dna fragments  (New England Biolabs)


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    NEBNext High Fidelity 2X PCR Master Mix
    Description:
    NEBNext High Fidelity 2X PCR Master Mix 250 rxns
    Catalog Number:
    m0541l
    Price:
    360
    Size:
    250 rxns
    Category:
    Thermostable DNA Polymerases
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    New England Biolabs dna fragments
    NEBNext High Fidelity 2X PCR Master Mix
    NEBNext High Fidelity 2X PCR Master Mix 250 rxns
    https://www.bioz.com/result/dna fragments/product/New England Biolabs
    Average 99 stars, based on 15367 article reviews
    Price from $9.99 to $1999.99
    dna fragments - by Bioz Stars, 2020-03
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    Images

    1) Product Images from "Epigenetic changes around the pX region and spontaneous HTLV-1 transcription are CTCF-independent"

    Article Title: Epigenetic changes around the pX region and spontaneous HTLV-1 transcription are CTCF-independent

    Journal: Wellcome Open Research

    doi: 10.12688/wellcomeopenres.14741.2

    DNA methylation across the body of the HTLV-1 provirus. ( a ) Upper panel: count of CpG dinucleotides in a window of 350 bp in the HTLV-1 reference genome (L36905). Lower panel: schematic diagram of HTLV-1 provirus indicating the two LTRs and the 9 loci examined by MeDIP. ( b ) DNA methylation on the HTLV-1 provirus in the Tax + and Tax – populations from two HTLV-1-infected T cell clones (Clones TBX4B and 11.65). ( c ) DNA methylation on the HTLV-1 provirus in the CADM1 + Tax + and CADM1 + Tax – populations in PBMCs from two unrelated individuals (Patients TDZ and TED). The asterisk (*) indicates that the PCR failed to amplify.
    Figure Legend Snippet: DNA methylation across the body of the HTLV-1 provirus. ( a ) Upper panel: count of CpG dinucleotides in a window of 350 bp in the HTLV-1 reference genome (L36905). Lower panel: schematic diagram of HTLV-1 provirus indicating the two LTRs and the 9 loci examined by MeDIP. ( b ) DNA methylation on the HTLV-1 provirus in the Tax + and Tax – populations from two HTLV-1-infected T cell clones (Clones TBX4B and 11.65). ( c ) DNA methylation on the HTLV-1 provirus in the CADM1 + Tax + and CADM1 + Tax – populations in PBMCs from two unrelated individuals (Patients TDZ and TED). The asterisk (*) indicates that the PCR failed to amplify.

    Techniques Used: DNA Methylation Assay, Methylated DNA Immunoprecipitation, Infection, Clone Assay, Polymerase Chain Reaction

    2) Product Images from "Transcriptome-wide identification of novel circular RNAs in soybean in response to low-phosphorus stress"

    Article Title: Transcriptome-wide identification of novel circular RNAs in soybean in response to low-phosphorus stress

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0227243

    Validation of a circRNAs (novel_circ_000237) by qPCR and Sanger sequencing. M, DL500 marker and the red arrow represent 100 bp. The yellow arrows denote the divergent primers for PCR amplification orientation.
    Figure Legend Snippet: Validation of a circRNAs (novel_circ_000237) by qPCR and Sanger sequencing. M, DL500 marker and the red arrow represent 100 bp. The yellow arrows denote the divergent primers for PCR amplification orientation.

    Techniques Used: Real-time Polymerase Chain Reaction, Sequencing, Marker, Polymerase Chain Reaction, Amplification

    3) Product Images from "Epigenetic changes around the pX region and spontaneous HTLV-1 transcription are CTCF-independent"

    Article Title: Epigenetic changes around the pX region and spontaneous HTLV-1 transcription are CTCF-independent

    Journal: Wellcome Open Research

    doi: 10.12688/wellcomeopenres.14741.2

    DNA methylation across the body of the HTLV-1 provirus. ( a ) Upper panel: count of CpG dinucleotides in a window of 350 bp in the HTLV-1 reference genome (L36905). Lower panel: schematic diagram of HTLV-1 provirus indicating the two LTRs and the 9 loci examined by MeDIP. ( b ) DNA methylation on the HTLV-1 provirus in the Tax + and Tax – populations from two HTLV-1-infected T cell clones (Clones TBX4B and 11.65). ( c ) DNA methylation on the HTLV-1 provirus in the CADM1 + Tax + and CADM1 + Tax – populations in PBMCs from two unrelated individuals (Patients TDZ and TED). The asterisk (*) indicates that the PCR failed to amplify.
    Figure Legend Snippet: DNA methylation across the body of the HTLV-1 provirus. ( a ) Upper panel: count of CpG dinucleotides in a window of 350 bp in the HTLV-1 reference genome (L36905). Lower panel: schematic diagram of HTLV-1 provirus indicating the two LTRs and the 9 loci examined by MeDIP. ( b ) DNA methylation on the HTLV-1 provirus in the Tax + and Tax – populations from two HTLV-1-infected T cell clones (Clones TBX4B and 11.65). ( c ) DNA methylation on the HTLV-1 provirus in the CADM1 + Tax + and CADM1 + Tax – populations in PBMCs from two unrelated individuals (Patients TDZ and TED). The asterisk (*) indicates that the PCR failed to amplify.

    Techniques Used: DNA Methylation Assay, Methylated DNA Immunoprecipitation, Infection, Clone Assay, Polymerase Chain Reaction

    4) Product Images from "Epigenetic changes around the pX region and spontaneous HTLV-1 transcription are CTCF-independent"

    Article Title: Epigenetic changes around the pX region and spontaneous HTLV-1 transcription are CTCF-independent

    Journal: Wellcome Open Research

    doi: 10.12688/wellcomeopenres.14741.2

    DNA methylation across the body of the HTLV-1 provirus. ( a ) Upper panel: count of CpG dinucleotides in a window of 350 bp in the HTLV-1 reference genome (L36905). Lower panel: schematic diagram of HTLV-1 provirus indicating the two LTRs and the 9 loci examined by MeDIP. ( b ) DNA methylation on the HTLV-1 provirus in the Tax + and Tax – populations from two HTLV-1-infected T cell clones (Clones TBX4B and 11.65). ( c ) DNA methylation on the HTLV-1 provirus in the CADM1 + Tax + and CADM1 + Tax – populations in PBMCs from two unrelated individuals (Patients TDZ and TED). The asterisk (*) indicates that the PCR failed to amplify.
    Figure Legend Snippet: DNA methylation across the body of the HTLV-1 provirus. ( a ) Upper panel: count of CpG dinucleotides in a window of 350 bp in the HTLV-1 reference genome (L36905). Lower panel: schematic diagram of HTLV-1 provirus indicating the two LTRs and the 9 loci examined by MeDIP. ( b ) DNA methylation on the HTLV-1 provirus in the Tax + and Tax – populations from two HTLV-1-infected T cell clones (Clones TBX4B and 11.65). ( c ) DNA methylation on the HTLV-1 provirus in the CADM1 + Tax + and CADM1 + Tax – populations in PBMCs from two unrelated individuals (Patients TDZ and TED). The asterisk (*) indicates that the PCR failed to amplify.

    Techniques Used: DNA Methylation Assay, Methylated DNA Immunoprecipitation, Infection, Clone Assay, Polymerase Chain Reaction

    5) Product Images from "Epigenetic changes around the pX region and spontaneous HTLV-1 transcription are CTCF-independent"

    Article Title: Epigenetic changes around the pX region and spontaneous HTLV-1 transcription are CTCF-independent

    Journal: Wellcome Open Research

    doi: 10.12688/wellcomeopenres.14741.2

    DNA methylation across the body of the HTLV-1 provirus. ( a ) Upper panel: count of CpG dinucleotides in a window of 350 bp in the HTLV-1 reference genome (L36905). Lower panel: schematic diagram of HTLV-1 provirus indicating the two LTRs and the 9 loci examined by MeDIP. ( b ) DNA methylation on the HTLV-1 provirus in the Tax + and Tax – populations from two HTLV-1-infected T cell clones (Clones TBX4B and 11.65). ( c ) DNA methylation on the HTLV-1 provirus in the CADM1 + Tax + and CADM1 + Tax – populations in PBMCs from two unrelated individuals (Patients TDZ and TED). The asterisk (*) indicates that the PCR failed to amplify.
    Figure Legend Snippet: DNA methylation across the body of the HTLV-1 provirus. ( a ) Upper panel: count of CpG dinucleotides in a window of 350 bp in the HTLV-1 reference genome (L36905). Lower panel: schematic diagram of HTLV-1 provirus indicating the two LTRs and the 9 loci examined by MeDIP. ( b ) DNA methylation on the HTLV-1 provirus in the Tax + and Tax – populations from two HTLV-1-infected T cell clones (Clones TBX4B and 11.65). ( c ) DNA methylation on the HTLV-1 provirus in the CADM1 + Tax + and CADM1 + Tax – populations in PBMCs from two unrelated individuals (Patients TDZ and TED). The asterisk (*) indicates that the PCR failed to amplify.

