dna repair  (New England Biolabs)


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  • 99
    Name:
    NEBNext FFPE DNA Repair Mix
    Description:

    Catalog Number:
    M6630L
    Price:
    None
    Score:
    85
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    Structured Review

    New England Biolabs dna repair
    Genome-wide base composition bias curves in Illumina reads from PCR-free human <t>DNA</t> libraries. ( a ) The GC-bias curves from libraries (in duplicate) produced by the immobilized enzyme method (IM-1 and IM-2 in blue), for end repair for 30 min at 20 °C and 3′ A-tailing at 37 °C in contrast to the data from the libraries generated by the soluble enzyme method, with 3′ A-tailing at 65 °C, using enzyme mixture PKT (PKT-1 and PKT-2 in purple). ( b ) The GC-bias data of the immobilized enzyme method compared to the data from the duplicate libraries generated by Illumina TruSeq DNA PCR-free LT Library Preparation Kit (Illumina), Kapa Hyper Prep Kit (Kapa) or <t>NEBNext</t> Ultra II DNA Library Prep Kit for Illumina (Ultra) according to the protocols of the manufacturers. The Illumina protocol carries out end repair for 30 min at 30 °C and 3′ A-tailing for 30 min at 37 °C, followed by incubation at 70 °C for 5 min, and includes a clean-up and size selection step between end repair and 3′ A-tailing. The Kapa Hyper and NEBNext Ultra workflows include an enzyme mixture to perform end repair for 30 min at 20 °C, followed by 3′ A-tailing for 30 min at 65 °C.

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    Images

    1) Product Images from "Solid-phase enzyme catalysis of DNA end repair and 3′ A-tailing reduces GC-bias in next-generation sequencing of human genomic DNA"

    Article Title: Solid-phase enzyme catalysis of DNA end repair and 3′ A-tailing reduces GC-bias in next-generation sequencing of human genomic DNA

    Journal: Scientific Reports

    doi: 10.1038/s41598-018-34079-2

    Genome-wide base composition bias curves in Illumina reads from PCR-free human DNA libraries. ( a ) The GC-bias curves from libraries (in duplicate) produced by the immobilized enzyme method (IM-1 and IM-2 in blue), for end repair for 30 min at 20 °C and 3′ A-tailing at 37 °C in contrast to the data from the libraries generated by the soluble enzyme method, with 3′ A-tailing at 65 °C, using enzyme mixture PKT (PKT-1 and PKT-2 in purple). ( b ) The GC-bias data of the immobilized enzyme method compared to the data from the duplicate libraries generated by Illumina TruSeq DNA PCR-free LT Library Preparation Kit (Illumina), Kapa Hyper Prep Kit (Kapa) or NEBNext Ultra II DNA Library Prep Kit for Illumina (Ultra) according to the protocols of the manufacturers. The Illumina protocol carries out end repair for 30 min at 30 °C and 3′ A-tailing for 30 min at 37 °C, followed by incubation at 70 °C for 5 min, and includes a clean-up and size selection step between end repair and 3′ A-tailing. The Kapa Hyper and NEBNext Ultra workflows include an enzyme mixture to perform end repair for 30 min at 20 °C, followed by 3′ A-tailing for 30 min at 65 °C.
    Figure Legend Snippet: Genome-wide base composition bias curves in Illumina reads from PCR-free human DNA libraries. ( a ) The GC-bias curves from libraries (in duplicate) produced by the immobilized enzyme method (IM-1 and IM-2 in blue), for end repair for 30 min at 20 °C and 3′ A-tailing at 37 °C in contrast to the data from the libraries generated by the soluble enzyme method, with 3′ A-tailing at 65 °C, using enzyme mixture PKT (PKT-1 and PKT-2 in purple). ( b ) The GC-bias data of the immobilized enzyme method compared to the data from the duplicate libraries generated by Illumina TruSeq DNA PCR-free LT Library Preparation Kit (Illumina), Kapa Hyper Prep Kit (Kapa) or NEBNext Ultra II DNA Library Prep Kit for Illumina (Ultra) according to the protocols of the manufacturers. The Illumina protocol carries out end repair for 30 min at 30 °C and 3′ A-tailing for 30 min at 37 °C, followed by incubation at 70 °C for 5 min, and includes a clean-up and size selection step between end repair and 3′ A-tailing. The Kapa Hyper and NEBNext Ultra workflows include an enzyme mixture to perform end repair for 30 min at 20 °C, followed by 3′ A-tailing for 30 min at 65 °C.

    Techniques Used: Genome Wide, Polymerase Chain Reaction, Gas Chromatography, Produced, Generated, Incubation, Selection

    Related Articles

    Centrifugation:

    Article Title: Nanopore sequencing and assembly of a human genome with ultra-long reads
    Article Snippet: 1.5–2.5 μg human genomic DNA was sheared in a Covaris g-TUBE centrifuged at 5,000–6,000 r.p.m. in an Eppendorf 5424 (or equivalent) centrifuge for 2 × 1 min, inverting the tube between centrifugation steps. .. DNA repair (NEBNext FFPE DNA Repair Mix, NEB M6630) was performed on purchased DNA but not on freshly extracted DNA.

