluted 1:87.6 and 1.5 µL of the diluted products were used for qPCR analysis.

    Article Title: Single-cell full-length total RNA sequencing uncovers dynamics of recursive splicing and enhancer RNAs
    Article Snippet: .. The PCR conditions were as follows: activation at 95 °C for 2 mi n; 14 cycles of denaturation at 95 °C for 30 s; annealing at 60 °C for 90 s; and extension at 72 °C for 60 s. After PCR preamplification, the PCR products were added 8 µL of primer digestion mix (32 U of Exonuclease I (NEB) and 1× Exonuclease I Reaction Buffer (NEB) in nuclease-free water) and incubated at 37 °C for 30 min and 80 °C for 15 min. .. The final products were diluted 1:87.6 and 1.5 µL of the diluted products were used for qPCR analysis.

    Marker:

    Article Title: Novel DNA methylation biomarkers show high sensitivity and specificity for blood-based detection of colorectal cancer—a clinical biomarker discovery and validation study
    Article Snippet: GeneRuler™ 100 bp Plus DNA Ladder (Thermo Scientific) was used as a molecular marker. .. To remove excess primers and dNTPs, 2 μl PCR products were treated with 1 μl FastAP Thermosensitive Alkaline Phosphatase (1 μmol/μl) (Life Technologies) and 1 μl EXO1 (20,000 U/ml) (New England Biolabs) diluted 1:4 in 10x Exonuclease I Reaction buffer (New England Biolabs).

    FACS:

    Article Title: Limited immune surveillance in lymphoid tissue by cytolytic CD4+ T cells during health and HIV disease
    Article Snippet: After FACS sorting, PCR plates were frozen and kept in -80°C until usage. .. Exonuclease mixture, containing of Exonuclease I (New England BioLabs), 10× Exonuclease I Reaction Buffer (New England BioLabs) and H2 O was added to each well to remove excessive primers and the plate was run on a thermocycler (37°C for 30 min, 80°C for 15 min).

    Article Title: Infantile Hemangioma Originates From A Dysregulated But Not Fully Transformed Multipotent Stem Cell
    Article Snippet: Single-Cell Gene Expression To survey gene expression profiles by a HT approach, fluorescence-activated cell sorting was performed using BD Aria II, 100 cells per well and for single-cell samples 1 cell directly flow sorted into the wells of 96-well plates containing Platinum Taq reverse transcriptase, polymerase master mix (Invitrogen) and primers for each gene target (SABiosciences) as per Fluidigm manufacturers Protocol 41 instructions and as described in the Nature Methods workflow for single-cell profiling . .. The Exo I treatment method (total volume 3.5 μl [per 9 μl RT-STA solution]) consisted of: 2.52 μl Nuclease Free H2O, 0.36 μl Exonuclease I Reaction Buffer (10X), and 0.72 μl Exonuclease I (20units/μl) (New England BioLabs).

    Article Title: T-bet+ B cells are induced by human viral infections and dominate the HIV gp140 response
    Article Snippet: Paragraph title: Cell sorting and Fluidigm Biomark transcript analysis. ... To remove excess primers, 4 μl of an exonuclease mixture, containing 0.8 μl 20 units/μl of Exonuclease I (New England BioLabs), 0.4 μl 10× Exonuclease I Reaction Buffer (New England BioLabs), and 2.8 μl H2 O was added to each well.

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    New England Biolabs λ exonuclease reaction buffer
    ChIP-exo 5.0 increases library yield. a Schematic of ChIP-exo 5.0. The purple triangle indicates the location of the Read_1 start site, which is also the <t>λ</t> exonuclease stop site. b 2% agarose gel of the electrophoresed library following 18 cycles of PCR for various S. cerevisiae transcription factors assayed by ChIP-exo 1.1 or 5.0. Following ChIP, the sample was split and libraries prepared using the indicated protocols. After splitting the sample, each reaction contained a 50 ml cell equivalent (OD 600 = 0.8) of yeast chromatin, which is five-fold less than the amount optimized for ChIP-exo 1.1. ChIP-exo 5.0 produced greater library yield for all samples. c Heatmaps comparing ChIP-exo 1.1 and 5.0 at the 975 Reb1 primary motifs in a 200 bp window. d Composite plot of data from panel ( c )
    λ Exonuclease Reaction Buffer, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 15 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    ChIP-exo 5.0 increases library yield. a Schematic of ChIP-exo 5.0. The purple triangle indicates the location of the Read_1 start site, which is also the λ exonuclease stop site. b 2% agarose gel of the electrophoresed library following 18 cycles of PCR for various S. cerevisiae transcription factors assayed by ChIP-exo 1.1 or 5.0. Following ChIP, the sample was split and libraries prepared using the indicated protocols. After splitting the sample, each reaction contained a 50 ml cell equivalent (OD 600 = 0.8) of yeast chromatin, which is five-fold less than the amount optimized for ChIP-exo 1.1. ChIP-exo 5.0 produced greater library yield for all samples. c Heatmaps comparing ChIP-exo 1.1 and 5.0 at the 975 Reb1 primary motifs in a 200 bp window. d Composite plot of data from panel ( c )