    Techniques Used: DNA Methylation Assay, Methylated DNA Immunoprecipitation, Infection, Clone Assay, Polymerase Chain Reaction

    Related Articles

    Clone Assay:

    Article Title: Random-sequence genetic oligomer pools display an innate potential for ligation and recombination
    Article Snippet: To prepare C120 /C320 -N20 > p pre-ligation for sequencing, the > p group was removed by dephosphorylation using T4-Polynucleotide Kinase (NEB) followed by adapter ligation using miRNA Cloning Linker 1 (IDT) as described in the previous section. cDNA was generated by RT-PCR using primers C1_fw (or C3_fw) and miRNA_CL1_rev in a SuperScript III One-Step RT-PCR reaction supplemented with Platinum Taq DNA Polymerase (ThermoFisher Scientific) according to the supplier’s instructions. .. Libraries for Illumina deep sequencing were prepared by PCR using the NEBNext High-Fidelity 2X PCR Master Mix (NEB) using the primers P3_miRNA_CL1_rv and P5_C1 (or P5_C3) according to the supplier’s protocol.

    Article Title: Systematic interrogation of β-catenin co-dependent genes facilitated by integrated genetic and proteomic approaches
    Article Snippet: Vectors containing an sgRNA in position 1 were pooled and digested with BsmBI (Thermo Scientific) and a pool of the same 115 sgRNAs was ligated into the BsmBI cloning sites (exactly as described above for single sgRNA pooled library). .. Following puromycin selection (2 μg/ml), genomic DNA (5μg) extracted at 3 or 28 DPI was PCR amplified using NEBNext® High-Fidelity 2X PCR Master Mix (New England Biolabs) and F/RPCR1_2sg primers ( ).

    Centrifugation:

    Article Title: TRIM28-Regulated Transposon Repression Is Required for Human Germline Competency and Not Primed or Naive Human Pluripotency
    Article Snippet: .. After centrifugation, the cell pellet was resuspended in transposase buffer, which contained the Tn5 transposase enzyme and tagmentation buffer (Nextera DNA library prep kit: Illumina, catalog no. 15028212), and incubated at 37°C for 30 min. After purification by a MinElute PCR Purification Kit 250 (Qiagen 28006), P1 barcode (25 μM) and appropriate P2 barcodes (25 μM) were added to the DNA and run for five cycles of PCR reaction (NEBNext High-Fidelity 2× PCR Master Mix; NEB, M0541S). .. Ten percent of the PCR product was taken out for real-time PCR analysis to determine the amplification cycles of the library, followed immediately by PCR amplification of the remaining library DNA.

    Amplification:

    Article Title: Microbiological Contamination at Workplaces in a Combined Heat and Power (CHP) Station Processing Plant Biomass
    Article Snippet: .. PCR reactions (both for amplification of 16S rRNA gene fragments and ITS1 regions) were prepared in 50 μL volume containing 5 μL of DNA as template, 25 μL NEBNext® Hot Start High-Fidelity 2× PCR Master Mix (New England BioLabs, Ipswich, MA, USA) and 10 pmol of 341F and 785R (16S rRNA gene) or IT1F12 and ITS2 (ITS1 region) primers. .. Amplification of bacterial and fungal fragments was performed under the same conditions: an initial denaturation at 98 °C for 30 s was followed by 15 cycles of denaturation at 98 °C for 10 s, annealing at 52 °C for 75 s, extension at 65 °C for 75 s, and a final extension at 65 °C for 5 min. DNA libraries were constructed using Nextera Index kit following the manufacturer’s library preparation protocol for short amplicons (2 × 250 bp).

    Article Title: Cooperative interactions enable singular olfactory receptor expression in mouse olfactory neurons
    Article Snippet: .. The transposed DNA was then amplified using barcoded primers and NEBNext High Fidelity 2x PCR Master Mix (NEB). .. Amplified libraries were purified using Ampure XP beads (Beckman Coulter) at a ratio of 1.6 ul of beads per 1 uL of library and eluted in 30 uL of elution buffer (10 mM Tris-HCl pH 8, 0.1 mM EDTA).

    Article Title: A global regulatory system links virulence and antibiotic resistance to envelope homeostasis in Acinetobacter baumannii
    Article Snippet: .. Genomic library preparation for Illumina sequencing was performed using the small-volume Nextera tagmentation method [ ], with the following modifications: NEBNext High-Fidelity 2X PCR Master Mix (NEB) was used for amplification; adapter addition/library amplification was followed by 5 cycles of reconditioning PCR with adapter-specific primers; library QC/quantitation was performed directly after PCR by visualizing a sample of each library separated on a 2% agarose/TAE gel; multiplexed PCRs were purified by using a single Qiagen PCR purification column and size-selected via a PippinHT. ..

    Article Title: PCR amplification of a triple-repeat genetic target directly from whole blood in 15 minutes as a proof-of-principle PCR study for direct sample analysis for a clinically relevant target
    Article Snippet: Conversely, many high-fidelity polymerases that successfully amplified target from pure DNA samples, using the Philisa 15-minute cycling protocol, did not amplify target when whole blood was added directly to the master mix. .. For the purposes of our study, the NEBNext High-Fidelity 2X PCR Master Mix was used because it produced PCR product successfully for both rapid cycling and direct-blood analysis when paired with the Philisa Thermal Cycler.

    Article Title: Transcriptomic but not genomic variability confers phenotype of breast cancer stem cells
    Article Snippet: .. G. PCR amplification: PCR enrichment was conducted by using 22 μL Adapter-ligated DNA, 25 μL of 2 × NEB Next high-fidelity PCR master buffer (NEB, Cat. M0541L), 1.5 μL of 10 μmol/L MUP primer, 1.5 μL of 10 μmol/L barcode primer. ..

    Article Title: Chromatinized Protein Kinase C-θ Directly Regulates Inducible Genes in Epithelial to Mesenchymal Transition and Breast Cancer Stem Cells
    Article Snippet: AMPure beads were used to isolate library fragments ranging between 175 and 225 bp in size, and the purified DNA was combined with the NEBNext High-Fidelity 2X PCR master mix, 25 μM Universal PCR primer, and 25 μM index primer for multiplexing purposes (NEBNext multiplex oligonucleotides for Illumina, NEB number E7335L; New England BioLabs Inc.). .. A total of 15 cycles were used for PCR amplification of the adaptor-ligated DNA.

    Article Title: Systematic interrogation of β-catenin co-dependent genes facilitated by integrated genetic and proteomic approaches
    Article Snippet: .. Following puromycin selection (2 μg/ml), genomic DNA (5μg) extracted at 3 or 28 DPI was PCR amplified using NEBNext® High-Fidelity 2X PCR Master Mix (New England Biolabs) and F/RPCR1_2sg primers ( ). .. This mixture (5μl) was used for a second PCR amplification using barcoded-staggered primers ( ).

    Article Title: TRIM28-Regulated Transposon Repression Is Required for Human Germline Competency and Not Primed or Naive Human Pluripotency
    Article Snippet: After centrifugation, the cell pellet was resuspended in transposase buffer, which contained the Tn5 transposase enzyme and tagmentation buffer (Nextera DNA library prep kit: Illumina, catalog no. 15028212), and incubated at 37°C for 30 min. After purification by a MinElute PCR Purification Kit 250 (Qiagen 28006), P1 barcode (25 μM) and appropriate P2 barcodes (25 μM) were added to the DNA and run for five cycles of PCR reaction (NEBNext High-Fidelity 2× PCR Master Mix; NEB, M0541S). .. Ten percent of the PCR product was taken out for real-time PCR analysis to determine the amplification cycles of the library, followed immediately by PCR amplification of the remaining library DNA.

    Article Title: Smokeless tobacco impacts oral microbiota in a Syrian Golden hamster cheek pouch carcinogenesis model
    Article Snippet: Each PCR reaction contained 1 μl 5 μM specified 16 s F primer , 1 μl 5 μM specified 16 s R primer , 1 μl gDNA, 10 μl NEBNext High-Fidelity 2X PCR Master Mix (New England BioLabs Inc., Ipswich, MA), and 7 μl deionized H2 O. .. Illumina-specific barcoded primers ( ) (Integrated DNA Technologies, Coralville, IA) were then added in a second PCR reaction that consisted of 15 μl NEBNext High-Fidelity 2X Master Mix, 5 μl primers, and 10 μl DNA from the first amplification.

    Article Title: Guidance of regulatory T cell development by Satb1-dependent super-enhancer establishment
    Article Snippet: .. Purified DNA was amplified using NEBNext High-Fidelity PCR Master Mix (New England BioLabs) with the following primers: forward 5′-TCGTCGGCAGCGTCAGATGTG-3′ and reverse 5′-GTCTCGTGGGCTCGGAGATGT-3′. .. PCR fragments were size-selected (150–500 bp) with Agencourt AMPure XP (Beckman Coulter), sonicated by Covaris Focused-ultrasonicator S220, subjected to library preparation with KAPA Library Preparation Kit, and sequenced by Ion Proton.

    Synthesized:

    Article Title: Transcriptome-wide identification of novel circular RNAs in soybean in response to low-phosphorus stress
    Article Snippet: Second-strand cDNA was subsequently synthesized by DNA polymerase I, second-strand synthesis reaction buffer, dUTP, and RNase H. The double-stranded cDNA was then purified and collected via AMPure XP beads for the subsequent steps. .. Quantitative polymerase chain reaction (qPCR) was subsequently conducted in conjunction with universal PCR primers, NEBNext High-Fidelity PCR Master Mix, and Index (X) primer.