    Article Title: De novo yeast genome assemblies from MinION, PacBio and MiSeq platforms
    Article Snippet: Two ug of genomic DNA was sheared to approximately 18,000 bp by centrifugation at 4000 rpm in a gTUBE. .. Sequencing libraries were prepared according to the SQK-MAP006 or SQK-NSK007 Sequencing Kit protocol, including the NEBNext FFPE DNA repair step.

    Article Title: Nanopore DNA Sequencing and Genome Assembly on the International Space Station
    Article Snippet: Aliquots of DNA for mouse, E. coli , and lambda libraries were sheared individually using Covaris g-TUBEs (Covaris, Boston, MA) by centrifugation at 4,800 × g for 2 min to produce fragments that were predominantly 8 kb. .. Mixed DNA samples then underwent treatment to repair residual nicks, gaps, and blocked 3′ ends using Formalin-Fixed, Paraffin-Embedded (FFPE) DNA Repair Mix (New England Biolabs), with modifications to the manufacturer’s protocol to include a 0.5X Agencourt AMPure XP system magnetic bead clean-up (Beckman Coulter Genomics, Brea, CA) to remove small fragments of DNA.

    Amplification:

    Article Title: Artifactual mutations resulting from DNA lesions limit detection levels in ultrasensitive sequencing applications
    Article Snippet: Although, we showed that Fpg can effectively eliminate rare 8-oxoG products even in larger amounts of DNA like genomic DNA, it is very important to consider that after Fpg treatment only the complementary strand without the 8-oxoG gets amplified, which could introduce other types of biases at nearby amplifiable lesions on the non 8-oxoG containing strand. .. In order to overcome this issue, instead of Fpg, DNA repair mixes could be used, such as available for formalin fixed, paraffin embedded (FFPE) samples (e.g. NEBNext FFPE DNA Repair Mix from NEB) or forensic samples (e.g. PreCR Repair Mix ), which include Fpg and other enzymes for the repair of generated abasic sites generating an amplifiable DNA template.

    Lambda DNA Preparation:

    Article Title: Nanopore DNA Sequencing and Genome Assembly on the International Space Station
    Article Snippet: Sheared mouse, E. coli , and lambda DNA samples were quantified using a Qubit 2.0 fluorometer (Thermo Fisher Scientific, Waltham, MA) and combined in equal abundances, targeting 1.5 µg total (0.5 µg each). .. Mixed DNA samples then underwent treatment to repair residual nicks, gaps, and blocked 3′ ends using Formalin-Fixed, Paraffin-Embedded (FFPE) DNA Repair Mix (New England Biolabs), with modifications to the manufacturer’s protocol to include a 0.5X Agencourt AMPure XP system magnetic bead clean-up (Beckman Coulter Genomics, Brea, CA) to remove small fragments of DNA.

    Construct:

    Article Title: Nanopore sequencing of complex genomic rearrangements in yeast reveals mechanisms of repeat-mediated double-strand break repair
    Article Snippet: Using the ONT (Oxford Nanopore Technologies) Ligation Sequencing Kit 1D (SQK-LSK108) in combination with the Native Barcoding Kit 1D (EXP-NBD103), 1.5 µg of purified DNA was used to construct barcoded sequencing libraries. .. All procedures recommended in the ONT-provided protocol were followed, including nick repair (NEBNext FFPE Repair mix, New England Biolabs).

    Incubation:

    Article Title: Nanopore sequencing and assembly of a human genome with ultra-long reads
    Article Snippet: DNA repair (NEBNext FFPE DNA Repair Mix, NEB M6630) was performed on purchased DNA but not on freshly extracted DNA. .. 8.5 μl nuclease-free water (NFW), 6.5 μl FFPE Repair Buffer and 2 μl FFPE DNA Repair Mix were added to the 46 μl sheared DNA.

    Article Title: In Situ Field Sequencing and Life Detection in Remote (79°26′N) Canadian High Arctic Permafrost Ice Wedge Microbial Communities
    Article Snippet: These kits were used according to manufacturer's instruction, but with the omission of the DNA shearing step. .. Briefly, 46 μL of DNA extract was used for a DNA repair step using NEBNext FFPE Repair Mix (New England Biolabs, Ipswich, MA), followed by clean-up using AMPure XP beads incubated with mixing at room temperature for 5 min. .. Samples were ethanol cleaned while on a magnetic rack.

    Article Title: De Novo Assembly of a New Solanum pennellii Accession Using Nanopore Sequencing
    Article Snippet: Subsequently, up to 5 μg of DNA was used for NEBNext FFPE DNA Repair (New England Biolabs) in a total volume of 155 μL including 16.3 μL NEBNext FFPE DNA Repair Buffer and 5 μL NEBNext FFPE DNA Repair Mix. .. Subsequently, up to 5 μg of DNA was used for NEBNext FFPE DNA Repair (New England Biolabs) in a total volume of 155 μL including 16.3 μL NEBNext FFPE DNA Repair Buffer and 5 μL NEBNext FFPE DNA Repair Mix.

    Article Title: High contiguity Arabidopsis thaliana genome assembly with a single nanopore flow cell
    Article Snippet: The resulting pellet was washed with freshly prepared ice-cold 70% ethanol, dried, and resuspended in 350 μL 1× TE buffer (10 mM Tris-HCl, 1 mM EDTA, pH 8.0) with 5 μL RNase A (Qiagen, Hilden) at 37 °C for 30 min, followed by incubation at 4 °C overnight. .. Briefly, approximately 2 μg of purified DNA, without exogenous shearing or size selection, was repaired with NEBNext FFPE Repair Mix for 60 min at 20 °C.