    Journal: Nature Communications

    Article Title: Simplified ChIP-exo assays

    doi: 10.1038/s41467-018-05265-7

    Figure Lengend Snippet: ChIP-exo 5.0 increases library yield. a Schematic of ChIP-exo 5.0. The purple triangle indicates the location of the Read_1 start site, which is also the λ exonuclease stop site. b 2% agarose gel of the electrophoresed library following 18 cycles of PCR for various S. cerevisiae transcription factors assayed by ChIP-exo 1.1 or 5.0. Following ChIP, the sample was split and libraries prepared using the indicated protocols. After splitting the sample, each reaction contained a 50 ml cell equivalent (OD 600 = 0.8) of yeast chromatin, which is five-fold less than the amount optimized for ChIP-exo 1.1. ChIP-exo 5.0 produced greater library yield for all samples. c Heatmaps comparing ChIP-exo 1.1 and 5.0 at the 975 Reb1 primary motifs in a 200 bp window. d Composite plot of data from panel ( c )

    Article Snippet: The λ exonuclease digestion (100 µl) containing: 20 U λ exonuclease (NEB), 1 × λ exonuclease reaction buffer (NEB), 0.1% Triton-X 100, and 5% DMSO was incubated for 30 min at 37 °C; then washed with 10 mM Tris-HCl, pH 8.0 at 4 °C.

    Techniques: Chromatin Immunoprecipitation, Agarose Gel Electrophoresis, Polymerase Chain Reaction, Produced

    ChIP-exo 5.0 increases library yield. a Schematic of ChIP-exo 5.0. The purple triangle indicates the location of the Read_1 start site, which is also the λ exonuclease stop site. b 2% agarose gel of the electrophoresed library following 18 cycles of PCR for various S. cerevisiae transcription factors assayed by ChIP-exo 1.1 or 5.0. Following ChIP, the sample was split and libraries prepared using the indicated protocols. After splitting the sample, each reaction contained a 50 ml cell equivalent (OD 600 = 0.8) of yeast chromatin, which is five-fold less than the amount optimized for ChIP-exo 1.1. ChIP-exo 5.0 produced greater library yield for all samples. c Heatmaps comparing ChIP-exo 1.1 and 5.0 at the 975 Reb1 primary motifs in a 200 bp window. d Composite plot of data from panel ( c )

    Journal: Nature Communications

    Article Title: Simplified ChIP-exo assays

    doi: 10.1038/s41467-018-05265-7

    Figure Lengend Snippet: ChIP-exo 5.0 increases library yield. a Schematic of ChIP-exo 5.0. The purple triangle indicates the location of the Read_1 start site, which is also the λ exonuclease stop site. b 2% agarose gel of the electrophoresed library following 18 cycles of PCR for various S. cerevisiae transcription factors assayed by ChIP-exo 1.1 or 5.0. Following ChIP, the sample was split and libraries prepared using the indicated protocols. After splitting the sample, each reaction contained a 50 ml cell equivalent (OD 600 = 0.8) of yeast chromatin, which is five-fold less than the amount optimized for ChIP-exo 1.1. ChIP-exo 5.0 produced greater library yield for all samples. c Heatmaps comparing ChIP-exo 1.1 and 5.0 at the 975 Reb1 primary motifs in a 200 bp window. d Composite plot of data from panel ( c )

    Article Snippet: The λ exonuclease digestion (100 µl) containing: 20 U λ exonuclease (NEB), 1 × λ exonuclease reaction buffer (NEB), 0.1% Triton-X 100, and 5% DMSO was incubated for 30 min at 37 °C; then washed with 10 mM Tris-HCl, pH 8.0 at 4 °C.

    Techniques: Chromatin Immunoprecipitation, Agarose Gel Electrophoresis, Polymerase Chain Reaction, Produced