    Neutralization:

    Article Title: Efficient Genome-Wide Sequencing and Low-Coverage Pedigree Analysis from Noninvasively Collected Samples
    Article Snippet: The beads were then washed three times with 500 μl high-stringency wash buffer (0.1× SSC, 0.1% SDS) with a 10-min room temperature incubation between each wash. After the final wash, the enriched fDNA fraction was eluted from the beads with 50 μl elution buffer (0.1 M NaOH), transferred to a new tube containing 70 μl “neutralization buffer” (1 M Tris-HCl, pH 7.5), purified with 1.8× AMPure beads, and eluted in a 30-μl volume. .. A final PCR was carried out in a 50-μl reaction volume, using 23 μl of the posthybridization fDNA and either (1) 25 μl 2× KAPA High Fidelity master mix and 2 μl TruSeq universal primer (capture 1) or (2) 25 μl 2× NEBNext High Fidelity PCR master mix, 1 μl universal PCR primer, and 1 μl NEB indexing primer (capture 2).

    Construct:

    Article Title: Microbiological Contamination at Workplaces in a Combined Heat and Power (CHP) Station Processing Plant Biomass
    Article Snippet: PCR reactions (both for amplification of 16S rRNA gene fragments and ITS1 regions) were prepared in 50 μL volume containing 5 μL of DNA as template, 25 μL NEBNext® Hot Start High-Fidelity 2× PCR Master Mix (New England BioLabs, Ipswich, MA, USA) and 10 pmol of 341F and 785R (16S rRNA gene) or IT1F12 and ITS2 (ITS1 region) primers. .. Amplification of bacterial and fungal fragments was performed under the same conditions: an initial denaturation at 98 °C for 30 s was followed by 15 cycles of denaturation at 98 °C for 10 s, annealing at 52 °C for 75 s, extension at 65 °C for 75 s, and a final extension at 65 °C for 5 min. DNA libraries were constructed using Nextera Index kit following the manufacturer’s library preparation protocol for short amplicons (2 × 250 bp).

    Article Title: Transcriptome-wide identification of novel circular RNAs in soybean in response to low-phosphorus stress
    Article Snippet: Quantitative polymerase chain reaction (qPCR) was subsequently conducted in conjunction with universal PCR primers, NEBNext High-Fidelity PCR Master Mix, and Index (X) primer. .. The qualified libraries were then constructed and sequenced on an Illumina HiSeq 4000 platform, and long paired-end reads (150 bp) were generated.

    Electrophoresis:

    Article Title: Transcriptomic but not genomic variability confers phenotype of breast cancer stem cells
    Article Snippet: G. PCR amplification: PCR enrichment was conducted by using 22 μL Adapter-ligated DNA, 25 μL of 2 × NEB Next high-fidelity PCR master buffer (NEB, Cat. M0541L), 1.5 μL of 10 μmol/L MUP primer, 1.5 μL of 10 μmol/L barcode primer. .. H. Extraction and purification of the final library: electrophoresis was conducted in agarose gel with the PCR products from the last step and then extracting and purifying DNA from agarose gel using the gel extraction kit (TIANGEN, Cat. DP214-03).

    Incubation:

    Article Title: Efficient Genome-Wide Sequencing and Low-Coverage Pedigree Analysis from Noninvasively Collected Samples
    Article Snippet: The beads were then washed three times with 500 μl high-stringency wash buffer (0.1× SSC, 0.1% SDS) with a 10-min room temperature incubation between each wash. After the final wash, the enriched fDNA fraction was eluted from the beads with 50 μl elution buffer (0.1 M NaOH), transferred to a new tube containing 70 μl “neutralization buffer” (1 M Tris-HCl, pH 7.5), purified with 1.8× AMPure beads, and eluted in a 30-μl volume. .. A final PCR was carried out in a 50-μl reaction volume, using 23 μl of the posthybridization fDNA and either (1) 25 μl 2× KAPA High Fidelity master mix and 2 μl TruSeq universal primer (capture 1) or (2) 25 μl 2× NEBNext High Fidelity PCR master mix, 1 μl universal PCR primer, and 1 μl NEB indexing primer (capture 2).

    Article Title: Transcriptomic but not genomic variability confers phenotype of breast cancer stem cells
    Article Snippet: Beads were washed twice with 200 μL of 80% ethanol for 30 s at room temperature and dried at room temperature for 5–10 min. Then, 25 μL Buffer EB was added to the beads, mixed up and down for ten times, incubated for 2 min at room temperature, and put on magnetic stand at room temperature for about 5 min. After that, 22 μL of supernatant was transferred to a new PCR tube. .. G. PCR amplification: PCR enrichment was conducted by using 22 μL Adapter-ligated DNA, 25 μL of 2 × NEB Next high-fidelity PCR master buffer (NEB, Cat. M0541L), 1.5 μL of 10 μmol/L MUP primer, 1.5 μL of 10 μmol/L barcode primer.

    Article Title: TRIM28-Regulated Transposon Repression Is Required for Human Germline Competency and Not Primed or Naive Human Pluripotency
    Article Snippet: .. After centrifugation, the cell pellet was resuspended in transposase buffer, which contained the Tn5 transposase enzyme and tagmentation buffer (Nextera DNA library prep kit: Illumina, catalog no. 15028212), and incubated at 37°C for 30 min. After purification by a MinElute PCR Purification Kit 250 (Qiagen 28006), P1 barcode (25 μM) and appropriate P2 barcodes (25 μM) were added to the DNA and run for five cycles of PCR reaction (NEBNext High-Fidelity 2× PCR Master Mix; NEB, M0541S). .. Ten percent of the PCR product was taken out for real-time PCR analysis to determine the amplification cycles of the library, followed immediately by PCR amplification of the remaining library DNA.

    Gel Extraction:

    Article Title: Transcriptomic but not genomic variability confers phenotype of breast cancer stem cells
    Article Snippet: G. PCR amplification: PCR enrichment was conducted by using 22 μL Adapter-ligated DNA, 25 μL of 2 × NEB Next high-fidelity PCR master buffer (NEB, Cat. M0541L), 1.5 μL of 10 μmol/L MUP primer, 1.5 μL of 10 μmol/L barcode primer. .. H. Extraction and purification of the final library: electrophoresis was conducted in agarose gel with the PCR products from the last step and then extracting and purifying DNA from agarose gel using the gel extraction kit (TIANGEN, Cat. DP214-03).

    Hybridization:

    Article Title: Efficient Genome-Wide Sequencing and Low-Coverage Pedigree Analysis from Noninvasively Collected Samples
    Article Snippet: Next, the entire fDNA/RNA hybridization mix was added to the 200-μl Dynal MyOne Streptavidin T1 bead and binding buffer slurry. .. A final PCR was carried out in a 50-μl reaction volume, using 23 μl of the posthybridization fDNA and either (1) 25 μl 2× KAPA High Fidelity master mix and 2 μl TruSeq universal primer (capture 1) or (2) 25 μl 2× NEBNext High Fidelity PCR master mix, 1 μl universal PCR primer, and 1 μl NEB indexing primer (capture 2).

    Article Title: Transcriptome-wide identification of novel circular RNAs in soybean in response to low-phosphorus stress
    Article Snippet: Afterward, to prepare for hybridization, the 3′ ends of the DNA fragments were repaired, and the hairpin loop structures were ligated to the fragments. .. Quantitative polymerase chain reaction (qPCR) was subsequently conducted in conjunction with universal PCR primers, NEBNext High-Fidelity PCR Master Mix, and Index (X) primer.

    Ligation:

    Article Title: Transcriptomic but not genomic variability confers phenotype of breast cancer stem cells
    Article Snippet: E. Adapter ligation: The mixture of 25 μL A-tailed DNA, 1 μL of 50 μmol/L multiplexing adapter, 3 μL of 10× T4 DNA ligase buffer (NEB, Cat. B0202), and 1 μL of 400 units/μL T4 DNA ligase (NEB, Cat. M0202L) was used in adapter ligation step followed by incubation at 16 °C overnight. .. G. PCR amplification: PCR enrichment was conducted by using 22 μL Adapter-ligated DNA, 25 μL of 2 × NEB Next high-fidelity PCR master buffer (NEB, Cat. M0541L), 1.5 μL of 10 μmol/L MUP primer, 1.5 μL of 10 μmol/L barcode primer.

    Article Title: Chromatinized Protein Kinase C-θ Directly Regulates Inducible Genes in Epithelial to Mesenchymal Transition and Breast Cancer Stem Cells
    Article Snippet: Subsequent adaptor ligation of the dA-tailed DNA was performed with 1.5 μM NEBNext adaptor primers, Quick Ligation reaction buffer, and Quick T4 DNA ligase. .. AMPure beads were used to isolate library fragments ranging between 175 and 225 bp in size, and the purified DNA was combined with the NEBNext High-Fidelity 2X PCR master mix, 25 μM Universal PCR primer, and 25 μM index primer for multiplexing purposes (NEBNext multiplex oligonucleotides for Illumina, NEB number E7335L; New England BioLabs Inc.).