    Formalin-fixed Paraffin-Embedded:

    Article Title: de novo assembly and population genomic survey of natural yeast isolates with the Oxford Nanopore MinION sequencer
    Article Snippet: Two μg of genomic DNA was sheared to approximately 8000 bp with g-TUBE. .. After clean-up using 1x AMPure XP beads, sequencing libraries were prepared according to the SQK-MAP005 or SQK-MAP006 Sequencing Kit protocol, including the PreCR treatment for the SQK-MAP005 protocol or the NEBNext FFPE DNA repair step for the SQK-MAP006 protocol. .. The following protocol was applied to some samples (Supplementary File 3).

    Article Title: Nanopore sequencing and assembly of a human genome with ultra-long reads
    Article Snippet: 1.5–2.5 μg human genomic DNA was sheared in a Covaris g-TUBE centrifuged at 5,000–6,000 r.p.m. in an Eppendorf 5424 (or equivalent) centrifuge for 2 × 1 min, inverting the tube between centrifugation steps. .. DNA repair (NEBNext FFPE DNA Repair Mix, NEB M6630) was performed on purchased DNA but not on freshly extracted DNA. .. 8.5 μl nuclease-free water (NFW), 6.5 μl FFPE Repair Buffer and 2 μl FFPE DNA Repair Mix were added to the 46 μl sheared DNA.

    Article Title: Antimicrobial resistance prediction and phylogenetic analysis of Neisseria gonorrhoeae isolates using the Oxford Nanopore MinION sequencer
    Article Snippet: Three µg of genomic DNA was sheared to an average fragment length of 8 kb with g-TUBES (Covaris, Woburn, WA, USA). .. Sequencing libraries were prepared according to the 2D library preparation protocol with the SQK-LSK208 2D ligation kit (Oxford Nanopore Technologies), including the DNA repair step with the NEBNext FFPE DNA repair module (New England Biolabs, Ipswich, MA, USA). .. The sequencing libraries were purified using MyOne C1 beads (Thermo Fisher Scientific), and 6 μl of sequencing library were loaded onto a R9.4 SpotON flow cell and sequenced with the MinION Mk 1B sequencing device (Oxford Nanopore Technologies) for 24 hours, including a top-up with additional 6 µl of DNA library after the first 6 hours of the sequencing run.

    Article Title: De novo yeast genome assemblies from MinION, PacBio and MiSeq platforms
    Article Snippet: Two ug of genomic DNA was sheared to approximately 18,000 bp by centrifugation at 4000 rpm in a gTUBE. .. Sequencing libraries were prepared according to the SQK-MAP006 or SQK-NSK007 Sequencing Kit protocol, including the NEBNext FFPE DNA repair step. .. The sequencing mix was prepared with 6 uL of the DNA library, water, the Fuel Mix and the running buffer according to the SQK-MAP006 or the SQK-MAP007 protocols.

    Article Title: Hybrid genome assembly and annotation of Paenibacillus pasadenensis strain R16 reveals insights on endophytic life style and antifungal activity
    Article Snippet: The library preparation was performed using components from the Genomic DNA Sequencing Kit SQK-MAP006 (Oxford Nanopore Technologies) and following Version R9 of the Oxford Nanopore protocol MAP006. .. DNA nicks were repaired using the FFPE DNA repair mix (New England Biolabs, NEB) and End-repair and dA-tailing were performed using the Ultra II End Prep Module (New Englands Biolabs, NEB Ipswich, USA) according to manufacturer’s instructions. .. The adapter utilized consisted of a linear double strand sequence and a harpin sequence that links the positive and negative strand of each fragment to allow the sequencing of both strands (2D reads).

    Article Title: Highly Contiguous Genome Assemblies of 15 Drosophila Species Generated Using Nanopore Sequencing
    Article Snippet: Water or TE was added to DNA for a total volume of 46 µL. .. For 4 of 21 library preps (Table S1), the FFPE repair and dA-Tailing steps were combined in the following reaction mix: 46.5 µL of genomic DNA in TE, 3.5 µL of UltraII EP Buffer (NEB), 3.5 µL of FFPE DNA Repair Buffer (NEB), 3 µL of UltraII EP Enzyme (NEB), 3 µL of FFPE Repair Mix (NEB), and 0.5 µL of 100x NAD+ (NEB). .. The combined reaction was prepared in a 200-µL PCR tube and run at 20° for 1 hr followed by 65° for 30 min in a thermocycler.

    Article Title: de novo assembly and population genomic survey of natural yeast isolates with the Oxford Nanopore MinION sequencer
    Article Snippet: Five hundred ng of genomic DNA was sheared to approximately 8000 bp with g-TUBE. .. After clean-up using 1x AMPure XP beads and the NEBNext FFPE DNA repair step, 100 ng of DNA was prepared according to the Low Input Expansion Pack Protocol for genomic DNA. .. The sequencing mix was prepared with 8 μl of the DNA library, water, the fuel mix, and the running buffer according to the SQK-MAP005 or the SQK-MAP006 protocols.