    Article Title: Random-sequence genetic oligomer pools display an innate potential for ligation and recombination
    Article Snippet: To prepare C120 /C320 -N20 > p pre-ligation for sequencing, the > p group was removed by dephosphorylation using T4-Polynucleotide Kinase (NEB) followed by adapter ligation using miRNA Cloning Linker 1 (IDT) as described in the previous section. cDNA was generated by RT-PCR using primers C1_fw (or C3_fw) and miRNA_CL1_rev in a SuperScript III One-Step RT-PCR reaction supplemented with Platinum Taq DNA Polymerase (ThermoFisher Scientific) according to the supplier’s instructions. .. Libraries for Illumina deep sequencing were prepared by PCR using the NEBNext High-Fidelity 2X PCR Master Mix (NEB) using the primers P3_miRNA_CL1_rv and P5_C1 (or P5_C3) according to the supplier’s protocol.

    Article Title: Systematic interrogation of β-catenin co-dependent genes facilitated by integrated genetic and proteomic approaches
    Article Snippet: Following ligation the library was electroporated into Stbl4 cells (Life Technologies) grown at 30°C. .. Following puromycin selection (2 μg/ml), genomic DNA (5μg) extracted at 3 or 28 DPI was PCR amplified using NEBNext® High-Fidelity 2X PCR Master Mix (New England Biolabs) and F/RPCR1_2sg primers ( ).

    Genomic Sequencing:

    Article Title: Smokeless tobacco impacts oral microbiota in a Syrian Golden hamster cheek pouch carcinogenesis model
    Article Snippet: 16S rRNA library preparation and sequencing were conducted at the Genomic Sequencing and Analysis Facility of the University of Texas at Austin. .. Each PCR reaction contained 1 μl 5 μM specified 16 s F primer , 1 μl 5 μM specified 16 s R primer , 1 μl gDNA, 10 μl NEBNext High-Fidelity 2X PCR Master Mix (New England BioLabs Inc., Ipswich, MA), and 7 μl deionized H2 O.

    Reverse Transcription Polymerase Chain Reaction:

    Article Title: Random-sequence genetic oligomer pools display an innate potential for ligation and recombination
    Article Snippet: To prepare C120 /C320 -N20 > p pre-ligation for sequencing, the > p group was removed by dephosphorylation using T4-Polynucleotide Kinase (NEB) followed by adapter ligation using miRNA Cloning Linker 1 (IDT) as described in the previous section. cDNA was generated by RT-PCR using primers C1_fw (or C3_fw) and miRNA_CL1_rev in a SuperScript III One-Step RT-PCR reaction supplemented with Platinum Taq DNA Polymerase (ThermoFisher Scientific) according to the supplier’s instructions. .. Libraries for Illumina deep sequencing were prepared by PCR using the NEBNext High-Fidelity 2X PCR Master Mix (NEB) using the primers P3_miRNA_CL1_rv and P5_C1 (or P5_C3) according to the supplier’s protocol.

    Generated:

    Article Title: Transcriptomic but not genomic variability confers phenotype of breast cancer stem cells
    Article Snippet: G. PCR amplification: PCR enrichment was conducted by using 22 μL Adapter-ligated DNA, 25 μL of 2 × NEB Next high-fidelity PCR master buffer (NEB, Cat. M0541L), 1.5 μL of 10 μmol/L MUP primer, 1.5 μL of 10 μmol/L barcode primer. .. Quality control and sequencing of the final library: The quality and concentration of DNA fragments in the DNA libraries generated were assessed using High-Sensitivity Bioanalyzer.

    Article Title: Transcriptome-wide identification of novel circular RNAs in soybean in response to low-phosphorus stress
    Article Snippet: Quantitative polymerase chain reaction (qPCR) was subsequently conducted in conjunction with universal PCR primers, NEBNext High-Fidelity PCR Master Mix, and Index (X) primer. .. The qualified libraries were then constructed and sequenced on an Illumina HiSeq 4000 platform, and long paired-end reads (150 bp) were generated.

    Article Title: Random-sequence genetic oligomer pools display an innate potential for ligation and recombination
    Article Snippet: To prepare C120 /C320 -N20 > p pre-ligation for sequencing, the > p group was removed by dephosphorylation using T4-Polynucleotide Kinase (NEB) followed by adapter ligation using miRNA Cloning Linker 1 (IDT) as described in the previous section. cDNA was generated by RT-PCR using primers C1_fw (or C3_fw) and miRNA_CL1_rev in a SuperScript III One-Step RT-PCR reaction supplemented with Platinum Taq DNA Polymerase (ThermoFisher Scientific) according to the supplier’s instructions. .. Libraries for Illumina deep sequencing were prepared by PCR using the NEBNext High-Fidelity 2X PCR Master Mix (NEB) using the primers P3_miRNA_CL1_rv and P5_C1 (or P5_C3) according to the supplier’s protocol.

    Sequencing:

    Article Title: Microbiological Contamination at Workplaces in a Combined Heat and Power (CHP) Station Processing Plant Biomass
    Article Snippet: Paragraph title: PCR Amplification and Sequencing ... PCR reactions (both for amplification of 16S rRNA gene fragments and ITS1 regions) were prepared in 50 μL volume containing 5 μL of DNA as template, 25 μL NEBNext® Hot Start High-Fidelity 2× PCR Master Mix (New England BioLabs, Ipswich, MA, USA) and 10 pmol of 341F and 785R (16S rRNA gene) or IT1F12 and ITS2 (ITS1 region) primers.

    Article Title: A global regulatory system links virulence and antibiotic resistance to envelope homeostasis in Acinetobacter baumannii
    Article Snippet: .. Genomic library preparation for Illumina sequencing was performed using the small-volume Nextera tagmentation method [ ], with the following modifications: NEBNext High-Fidelity 2X PCR Master Mix (NEB) was used for amplification; adapter addition/library amplification was followed by 5 cycles of reconditioning PCR with adapter-specific primers; library QC/quantitation was performed directly after PCR by visualizing a sample of each library separated on a 2% agarose/TAE gel; multiplexed PCRs were purified by using a single Qiagen PCR purification column and size-selected via a PippinHT. ..

    Article Title: Transcriptomic but not genomic variability confers phenotype of breast cancer stem cells
    Article Snippet: Paragraph title: Library preparation and sequencing for amplicons ... G. PCR amplification: PCR enrichment was conducted by using 22 μL Adapter-ligated DNA, 25 μL of 2 × NEB Next high-fidelity PCR master buffer (NEB, Cat. M0541L), 1.5 μL of 10 μmol/L MUP primer, 1.5 μL of 10 μmol/L barcode primer.

    Article Title: Transcriptome-wide identification of novel circular RNAs in soybean in response to low-phosphorus stress
    Article Snippet: Paragraph title: Library construction and Illumina sequencing ... Quantitative polymerase chain reaction (qPCR) was subsequently conducted in conjunction with universal PCR primers, NEBNext High-Fidelity PCR Master Mix, and Index (X) primer.

    Article Title: Random-sequence genetic oligomer pools display an innate potential for ligation and recombination
    Article Snippet: .. Libraries for Illumina deep sequencing were prepared by PCR using the NEBNext High-Fidelity 2X PCR Master Mix (NEB) using the primers P3_miRNA_CL1_rv and P5_C1 (or P5_C3) according to the supplier’s protocol. .. DNA libraries from the 5’OH-N20 -C220 and 5’OH-N20 -C420 pre-ligation semi-random RNA pools were generated using a two-step protocol: First, single stranded cDNA was prepared using C2_rv/C4_rv using the SuperScript III according to the supplier’s instructions.

    Article Title: Systematic interrogation of β-catenin co-dependent genes facilitated by integrated genetic and proteomic approaches
    Article Snippet: To generate a pooled double sgRNA library, 115 sgRNAs were individually PCR amplified together with the S. Pyogenes tracer sequence and inserted into position 1 (AgeI/EcoRI restriction sites). .. Following puromycin selection (2 μg/ml), genomic DNA (5μg) extracted at 3 or 28 DPI was PCR amplified using NEBNext® High-Fidelity 2X PCR Master Mix (New England Biolabs) and F/RPCR1_2sg primers ( ).

    Article Title: TRIM28-Regulated Transposon Repression Is Required for Human Germline Competency and Not Primed or Naive Human Pluripotency
    Article Snippet: After centrifugation, the cell pellet was resuspended in transposase buffer, which contained the Tn5 transposase enzyme and tagmentation buffer (Nextera DNA library prep kit: Illumina, catalog no. 15028212), and incubated at 37°C for 30 min. After purification by a MinElute PCR Purification Kit 250 (Qiagen 28006), P1 barcode (25 μM) and appropriate P2 barcodes (25 μM) were added to the DNA and run for five cycles of PCR reaction (NEBNext High-Fidelity 2× PCR Master Mix; NEB, M0541S). .. Libraries were subjected to paired-end 50-bp sequencing on a Hi Seq 4000 sequencer with 4–6 indexed libraries per lane.

    Article Title: Smokeless tobacco impacts oral microbiota in a Syrian Golden hamster cheek pouch carcinogenesis model
    Article Snippet: Paragraph title: 2.3. 16S rRNA library preparation and sequencing ... Each PCR reaction contained 1 μl 5 μM specified 16 s F primer , 1 μl 5 μM specified 16 s R primer , 1 μl gDNA, 10 μl NEBNext High-Fidelity 2X PCR Master Mix (New England BioLabs Inc., Ipswich, MA), and 7 μl deionized H2 O.

    Article Title: Guidance of regulatory T cell development by Satb1-dependent super-enhancer establishment
    Article Snippet: Native chromatin transposed with sequencing adaptors using Nextera DNA Library Prep Kit (Illumina) was purified using a MinElute PCR Purification Kit (Qiagen). .. Purified DNA was amplified using NEBNext High-Fidelity PCR Master Mix (New England BioLabs) with the following primers: forward 5′-TCGTCGGCAGCGTCAGATGTG-3′ and reverse 5′-GTCTCGTGGGCTCGGAGATGT-3′.

    Sonication:

    Article Title: Guidance of regulatory T cell development by Satb1-dependent super-enhancer establishment
    Article Snippet: Purified DNA was amplified using NEBNext High-Fidelity PCR Master Mix (New England BioLabs) with the following primers: forward 5′-TCGTCGGCAGCGTCAGATGTG-3′ and reverse 5′-GTCTCGTGGGCTCGGAGATGT-3′. .. PCR fragments were size-selected (150–500 bp) with Agencourt AMPure XP (Beckman Coulter), sonicated by Covaris Focused-ultrasonicator S220, subjected to library preparation with KAPA Library Preparation Kit, and sequenced by Ion Proton.

    Binding Assay:

    Article Title: Efficient Genome-Wide Sequencing and Low-Coverage Pedigree Analysis from Noninvasively Collected Samples
    Article Snippet: Next, the entire fDNA/RNA hybridization mix was added to the 200-μl Dynal MyOne Streptavidin T1 bead and binding buffer slurry. .. A final PCR was carried out in a 50-μl reaction volume, using 23 μl of the posthybridization fDNA and either (1) 25 μl 2× KAPA High Fidelity master mix and 2 μl TruSeq universal primer (capture 1) or (2) 25 μl 2× NEBNext High Fidelity PCR master mix, 1 μl universal PCR primer, and 1 μl NEB indexing primer (capture 2).

    Article Title: Random-sequence genetic oligomer pools display an innate potential for ligation and recombination
    Article Snippet: In each reaction, 1.25 pM of total RNA from ligation or control reactions was used as RT-template with forward and reverse primers complementary to the respective constant C120 /C320 and C220 /C420 primer binding sites ( ). .. Libraries for Illumina deep sequencing were prepared by PCR using the NEBNext High-Fidelity 2X PCR Master Mix (NEB) using the primers P3_miRNA_CL1_rv and P5_C1 (or P5_C3) according to the supplier’s protocol.

    ChIP-sequencing:

    Article Title: Chromatinized Protein Kinase C-θ Directly Regulates Inducible Genes in Epithelial to Mesenchymal Transition and Breast Cancer Stem Cells
    Article Snippet: Paragraph title: ChIP-seq. ... AMPure beads were used to isolate library fragments ranging between 175 and 225 bp in size, and the purified DNA was combined with the NEBNext High-Fidelity 2X PCR master mix, 25 μM Universal PCR primer, and 25 μM index primer for multiplexing purposes (NEBNext multiplex oligonucleotides for Illumina, NEB number E7335L; New England BioLabs Inc.).

    Multiplexing:

    Article Title: Transcriptomic but not genomic variability confers phenotype of breast cancer stem cells
    Article Snippet: E. Adapter ligation: The mixture of 25 μL A-tailed DNA, 1 μL of 50 μmol/L multiplexing adapter, 3 μL of 10× T4 DNA ligase buffer (NEB, Cat. B0202), and 1 μL of 400 units/μL T4 DNA ligase (NEB, Cat. M0202L) was used in adapter ligation step followed by incubation at 16 °C overnight. .. G. PCR amplification: PCR enrichment was conducted by using 22 μL Adapter-ligated DNA, 25 μL of 2 × NEB Next high-fidelity PCR master buffer (NEB, Cat. M0541L), 1.5 μL of 10 μmol/L MUP primer, 1.5 μL of 10 μmol/L barcode primer.

    Article Title: Chromatinized Protein Kinase C-θ Directly Regulates Inducible Genes in Epithelial to Mesenchymal Transition and Breast Cancer Stem Cells
    Article Snippet: .. AMPure beads were used to isolate library fragments ranging between 175 and 225 bp in size, and the purified DNA was combined with the NEBNext High-Fidelity 2X PCR master mix, 25 μM Universal PCR primer, and 25 μM index primer for multiplexing purposes (NEBNext multiplex oligonucleotides for Illumina, NEB number E7335L; New England BioLabs Inc.). .. A total of 15 cycles were used for PCR amplification of the adaptor-ligated DNA.

    Functional Assay:

    Article Title: A global regulatory system links virulence and antibiotic resistance to envelope homeostasis in Acinetobacter baumannii
    Article Snippet: Genomic library preparation for Illumina sequencing was performed using the small-volume Nextera tagmentation method [ ], with the following modifications: NEBNext High-Fidelity 2X PCR Master Mix (NEB) was used for amplification; adapter addition/library amplification was followed by 5 cycles of reconditioning PCR with adapter-specific primers; library QC/quantitation was performed directly after PCR by visualizing a sample of each library separated on a 2% agarose/TAE gel; multiplexed PCRs were purified by using a single Qiagen PCR purification column and size-selected via a PippinHT. .. The predicted functional impact of substitution variants was determined by using PROVEAN [ ].

    Purification:

    Article Title: Efficient Genome-Wide Sequencing and Low-Coverage Pedigree Analysis from Noninvasively Collected Samples
    Article Snippet: The beads were then washed three times with 500 μl high-stringency wash buffer (0.1× SSC, 0.1% SDS) with a 10-min room temperature incubation between each wash. After the final wash, the enriched fDNA fraction was eluted from the beads with 50 μl elution buffer (0.1 M NaOH), transferred to a new tube containing 70 μl “neutralization buffer” (1 M Tris-HCl, pH 7.5), purified with 1.8× AMPure beads, and eluted in a 30-μl volume. .. A final PCR was carried out in a 50-μl reaction volume, using 23 μl of the posthybridization fDNA and either (1) 25 μl 2× KAPA High Fidelity master mix and 2 μl TruSeq universal primer (capture 1) or (2) 25 μl 2× NEBNext High Fidelity PCR master mix, 1 μl universal PCR primer, and 1 μl NEB indexing primer (capture 2).

    Article Title: Cooperative interactions enable singular olfactory receptor expression in mouse olfactory neurons
    Article Snippet: Transposed DNA was column purified using a Qiagen MinElute PCR cleanup kit (Qiagen). .. The transposed DNA was then amplified using barcoded primers and NEBNext High Fidelity 2x PCR Master Mix (NEB).

    Article Title: A global regulatory system links virulence and antibiotic resistance to envelope homeostasis in Acinetobacter baumannii
    Article Snippet: .. Genomic library preparation for Illumina sequencing was performed using the small-volume Nextera tagmentation method [ ], with the following modifications: NEBNext High-Fidelity 2X PCR Master Mix (NEB) was used for amplification; adapter addition/library amplification was followed by 5 cycles of reconditioning PCR with adapter-specific primers; library QC/quantitation was performed directly after PCR by visualizing a sample of each library separated on a 2% agarose/TAE gel; multiplexed PCRs were purified by using a single Qiagen PCR purification column and size-selected via a PippinHT. ..

    Article Title: Transcriptomic but not genomic variability confers phenotype of breast cancer stem cells
    Article Snippet: D. Column purification: The PCR products were purified with Universal DNA Purification Kit (TIANGEN, Cat. DP214-03). .. G. PCR amplification: PCR enrichment was conducted by using 22 μL Adapter-ligated DNA, 25 μL of 2 × NEB Next high-fidelity PCR master buffer (NEB, Cat. M0541L), 1.5 μL of 10 μmol/L MUP primer, 1.5 μL of 10 μmol/L barcode primer.

    Article Title: Chromatinized Protein Kinase C-θ Directly Regulates Inducible Genes in Epithelial to Mesenchymal Transition and Breast Cancer Stem Cells
    Article Snippet: .. AMPure beads were used to isolate library fragments ranging between 175 and 225 bp in size, and the purified DNA was combined with the NEBNext High-Fidelity 2X PCR master mix, 25 μM Universal PCR primer, and 25 μM index primer for multiplexing purposes (NEBNext multiplex oligonucleotides for Illumina, NEB number E7335L; New England BioLabs Inc.). .. A total of 15 cycles were used for PCR amplification of the adaptor-ligated DNA.

    Article Title: Transcriptome-wide identification of novel circular RNAs in soybean in response to low-phosphorus stress
    Article Snippet: To select cDNA fragments that were 200 to 500 bp in length, the fragments in each of the libraries were purified by an AMPure XP bead system. .. Quantitative polymerase chain reaction (qPCR) was subsequently conducted in conjunction with universal PCR primers, NEBNext High-Fidelity PCR Master Mix, and Index (X) primer.