    Article Title: Nanopore DNA Sequencing and Genome Assembly on the International Space Station
    Article Snippet: Sheared mouse, E. coli , and lambda DNA samples were quantified using a Qubit 2.0 fluorometer (Thermo Fisher Scientific, Waltham, MA) and combined in equal abundances, targeting 1.5 µg total (0.5 µg each). .. Mixed DNA samples then underwent treatment to repair residual nicks, gaps, and blocked 3′ ends using Formalin-Fixed, Paraffin-Embedded (FFPE) DNA Repair Mix (New England Biolabs), with modifications to the manufacturer’s protocol to include a 0.5X Agencourt AMPure XP system magnetic bead clean-up (Beckman Coulter Genomics, Brea, CA) to remove small fragments of DNA. .. The repaired DNA was then prepared for sequencing according to Oxford Nanopore Technologies’ procedure for the SQK-MAP-006 Kit.

    Article Title: Nanopore sequencing of complex genomic rearrangements in yeast reveals mechanisms of repeat-mediated double-strand break repair
    Article Snippet: Using the ONT (Oxford Nanopore Technologies) Ligation Sequencing Kit 1D (SQK-LSK108) in combination with the Native Barcoding Kit 1D (EXP-NBD103), 1.5 µg of purified DNA was used to construct barcoded sequencing libraries. .. All procedures recommended in the ONT-provided protocol were followed, including nick repair (NEBNext FFPE Repair mix, New England Biolabs). .. The libraries were pooled and sequenced together on a single SpotON Flow Cell Mk I R9.4 (FLO-SPOTR9) for 48 h.

    Article Title: Biosynthesis of the antibiotic nonribosomal peptide penicillin in baker's yeast
    Article Snippet: Enriched assembly DNA for the multigene and single gene assemblies was quantified on a Qubit 2.0 fluorometer (Thermo Fisher Scientific) using a Qubit dsDNA HS Assay Kit (Thermo Fisher Scientific). .. The four samples were combined to a give an equimolar mix (assuming a molecular weight for each assembly based on the mean promoter length) with a total DNA content of 2.6 μg in 45 μl dH2 O. DNA underwent end repair using NEBNext FFPE DNA Repair Mix (M6630, New England Biolabs) according to the manufacturer's instructions. .. The repaired DNA was recovered using Agincourt AMPure XP beads (A63880, Beckman Coulter), washed twice in 200 μl 70% ethanol and eluted in 46 μl dH2 O. DNA was then dA-tailed using NEBNext Ultra II End Repair/dA-Tailing Module (E7546, New England Biolabs) according to the manufacturer's instructions and recovered using Agincourt AMPure XP beads as before, eluting in 30 μl dH2 O.

    Article Title: De novo whole-genome assembly of a wild type yeast isolate using nanopore sequencing
    Article Snippet: In short, high molecular weight DNA was sheared with a g-TUBE (Covaris) to an average fragment length of 20 kbp. .. The sheared DNA was repaired using the FFPE Repair Mix, according to the manufacturer’s instructions (New England Biolabs). .. After cleaning the DNA with bead extraction, using a ratio of 0.4:1 Ampure XP beads (Beckman Coulter) to DNA, the DNA ends were polished and an A overhang was added with the NEBNext End Prep Module (New England Biolabs).

    Article Title: De Novo Assembly of a New Solanum pennellii Accession Using Nanopore Sequencing
    Article Snippet: Bead binding was performed at room temperature and elution at 37°C. .. Subsequently, up to 5 μg of DNA was used for NEBNext FFPE DNA Repair (New England Biolabs) in a total volume of 155 μL including 16.3 μL NEBNext FFPE DNA Repair Buffer and 5 μL NEBNext FFPE DNA Repair Mix. .. To reduce DNA shearing during the following bead clean-up, the sample was split in two 77.5-μL aliquots that were each eluted in 50.5 μL nuclease free water.

    Article Title: Copy number analysis by low coverage whole genome sequencing using ultra low-input DNA from formalin-fixed paraffin embedded tumor tissue
    Article Snippet: A standard human genomic DNA was used as a positive control provided with the Genome Plex WGA kit (Sigma) and a no template control was used as a negative control. .. The NEB Next FFPE Repair kit (NEB M6630, New England® Biolabs Inc) was used for repairing 150 ng of total DNA, according to the manufacturer’s protocol with a minor change of eluting DNA in 30 μL instead of 40 μL. .. A total of 10 μL of eluted DNA (total 50 ng of repaired DNA) was used for WGA using the Sigma WGA kit as described above.

    Article Title: Rapid de novo assembly of the European eel genome from nanopore sequencing reads
    Article Snippet: Briefly, high molecular weight DNA was sheared with a g-TUBE (Covaris) to an average fragment length of 20 kbp. .. The sheared DNA was repaired using the FFPE repair mix according to the manufacturer’s instructions (New England Biolabs, Ipswich, USA). .. After cleaning up the DNA with an extraction using a ratio of 0.4:1 Ampure XP beads to DNA the DNA ends were polished and an A overhang was added with the the NEBNext End Prep Module and again cleaned up with an extraction using a ratio of 1:1 Ampure XP beads to DNA the DNA prior to ligation.