    Article Title: Random-sequence genetic oligomer pools display an innate potential for ligation and recombination
    Article Snippet: Libraries for Illumina deep sequencing were prepared by PCR using the NEBNext High-Fidelity 2X PCR Master Mix (NEB) using the primers P3_miRNA_CL1_rv and P5_C1 (or P5_C3) according to the supplier’s protocol. .. Next, miRNA Cloning Linker 1 was ligated to the purified cDNA and ds cDNA was generated using GoTaq Green mastermix (Promega).

    Article Title: TRIM28-Regulated Transposon Repression Is Required for Human Germline Competency and Not Primed or Naive Human Pluripotency
    Article Snippet: .. After centrifugation, the cell pellet was resuspended in transposase buffer, which contained the Tn5 transposase enzyme and tagmentation buffer (Nextera DNA library prep kit: Illumina, catalog no. 15028212), and incubated at 37°C for 30 min. After purification by a MinElute PCR Purification Kit 250 (Qiagen 28006), P1 barcode (25 μM) and appropriate P2 barcodes (25 μM) were added to the DNA and run for five cycles of PCR reaction (NEBNext High-Fidelity 2× PCR Master Mix; NEB, M0541S). .. Ten percent of the PCR product was taken out for real-time PCR analysis to determine the amplification cycles of the library, followed immediately by PCR amplification of the remaining library DNA.

    Article Title: Smokeless tobacco impacts oral microbiota in a Syrian Golden hamster cheek pouch carcinogenesis model
    Article Snippet: Each PCR reaction contained 1 μl 5 μM specified 16 s F primer , 1 μl 5 μM specified 16 s R primer , 1 μl gDNA, 10 μl NEBNext High-Fidelity 2X PCR Master Mix (New England BioLabs Inc., Ipswich, MA), and 7 μl deionized H2 O. .. Triplicate PCR reactions were combined and purified using the Agencourt AMPure XP system (Beckman Coulter Life Sciences, Indianapolis, IN).

    Article Title: Guidance of regulatory T cell development by Satb1-dependent super-enhancer establishment
    Article Snippet: .. Purified DNA was amplified using NEBNext High-Fidelity PCR Master Mix (New England BioLabs) with the following primers: forward 5′-TCGTCGGCAGCGTCAGATGTG-3′ and reverse 5′-GTCTCGTGGGCTCGGAGATGT-3′. .. PCR fragments were size-selected (150–500 bp) with Agencourt AMPure XP (Beckman Coulter), sonicated by Covaris Focused-ultrasonicator S220, subjected to library preparation with KAPA Library Preparation Kit, and sequenced by Ion Proton.

    Polymerase Chain Reaction:

    Article Title: Efficient Genome-Wide Sequencing and Low-Coverage Pedigree Analysis from Noninvasively Collected Samples
    Article Snippet: .. A final PCR was carried out in a 50-μl reaction volume, using 23 μl of the posthybridization fDNA and either (1) 25 μl 2× KAPA High Fidelity master mix and 2 μl TruSeq universal primer (capture 1) or (2) 25 μl 2× NEBNext High Fidelity PCR master mix, 1 μl universal PCR primer, and 1 μl NEB indexing primer (capture 2). .. After 12 PCR cycles the final reaction was purified with 1× AMPure beads, eluted in 20 μL H2 O, and visualized on a Bioanalyzer High Sensitivity DNA chip.

    Article Title: Microbiological Contamination at Workplaces in a Combined Heat and Power (CHP) Station Processing Plant Biomass
    Article Snippet: .. PCR reactions (both for amplification of 16S rRNA gene fragments and ITS1 regions) were prepared in 50 μL volume containing 5 μL of DNA as template, 25 μL NEBNext® Hot Start High-Fidelity 2× PCR Master Mix (New England BioLabs, Ipswich, MA, USA) and 10 pmol of 341F and 785R (16S rRNA gene) or IT1F12 and ITS2 (ITS1 region) primers. .. Amplification of bacterial and fungal fragments was performed under the same conditions: an initial denaturation at 98 °C for 30 s was followed by 15 cycles of denaturation at 98 °C for 10 s, annealing at 52 °C for 75 s, extension at 65 °C for 75 s, and a final extension at 65 °C for 5 min. DNA libraries were constructed using Nextera Index kit following the manufacturer’s library preparation protocol for short amplicons (2 × 250 bp).

    Article Title: Cooperative interactions enable singular olfactory receptor expression in mouse olfactory neurons
    Article Snippet: .. The transposed DNA was then amplified using barcoded primers and NEBNext High Fidelity 2x PCR Master Mix (NEB). .. Amplified libraries were purified using Ampure XP beads (Beckman Coulter) at a ratio of 1.6 ul of beads per 1 uL of library and eluted in 30 uL of elution buffer (10 mM Tris-HCl pH 8, 0.1 mM EDTA).

    Article Title: A global regulatory system links virulence and antibiotic resistance to envelope homeostasis in Acinetobacter baumannii
    Article Snippet: .. Genomic library preparation for Illumina sequencing was performed using the small-volume Nextera tagmentation method [ ], with the following modifications: NEBNext High-Fidelity 2X PCR Master Mix (NEB) was used for amplification; adapter addition/library amplification was followed by 5 cycles of reconditioning PCR with adapter-specific primers; library QC/quantitation was performed directly after PCR by visualizing a sample of each library separated on a 2% agarose/TAE gel; multiplexed PCRs were purified by using a single Qiagen PCR purification column and size-selected via a PippinHT. ..

    Article Title: PCR amplification of a triple-repeat genetic target directly from whole blood in 15 minutes as a proof-of-principle PCR study for direct sample analysis for a clinically relevant target
    Article Snippet: .. For the purposes of our study, the NEBNext High-Fidelity 2X PCR Master Mix was used because it produced PCR product successfully for both rapid cycling and direct-blood analysis when paired with the Philisa Thermal Cycler. .. Data from Figure indicate the assay could tolerate up to 30% whole blood in a given PCR reaction tube and the cycling time was completed in 15 minutes, including a 3-minute hot-start/cell lysis step.

    Article Title: Clinical impact of measurable residual disease monitoring by ultradeep next generation sequencing in NPM1 mutated acute myeloid leukemia
    Article Snippet: .. Assay setup was as follows: For a total reaction volume of 75 μl, 37.5 μl of NEBNext High-Fidelity 2X PCR Master Mix (New England BioLabs Inc., Massachusetts, USA), and 10 uM each forward and reverse primers were used to amplify 600ng of genomic DNA (approximately 1,00,000 genomic cell equivalents). .. PCR cycling conditions were as follows: Initial denaturation of 95°C for 15 minutes; then 35 cycles of denaturation at 94°C for 1 minute, annealing at 65°C for 1 minute, and extension at 72°C for 1 minute; followed by extension cycle of 72°C for 45 seconds.

    Article Title: A rapid and robust method for single cell chromatin accessibility profiling
    Article Snippet: .. Exceptions are the Tn5 transposase, which can be purchased from Illumina (Cat No. FC-121-1030), and the PCR master mix, which can be purchased from various vendors (we used the 2X NEBNext® High-Fidelity 2X PCR Master Mix from NEB). ..

    Article Title: Transcriptomic but not genomic variability confers phenotype of breast cancer stem cells
    Article Snippet: .. G. PCR amplification: PCR enrichment was conducted by using 22 μL Adapter-ligated DNA, 25 μL of 2 × NEB Next high-fidelity PCR master buffer (NEB, Cat. M0541L), 1.5 μL of 10 μmol/L MUP primer, 1.5 μL of 10 μmol/L barcode primer. ..

    Article Title: Chromatinized Protein Kinase C-θ Directly Regulates Inducible Genes in Epithelial to Mesenchymal Transition and Breast Cancer Stem Cells
    Article Snippet: .. AMPure beads were used to isolate library fragments ranging between 175 and 225 bp in size, and the purified DNA was combined with the NEBNext High-Fidelity 2X PCR master mix, 25 μM Universal PCR primer, and 25 μM index primer for multiplexing purposes (NEBNext multiplex oligonucleotides for Illumina, NEB number E7335L; New England BioLabs Inc.). .. A total of 15 cycles were used for PCR amplification of the adaptor-ligated DNA.

    Article Title: Transcriptome-wide identification of novel circular RNAs in soybean in response to low-phosphorus stress
    Article Snippet: .. Quantitative polymerase chain reaction (qPCR) was subsequently conducted in conjunction with universal PCR primers, NEBNext High-Fidelity PCR Master Mix, and Index (X) primer. .. To assess the library quality, an Agilent Bioanalyzer 2100 system was used to purify the PCR products (AMPure XP system).

    Article Title: Random-sequence genetic oligomer pools display an innate potential for ligation and recombination
    Article Snippet: .. Libraries for Illumina deep sequencing were prepared by PCR using the NEBNext High-Fidelity 2X PCR Master Mix (NEB) using the primers P3_miRNA_CL1_rv and P5_C1 (or P5_C3) according to the supplier’s protocol. .. DNA libraries from the 5’OH-N20 -C220 and 5’OH-N20 -C420 pre-ligation semi-random RNA pools were generated using a two-step protocol: First, single stranded cDNA was prepared using C2_rv/C4_rv using the SuperScript III according to the supplier’s instructions.

    Article Title: Systematic interrogation of β-catenin co-dependent genes facilitated by integrated genetic and proteomic approaches
    Article Snippet: .. Following puromycin selection (2 μg/ml), genomic DNA (5μg) extracted at 3 or 28 DPI was PCR amplified using NEBNext® High-Fidelity 2X PCR Master Mix (New England Biolabs) and F/RPCR1_2sg primers ( ). .. This mixture (5μl) was used for a second PCR amplification using barcoded-staggered primers ( ).

    Article Title: TRIM28-Regulated Transposon Repression Is Required for Human Germline Competency and Not Primed or Naive Human Pluripotency
    Article Snippet: .. After centrifugation, the cell pellet was resuspended in transposase buffer, which contained the Tn5 transposase enzyme and tagmentation buffer (Nextera DNA library prep kit: Illumina, catalog no. 15028212), and incubated at 37°C for 30 min. After purification by a MinElute PCR Purification Kit 250 (Qiagen 28006), P1 barcode (25 μM) and appropriate P2 barcodes (25 μM) were added to the DNA and run for five cycles of PCR reaction (NEBNext High-Fidelity 2× PCR Master Mix; NEB, M0541S). .. Ten percent of the PCR product was taken out for real-time PCR analysis to determine the amplification cycles of the library, followed immediately by PCR amplification of the remaining library DNA.

    Article Title: Smokeless tobacco impacts oral microbiota in a Syrian Golden hamster cheek pouch carcinogenesis model
    Article Snippet: .. Each PCR reaction contained 1 μl 5 μM specified 16 s F primer , 1 μl 5 μM specified 16 s R primer , 1 μl gDNA, 10 μl NEBNext High-Fidelity 2X PCR Master Mix (New England BioLabs Inc., Ipswich, MA), and 7 μl deionized H2 O. ..

    Article Title: Guidance of regulatory T cell development by Satb1-dependent super-enhancer establishment
    Article Snippet: .. Purified DNA was amplified using NEBNext High-Fidelity PCR Master Mix (New England BioLabs) with the following primers: forward 5′-TCGTCGGCAGCGTCAGATGTG-3′ and reverse 5′-GTCTCGTGGGCTCGGAGATGT-3′. .. PCR fragments were size-selected (150–500 bp) with Agencourt AMPure XP (Beckman Coulter), sonicated by Covaris Focused-ultrasonicator S220, subjected to library preparation with KAPA Library Preparation Kit, and sequenced by Ion Proton.

    De-Phosphorylation Assay:

    Article Title: Random-sequence genetic oligomer pools display an innate potential for ligation and recombination
    Article Snippet: To prepare C120 /C320 -N20 > p pre-ligation for sequencing, the > p group was removed by dephosphorylation using T4-Polynucleotide Kinase (NEB) followed by adapter ligation using miRNA Cloning Linker 1 (IDT) as described in the previous section. cDNA was generated by RT-PCR using primers C1_fw (or C3_fw) and miRNA_CL1_rev in a SuperScript III One-Step RT-PCR reaction supplemented with Platinum Taq DNA Polymerase (ThermoFisher Scientific) according to the supplier’s instructions. .. Libraries for Illumina deep sequencing were prepared by PCR using the NEBNext High-Fidelity 2X PCR Master Mix (NEB) using the primers P3_miRNA_CL1_rv and P5_C1 (or P5_C3) according to the supplier’s protocol.

    CRISPR:

    Article Title: Systematic interrogation of β-catenin co-dependent genes facilitated by integrated genetic and proteomic approaches
    Article Snippet: Paragraph title: Combinatorial CRISPR-Cas9 loss of function screen ... Following puromycin selection (2 μg/ml), genomic DNA (5μg) extracted at 3 or 28 DPI was PCR amplified using NEBNext® High-Fidelity 2X PCR Master Mix (New England Biolabs) and F/RPCR1_2sg primers ( ).

    IA:

    Article Title: Smokeless tobacco impacts oral microbiota in a Syrian Golden hamster cheek pouch carcinogenesis model
    Article Snippet: Each PCR reaction contained 1 μl 5 μM specified 16 s F primer , 1 μl 5 μM specified 16 s R primer , 1 μl gDNA, 10 μl NEBNext High-Fidelity 2X PCR Master Mix (New England BioLabs Inc., Ipswich, MA), and 7 μl deionized H2 O. .. Illumina-specific barcoded primers ( ) (Integrated DNA Technologies, Coralville, IA) were then added in a second PCR reaction that consisted of 15 μl NEBNext High-Fidelity 2X Master Mix, 5 μl primers, and 10 μl DNA from the first amplification.

    Chromatin Immunoprecipitation:

    Article Title: Efficient Genome-Wide Sequencing and Low-Coverage Pedigree Analysis from Noninvasively Collected Samples
    Article Snippet: A final PCR was carried out in a 50-μl reaction volume, using 23 μl of the posthybridization fDNA and either (1) 25 μl 2× KAPA High Fidelity master mix and 2 μl TruSeq universal primer (capture 1) or (2) 25 μl 2× NEBNext High Fidelity PCR master mix, 1 μl universal PCR primer, and 1 μl NEB indexing primer (capture 2). .. After 12 PCR cycles the final reaction was purified with 1× AMPure beads, eluted in 20 μL H2 O, and visualized on a Bioanalyzer High Sensitivity DNA chip.

    Article Title: Chromatinized Protein Kinase C-θ Directly Regulates Inducible Genes in Epithelial to Mesenchymal Transition and Breast Cancer Stem Cells
    Article Snippet: ChIP-DNA size was selected with AMPure XP beads (Beckman Coulter, Inc., part number ). .. AMPure beads were used to isolate library fragments ranging between 175 and 225 bp in size, and the purified DNA was combined with the NEBNext High-Fidelity 2X PCR master mix, 25 μM Universal PCR primer, and 25 μM index primer for multiplexing purposes (NEBNext multiplex oligonucleotides for Illumina, NEB number E7335L; New England BioLabs Inc.).

    Software:

    Article Title: A global regulatory system links virulence and antibiotic resistance to envelope homeostasis in Acinetobacter baumannii
    Article Snippet: Genomic library preparation for Illumina sequencing was performed using the small-volume Nextera tagmentation method [ ], with the following modifications: NEBNext High-Fidelity 2X PCR Master Mix (NEB) was used for amplification; adapter addition/library amplification was followed by 5 cycles of reconditioning PCR with adapter-specific primers; library QC/quantitation was performed directly after PCR by visualizing a sample of each library separated on a 2% agarose/TAE gel; multiplexed PCRs were purified by using a single Qiagen PCR purification column and size-selected via a PippinHT. .. Reads were aligned to the A . baumannii 17978-mff genome and variants identified by using Geneious software [ ] or BRESEQ [ ].

    Real-time Polymerase Chain Reaction:

    Article Title: Transcriptome-wide identification of novel circular RNAs in soybean in response to low-phosphorus stress
    Article Snippet: .. Quantitative polymerase chain reaction (qPCR) was subsequently conducted in conjunction with universal PCR primers, NEBNext High-Fidelity PCR Master Mix, and Index (X) primer. .. To assess the library quality, an Agilent Bioanalyzer 2100 system was used to purify the PCR products (AMPure XP system).

    Article Title: TRIM28-Regulated Transposon Repression Is Required for Human Germline Competency and Not Primed or Naive Human Pluripotency
    Article Snippet: After centrifugation, the cell pellet was resuspended in transposase buffer, which contained the Tn5 transposase enzyme and tagmentation buffer (Nextera DNA library prep kit: Illumina, catalog no. 15028212), and incubated at 37°C for 30 min. After purification by a MinElute PCR Purification Kit 250 (Qiagen 28006), P1 barcode (25 μM) and appropriate P2 barcodes (25 μM) were added to the DNA and run for five cycles of PCR reaction (NEBNext High-Fidelity 2× PCR Master Mix; NEB, M0541S). .. Ten percent of the PCR product was taken out for real-time PCR analysis to determine the amplification cycles of the library, followed immediately by PCR amplification of the remaining library DNA.

    Multiplex Assay:

    Article Title: Chromatinized Protein Kinase C-θ Directly Regulates Inducible Genes in Epithelial to Mesenchymal Transition and Breast Cancer Stem Cells
    Article Snippet: .. AMPure beads were used to isolate library fragments ranging between 175 and 225 bp in size, and the purified DNA was combined with the NEBNext High-Fidelity 2X PCR master mix, 25 μM Universal PCR primer, and 25 μM index primer for multiplexing purposes (NEBNext multiplex oligonucleotides for Illumina, NEB number E7335L; New England BioLabs Inc.). .. A total of 15 cycles were used for PCR amplification of the adaptor-ligated DNA.

    Selection:

    Article Title: Systematic interrogation of β-catenin co-dependent genes facilitated by integrated genetic and proteomic approaches
    Article Snippet: .. Following puromycin selection (2 μg/ml), genomic DNA (5μg) extracted at 3 or 28 DPI was PCR amplified using NEBNext® High-Fidelity 2X PCR Master Mix (New England Biolabs) and F/RPCR1_2sg primers ( ). .. This mixture (5μl) was used for a second PCR amplification using barcoded-staggered primers ( ).