    Article Title: High contiguity Arabidopsis thaliana genome assembly with a single nanopore flow cell
    Article Snippet: The purified DNA was then prepared for sequencing following the protocol in the genomic sequencing kit SQK-LSK108 (ONT, Oxford, UK). .. Briefly, approximately 2 μg of purified DNA, without exogenous shearing or size selection, was repaired with NEBNext FFPE Repair Mix for 60 min at 20 °C. .. The DNA was purified with 0.5× Ampure XP beads (Beckman Coulter).

    Article Title: De novo assembly and annotation of three Leptosphaeria genomes using Oxford Nanopore MinION sequencing
    Article Snippet: Briefly, 100 ng to 1.5 μg of genomic DNA was sheared to approximately 8 Kb with g-TUBE (Covaris, Woburn, MA, USA) and cleaned-up using AMPure XP beads (Beckmann Coulter Genomics). .. For some libraries, DNA fragments were repaired using the NEBNext FFPE Repair Mix (New England Biolabs). .. After end-reparation and 3′-adenylation with the NEBNext® UltraTM II End Repair/dA-Tailing Module (New England Biolabs), sequencing adapters provided by Oxford Nanopore Technologies (Oxford Nanopore Technologies Ltd, UK) were ligated using Blunt/TA Ligase Master Mix (New England Biolabs).

    Article Title: Comparison of bacterial genome assembly software for MinION data and their applicability to medical microbiology
    Article Snippet: DNA was extracted using the QiaAMP DNA Mini kit (Qiagen), and quantified using the Qubit fluorimeter (Life Technologies). .. Sample preparation was carried out using the Genomic DNA Sequencing Kit SQK-MAP-006 (Oxford Nanopore Technologies) following the manufacturers instructions, including the optional NEBNext FFPE DNA repair step (NEB). .. A 6 µl aliquot of pre-sequencing mix was combined with 4 µl Fuel Mix (Oxford Nanopore), 75 µl running buffer (Oxford Nanopore) and 66 µl water and added to the flow cell.

    Significance Assay:

    Article Title: Artifactual mutations resulting from DNA lesions limit detection levels in ultrasensitive sequencing applications
    Article Snippet: Fpg treatments of genomic DNA also showed a significant ∼4-fold reduction (P value = 0.01, Fisher’s exact test) in G- > T/C- > A and G- > GT/C- > CA transversions , but given already the low frequencies of this substitution, it is unclear if the remaining transversions in genomic DNA are still due to 8-oxoG lesions, other types of artifacts, or represent rare true mutations in the genome. .. In order to overcome this issue, instead of Fpg, DNA repair mixes could be used, such as available for formalin fixed, paraffin embedded (FFPE) samples (e.g. NEBNext FFPE DNA Repair Mix from NEB) or forensic samples (e.g. PreCR Repair Mix ), which include Fpg and other enzymes for the repair of generated abasic sites generating an amplifiable DNA template.

    Flow Cytometry:

    Article Title: Hybrid genome assembly and annotation of Paenibacillus pasadenensis strain R16 reveals insights on endophytic life style and antifungal activity
    Article Snippet: DNA nicks were repaired using the FFPE DNA repair mix (New England Biolabs, NEB) and End-repair and dA-tailing were performed using the Ultra II End Prep Module (New Englands Biolabs, NEB Ipswich, USA) according to manufacturer’s instructions. .. Prior to sequencing the MinION flowcell quality control was carried out in order to determine the number of available pores for the sequencing.

    Article Title: Highly Contiguous Genome Assemblies of 15 Drosophila Species Generated Using Nanopore Sequencing
    Article Snippet: For 4 of 21 library preps (Table S1), the FFPE repair and dA-Tailing steps were combined in the following reaction mix: 46.5 µL of genomic DNA in TE, 3.5 µL of UltraII EP Buffer (NEB), 3.5 µL of FFPE DNA Repair Buffer (NEB), 3 µL of UltraII EP Enzyme (NEB), 3 µL of FFPE Repair Mix (NEB), and 0.5 µL of 100x NAD+ (NEB). .. The combined reaction was prepared in a 200-µL PCR tube and run at 20° for 1 hr followed by 65° for 30 min in a thermocycler.

    Article Title: De novo whole-genome assembly of a wild type yeast isolate using nanopore sequencing
    Article Snippet: Subsequently, multiple R9 and R9.4 flow cells were used to sequence the DNA. .. The sheared DNA was repaired using the FFPE Repair Mix, according to the manufacturer’s instructions (New England Biolabs).

    Article Title: Rapid de novo assembly of the European eel genome from nanopore sequencing reads
    Article Snippet: Subsequently multiple R9 and R9.4 flow cells were used to sequence the DNA. .. The sheared DNA was repaired using the FFPE repair mix according to the manufacturer’s instructions (New England Biolabs, Ipswich, USA).

    Article Title: Comparison of bacterial genome assembly software for MinION data and their applicability to medical microbiology
    Article Snippet: Sample preparation was carried out using the Genomic DNA Sequencing Kit SQK-MAP-006 (Oxford Nanopore Technologies) following the manufacturers instructions, including the optional NEBNext FFPE DNA repair step (NEB). .. Sample preparation was carried out using the Genomic DNA Sequencing Kit SQK-MAP-006 (Oxford Nanopore Technologies) following the manufacturers instructions, including the optional NEBNext FFPE DNA repair step (NEB).