    Agarose Gel Electrophoresis:

    Article Title: Transcriptomic but not genomic variability confers phenotype of breast cancer stem cells
    Article Snippet: G. PCR amplification: PCR enrichment was conducted by using 22 μL Adapter-ligated DNA, 25 μL of 2 × NEB Next high-fidelity PCR master buffer (NEB, Cat. M0541L), 1.5 μL of 10 μmol/L MUP primer, 1.5 μL of 10 μmol/L barcode primer. .. H. Extraction and purification of the final library: electrophoresis was conducted in agarose gel with the PCR products from the last step and then extracting and purifying DNA from agarose gel using the gel extraction kit (TIANGEN, Cat. DP214-03).

    Random Hexamer Labeling:

    Article Title: Transcriptome-wide identification of novel circular RNAs in soybean in response to low-phosphorus stress
    Article Snippet: First-strand cDNA was then synthesized by reverse transcriptase and random hexamer primers. .. Quantitative polymerase chain reaction (qPCR) was subsequently conducted in conjunction with universal PCR primers, NEBNext High-Fidelity PCR Master Mix, and Index (X) primer.

    Produced:

    Article Title: PCR amplification of a triple-repeat genetic target directly from whole blood in 15 minutes as a proof-of-principle PCR study for direct sample analysis for a clinically relevant target
    Article Snippet: .. For the purposes of our study, the NEBNext High-Fidelity 2X PCR Master Mix was used because it produced PCR product successfully for both rapid cycling and direct-blood analysis when paired with the Philisa Thermal Cycler. .. Data from Figure indicate the assay could tolerate up to 30% whole blood in a given PCR reaction tube and the cycling time was completed in 15 minutes, including a 3-minute hot-start/cell lysis step.

    Concentration Assay:

    Article Title: PCR amplification of a triple-repeat genetic target directly from whole blood in 15 minutes as a proof-of-principle PCR study for direct sample analysis for a clinically relevant target
    Article Snippet: For the purposes of our study, the NEBNext High-Fidelity 2X PCR Master Mix was used because it produced PCR product successfully for both rapid cycling and direct-blood analysis when paired with the Philisa Thermal Cycler. .. Some qualitative loss of band intensity is observed during testing of whole blood concentrations at 30%, which likely results from increased inhibitor concentration scaled alongside increased template input.

    Article Title: Clinical impact of measurable residual disease monitoring by ultradeep next generation sequencing in NPM1 mutated acute myeloid leukemia
    Article Snippet: Assay setup was as follows: For a total reaction volume of 75 μl, 37.5 μl of NEBNext High-Fidelity 2X PCR Master Mix (New England BioLabs Inc., Massachusetts, USA), and 10 uM each forward and reverse primers were used to amplify 600ng of genomic DNA (approximately 1,00,000 genomic cell equivalents). .. Samples were deep sequenced after pooling in equimolar concentration on an Illumina MiSeq (Illumina, San Diego, CA, USA) next generation sequencer using a 150 bp paired end V2 chemistry.

    Article Title: Transcriptomic but not genomic variability confers phenotype of breast cancer stem cells
    Article Snippet: G. PCR amplification: PCR enrichment was conducted by using 22 μL Adapter-ligated DNA, 25 μL of 2 × NEB Next high-fidelity PCR master buffer (NEB, Cat. M0541L), 1.5 μL of 10 μmol/L MUP primer, 1.5 μL of 10 μmol/L barcode primer. .. Quality control and sequencing of the final library: The quality and concentration of DNA fragments in the DNA libraries generated were assessed using High-Sensitivity Bioanalyzer.

    Article Title: Smokeless tobacco impacts oral microbiota in a Syrian Golden hamster cheek pouch carcinogenesis model
    Article Snippet: All genomic DNA samples were normalized to an equal concentration (10 ng/μl), and then used for bacterial 16S rRNA library construction using primers for the V1-2 region and the V4 region in separate PCR amplifications. .. Each PCR reaction contained 1 μl 5 μM specified 16 s F primer , 1 μl 5 μM specified 16 s R primer , 1 μl gDNA, 10 μl NEBNext High-Fidelity 2X PCR Master Mix (New England BioLabs Inc., Ipswich, MA), and 7 μl deionized H2 O.

    DNA Purification:

    Article Title: Transcriptomic but not genomic variability confers phenotype of breast cancer stem cells
    Article Snippet: D. Column purification: The PCR products were purified with Universal DNA Purification Kit (TIANGEN, Cat. DP214-03). .. G. PCR amplification: PCR enrichment was conducted by using 22 μL Adapter-ligated DNA, 25 μL of 2 × NEB Next high-fidelity PCR master buffer (NEB, Cat. M0541L), 1.5 μL of 10 μmol/L MUP primer, 1.5 μL of 10 μmol/L barcode primer.

    Article Title: Chromatinized Protein Kinase C-θ Directly Regulates Inducible Genes in Epithelial to Mesenchymal Transition and Breast Cancer Stem Cells
    Article Snippet: AMPure beads were used to isolate library fragments ranging between 175 and 225 bp in size, and the purified DNA was combined with the NEBNext High-Fidelity 2X PCR master mix, 25 μM Universal PCR primer, and 25 μM index primer for multiplexing purposes (NEBNext multiplex oligonucleotides for Illumina, NEB number E7335L; New England BioLabs Inc.). .. The resultant ChIP-DNA library was subjected to DNA purification with AMPure beads.

    High Throughput Screening Assay:

    Article Title: Microbiological Contamination at Workplaces in a Combined Heat and Power (CHP) Station Processing Plant Biomass
    Article Snippet: The amplification of the fungal ITS1 region for high-throughput sequencing analysis was performed with primers given in . .. PCR reactions (both for amplification of 16S rRNA gene fragments and ITS1 regions) were prepared in 50 μL volume containing 5 μL of DNA as template, 25 μL NEBNext® Hot Start High-Fidelity 2× PCR Master Mix (New England BioLabs, Ipswich, MA, USA) and 10 pmol of 341F and 785R (16S rRNA gene) or IT1F12 and ITS2 (ITS1 region) primers.

    Lysis:

    Article Title: PCR amplification of a triple-repeat genetic target directly from whole blood in 15 minutes as a proof-of-principle PCR study for direct sample analysis for a clinically relevant target
    Article Snippet: For the purposes of our study, the NEBNext High-Fidelity 2X PCR Master Mix was used because it produced PCR product successfully for both rapid cycling and direct-blood analysis when paired with the Philisa Thermal Cycler. .. Data from Figure indicate the assay could tolerate up to 30% whole blood in a given PCR reaction tube and the cycling time was completed in 15 minutes, including a 3-minute hot-start/cell lysis step.

    Article Title: TRIM28-Regulated Transposon Repression Is Required for Human Germline Competency and Not Primed or Naive Human Pluripotency
    Article Snippet: ATAC-Seq Cells were lysed directly in lysis buffer (500 μL 1 M Tris [pH 7.4], 100 μL 5 M NaCl, 150 μL 1 M MgCl2 , 500 μL 10% NP-40 in 50 mL water). .. After centrifugation, the cell pellet was resuspended in transposase buffer, which contained the Tn5 transposase enzyme and tagmentation buffer (Nextera DNA library prep kit: Illumina, catalog no. 15028212), and incubated at 37°C for 30 min. After purification by a MinElute PCR Purification Kit 250 (Qiagen 28006), P1 barcode (25 μM) and appropriate P2 barcodes (25 μM) were added to the DNA and run for five cycles of PCR reaction (NEBNext High-Fidelity 2× PCR Master Mix; NEB, M0541S).

    Variant Assay:

    Article Title: Systematic interrogation of β-catenin co-dependent genes facilitated by integrated genetic and proteomic approaches
    Article Snippet: To minimize recombination events between repetitive sequences, we inserted a variant U6 promoter downstream of the U6 promoter in pXRP003 ( ). .. Following puromycin selection (2 μg/ml), genomic DNA (5μg) extracted at 3 or 28 DPI was PCR amplified using NEBNext® High-Fidelity 2X PCR Master Mix (New England Biolabs) and F/RPCR1_2sg primers ( ).

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    New England Biolabs high fidelity pcr master mix
    Validation of a circRNAs (novel_circ_000237) by <t>qPCR</t> and Sanger sequencing. M, DL500 marker and the red arrow represent 100 bp. The yellow arrows denote the divergent primers for <t>PCR</t> amplification orientation.
    High Fidelity Pcr Master Mix, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 140 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Validation of a circRNAs (novel_circ_000237) by qPCR and Sanger sequencing. M, DL500 marker and the red arrow represent 100 bp. The yellow arrows denote the divergent primers for PCR amplification orientation.

    Journal: PLoS ONE

    Article Title: Transcriptome-wide identification of novel circular RNAs in soybean in response to low-phosphorus stress

    doi: 10.1371/journal.pone.0227243

    Figure Lengend Snippet: Validation of a circRNAs (novel_circ_000237) by qPCR and Sanger sequencing. M, DL500 marker and the red arrow represent 100 bp. The yellow arrows denote the divergent primers for PCR amplification orientation.

    Article Snippet: Quantitative polymerase chain reaction (qPCR) was subsequently conducted in conjunction with universal PCR primers, NEBNext High-Fidelity PCR Master Mix, and Index (X) primer.

    Techniques: Real-time Polymerase Chain Reaction, Sequencing, Marker, Polymerase Chain Reaction, Amplification