    Ligation:

    Article Title: Nanopore sequencing and assembly of a human genome with ultra-long reads
    Article Snippet: Paragraph title: Library preparation (SQK-LSK108 1D ligation genomic DNA) ... DNA repair (NEBNext FFPE DNA Repair Mix, NEB M6630) was performed on purchased DNA but not on freshly extracted DNA.

    Article Title: Antimicrobial resistance prediction and phylogenetic analysis of Neisseria gonorrhoeae isolates using the Oxford Nanopore MinION sequencer
    Article Snippet: Three µg of genomic DNA was sheared to an average fragment length of 8 kb with g-TUBES (Covaris, Woburn, WA, USA). .. Sequencing libraries were prepared according to the 2D library preparation protocol with the SQK-LSK208 2D ligation kit (Oxford Nanopore Technologies), including the DNA repair step with the NEBNext FFPE DNA repair module (New England Biolabs, Ipswich, MA, USA). .. The sequencing libraries were purified using MyOne C1 beads (Thermo Fisher Scientific), and 6 μl of sequencing library were loaded onto a R9.4 SpotON flow cell and sequenced with the MinION Mk 1B sequencing device (Oxford Nanopore Technologies) for 24 hours, including a top-up with additional 6 µl of DNA library after the first 6 hours of the sequencing run.

    Article Title: Highly Contiguous Genome Assemblies of 15 Drosophila Species Generated Using Nanopore Sequencing
    Article Snippet: Libraries were prepared using the Ligation Sequencing Kit 1D (Oxford Nanopore) either according to or with slight modifications to the manufacturer’s protocol. .. For 4 of 21 library preps (Table S1), the FFPE repair and dA-Tailing steps were combined in the following reaction mix: 46.5 µL of genomic DNA in TE, 3.5 µL of UltraII EP Buffer (NEB), 3.5 µL of FFPE DNA Repair Buffer (NEB), 3 µL of UltraII EP Enzyme (NEB), 3 µL of FFPE Repair Mix (NEB), and 0.5 µL of 100x NAD+ (NEB).

    Article Title: In Situ Field Sequencing and Life Detection in Remote (79°26′N) Canadian High Arctic Permafrost Ice Wedge Microbial Communities
    Article Snippet: Briefly, 46 μL of DNA extract was used for a DNA repair step using NEBNext FFPE Repair Mix (New England Biolabs, Ipswich, MA), followed by clean-up using AMPure XP beads incubated with mixing at room temperature for 5 min. .. Briefly, 46 μL of DNA extract was used for a DNA repair step using NEBNext FFPE Repair Mix (New England Biolabs, Ipswich, MA), followed by clean-up using AMPure XP beads incubated with mixing at room temperature for 5 min.

    Article Title: Nanopore sequencing of complex genomic rearrangements in yeast reveals mechanisms of repeat-mediated double-strand break repair
    Article Snippet: Using the ONT (Oxford Nanopore Technologies) Ligation Sequencing Kit 1D (SQK-LSK108) in combination with the Native Barcoding Kit 1D (EXP-NBD103), 1.5 µg of purified DNA was used to construct barcoded sequencing libraries. .. All procedures recommended in the ONT-provided protocol were followed, including nick repair (NEBNext FFPE Repair mix, New England Biolabs).

    Article Title: De novo whole-genome assembly of a wild type yeast isolate using nanopore sequencing
    Article Snippet: The sheared DNA was repaired using the FFPE Repair Mix, according to the manufacturer’s instructions (New England Biolabs). .. The sheared DNA was repaired using the FFPE Repair Mix, according to the manufacturer’s instructions (New England Biolabs).

    Article Title: Rapid de novo assembly of the European eel genome from nanopore sequencing reads
    Article Snippet: The sheared DNA was repaired using the FFPE repair mix according to the manufacturer’s instructions (New England Biolabs, Ipswich, USA). .. The sheared DNA was repaired using the FFPE repair mix according to the manufacturer’s instructions (New England Biolabs, Ipswich, USA).

    Genomic Sequencing:

    Article Title: High contiguity Arabidopsis thaliana genome assembly with a single nanopore flow cell
    Article Snippet: The purified DNA was then prepared for sequencing following the protocol in the genomic sequencing kit SQK-LSK108 (ONT, Oxford, UK). .. Briefly, approximately 2 μg of purified DNA, without exogenous shearing or size selection, was repaired with NEBNext FFPE Repair Mix for 60 min at 20 °C.

    Introduce:

    Article Title: Artifactual mutations resulting from DNA lesions limit detection levels in ultrasensitive sequencing applications
    Article Snippet: Although, we showed that Fpg can effectively eliminate rare 8-oxoG products even in larger amounts of DNA like genomic DNA, it is very important to consider that after Fpg treatment only the complementary strand without the 8-oxoG gets amplified, which could introduce other types of biases at nearby amplifiable lesions on the non 8-oxoG containing strand. .. In order to overcome this issue, instead of Fpg, DNA repair mixes could be used, such as available for formalin fixed, paraffin embedded (FFPE) samples (e.g. NEBNext FFPE DNA Repair Mix from NEB) or forensic samples (e.g. PreCR Repair Mix ), which include Fpg and other enzymes for the repair of generated abasic sites generating an amplifiable DNA template.

    Generated:

    Article Title: Artifactual mutations resulting from DNA lesions limit detection levels in ultrasensitive sequencing applications
    Article Snippet: Although, we showed that Fpg can effectively eliminate rare 8-oxoG products even in larger amounts of DNA like genomic DNA, it is very important to consider that after Fpg treatment only the complementary strand without the 8-oxoG gets amplified, which could introduce other types of biases at nearby amplifiable lesions on the non 8-oxoG containing strand. .. In order to overcome this issue, instead of Fpg, DNA repair mixes could be used, such as available for formalin fixed, paraffin embedded (FFPE) samples (e.g. NEBNext FFPE DNA Repair Mix from NEB) or forensic samples (e.g. PreCR Repair Mix ), which include Fpg and other enzymes for the repair of generated abasic sites generating an amplifiable DNA template. .. However, this enzymatic repair is not 100% accurate and might also be a source of error.

    DNA Sequencing:

    Article Title: Hybrid genome assembly and annotation of Paenibacillus pasadenensis strain R16 reveals insights on endophytic life style and antifungal activity
    Article Snippet: The library preparation was performed using components from the Genomic DNA Sequencing Kit SQK-MAP006 (Oxford Nanopore Technologies) and following Version R9 of the Oxford Nanopore protocol MAP006. .. DNA nicks were repaired using the FFPE DNA repair mix (New England Biolabs, NEB) and End-repair and dA-tailing were performed using the Ultra II End Prep Module (New Englands Biolabs, NEB Ipswich, USA) according to manufacturer’s instructions.

    Article Title: Comparison of bacterial genome assembly software for MinION data and their applicability to medical microbiology
    Article Snippet: DNA was extracted using the QiaAMP DNA Mini kit (Qiagen), and quantified using the Qubit fluorimeter (Life Technologies). .. Sample preparation was carried out using the Genomic DNA Sequencing Kit SQK-MAP-006 (Oxford Nanopore Technologies) following the manufacturers instructions, including the optional NEBNext FFPE DNA repair step (NEB). .. A 6 µl aliquot of pre-sequencing mix was combined with 4 µl Fuel Mix (Oxford Nanopore), 75 µl running buffer (Oxford Nanopore) and 66 µl water and added to the flow cell.

    Sequencing:

    Article Title: de novo assembly and population genomic survey of natural yeast isolates with the Oxford Nanopore MinION sequencer
    Article Snippet: Two μg of genomic DNA was sheared to approximately 8000 bp with g-TUBE. .. After clean-up using 1x AMPure XP beads, sequencing libraries were prepared according to the SQK-MAP005 or SQK-MAP006 Sequencing Kit protocol, including the PreCR treatment for the SQK-MAP005 protocol or the NEBNext FFPE DNA repair step for the SQK-MAP006 protocol. .. The following protocol was applied to some samples (Supplementary File 3).

    Article Title: Antimicrobial resistance prediction and phylogenetic analysis of Neisseria gonorrhoeae isolates using the Oxford Nanopore MinION sequencer
    Article Snippet: Three µg of genomic DNA was sheared to an average fragment length of 8 kb with g-TUBES (Covaris, Woburn, WA, USA). .. Sequencing libraries were prepared according to the 2D library preparation protocol with the SQK-LSK208 2D ligation kit (Oxford Nanopore Technologies), including the DNA repair step with the NEBNext FFPE DNA repair module (New England Biolabs, Ipswich, MA, USA). .. The sequencing libraries were purified using MyOne C1 beads (Thermo Fisher Scientific), and 6 μl of sequencing library were loaded onto a R9.4 SpotON flow cell and sequenced with the MinION Mk 1B sequencing device (Oxford Nanopore Technologies) for 24 hours, including a top-up with additional 6 µl of DNA library after the first 6 hours of the sequencing run.

    Article Title: De novo yeast genome assemblies from MinION, PacBio and MiSeq platforms
    Article Snippet: Two ug of genomic DNA was sheared to approximately 18,000 bp by centrifugation at 4000 rpm in a gTUBE. .. Sequencing libraries were prepared according to the SQK-MAP006 or SQK-NSK007 Sequencing Kit protocol, including the NEBNext FFPE DNA repair step. .. The sequencing mix was prepared with 6 uL of the DNA library, water, the Fuel Mix and the running buffer according to the SQK-MAP006 or the SQK-MAP007 protocols.

    Article Title: Hybrid genome assembly and annotation of Paenibacillus pasadenensis strain R16 reveals insights on endophytic life style and antifungal activity
    Article Snippet: Paragraph title: ONT libraries preparation and sequencing ... DNA nicks were repaired using the FFPE DNA repair mix (New England Biolabs, NEB) and End-repair and dA-tailing were performed using the Ultra II End Prep Module (New Englands Biolabs, NEB Ipswich, USA) according to manufacturer’s instructions.

    Article Title: Highly Contiguous Genome Assemblies of 15 Drosophila Species Generated Using Nanopore Sequencing
    Article Snippet: Paragraph title: Nanopore library preparation, sequencing, and base calling ... For 4 of 21 library preps (Table S1), the FFPE repair and dA-Tailing steps were combined in the following reaction mix: 46.5 µL of genomic DNA in TE, 3.5 µL of UltraII EP Buffer (NEB), 3.5 µL of FFPE DNA Repair Buffer (NEB), 3 µL of UltraII EP Enzyme (NEB), 3 µL of FFPE Repair Mix (NEB), and 0.5 µL of 100x NAD+ (NEB).

    Article Title: In Situ Field Sequencing and Life Detection in Remote (79°26′N) Canadian High Arctic Permafrost Ice Wedge Microbial Communities
    Article Snippet: Paragraph title: MinION sequencing and analysis ... Briefly, 46 μL of DNA extract was used for a DNA repair step using NEBNext FFPE Repair Mix (New England Biolabs, Ipswich, MA), followed by clean-up using AMPure XP beads incubated with mixing at room temperature for 5 min.

    Article Title: Nanopore DNA Sequencing and Genome Assembly on the International Space Station
    Article Snippet: Paragraph title: MinION Library Preparation and Sequencing (Ground and ISS) ... Mixed DNA samples then underwent treatment to repair residual nicks, gaps, and blocked 3′ ends using Formalin-Fixed, Paraffin-Embedded (FFPE) DNA Repair Mix (New England Biolabs), with modifications to the manufacturer’s protocol to include a 0.5X Agencourt AMPure XP system magnetic bead clean-up (Beckman Coulter Genomics, Brea, CA) to remove small fragments of DNA.

    Article Title: Nanopore sequencing of complex genomic rearrangements in yeast reveals mechanisms of repeat-mediated double-strand break repair
    Article Snippet: Using the ONT (Oxford Nanopore Technologies) Ligation Sequencing Kit 1D (SQK-LSK108) in combination with the Native Barcoding Kit 1D (EXP-NBD103), 1.5 µg of purified DNA was used to construct barcoded sequencing libraries. .. All procedures recommended in the ONT-provided protocol were followed, including nick repair (NEBNext FFPE Repair mix, New England Biolabs).

    Article Title: De novo whole-genome assembly of a wild type yeast isolate using nanopore sequencing
    Article Snippet: Paragraph title: MinION library preparation, sequencing and quality control ... The sheared DNA was repaired using the FFPE Repair Mix, according to the manufacturer’s instructions (New England Biolabs).

    Article Title: Artifactual mutations resulting from DNA lesions limit detection levels in ultrasensitive sequencing applications
    Article Snippet: An elevated number of G- > T and G- > GT mutations (observed either as G- > T or C- > A or G- > GT or C- > CA, depending on which strand was used as the reference sequence) was observed with an effective mutation frequency of 1.5 × 10− 6 and 1.6 × 10− 4 in genomic DNA and the oxidized HSI_vector, respectively ( ). .. In order to overcome this issue, instead of Fpg, DNA repair mixes could be used, such as available for formalin fixed, paraffin embedded (FFPE) samples (e.g. NEBNext FFPE DNA Repair Mix from NEB) or forensic samples (e.g. PreCR Repair Mix ), which include Fpg and other enzymes for the repair of generated abasic sites generating an amplifiable DNA template.

    Article Title: Rapid de novo assembly of the European eel genome from nanopore sequencing reads
    Article Snippet: Paragraph title: MinION library preparation and sequencing ... The sheared DNA was repaired using the FFPE repair mix according to the manufacturer’s instructions (New England Biolabs, Ipswich, USA).

    Article Title: High contiguity Arabidopsis thaliana genome assembly with a single nanopore flow cell
    Article Snippet: Paragraph title: Oxford Nanopore MinION sequencing ... Briefly, approximately 2 μg of purified DNA, without exogenous shearing or size selection, was repaired with NEBNext FFPE Repair Mix for 60 min at 20 °C.

    Article Title: De novo assembly and annotation of three Leptosphaeria genomes using Oxford Nanopore MinION sequencing
    Article Snippet: MinION sequencing libraries were prepared according to the nanopore Sequencing Kit protocol SQK-MAP006 or the Low Input Expansion Pack protocol for genomic DNA. .. For some libraries, DNA fragments were repaired using the NEBNext FFPE Repair Mix (New England Biolabs).

    Article Title: Comparison of bacterial genome assembly software for MinION data and their applicability to medical microbiology
    Article Snippet: Paragraph title: MinION sequencing and bioinformatic analysis. ... Sample preparation was carried out using the Genomic DNA Sequencing Kit SQK-MAP-006 (Oxford Nanopore Technologies) following the manufacturers instructions, including the optional NEBNext FFPE DNA repair step (NEB).

    Binding Assay:

    Article Title: De Novo Assembly of a New Solanum pennellii Accession Using Nanopore Sequencing
    Article Snippet: Subsequently, up to 5 μg of DNA was used for NEBNext FFPE DNA Repair (New England Biolabs) in a total volume of 155 μL including 16.3 μL NEBNext FFPE DNA Repair Buffer and 5 μL NEBNext FFPE DNA Repair Mix. .. Subsequently, up to 5 μg of DNA was used for NEBNext FFPE DNA Repair (New England Biolabs) in a total volume of 155 μL including 16.3 μL NEBNext FFPE DNA Repair Buffer and 5 μL NEBNext FFPE DNA Repair Mix.

    Molecular Weight:

    Article Title: Biosynthesis of the antibiotic nonribosomal peptide penicillin in baker's yeast
    Article Snippet: Enriched assembly DNA for the multigene and single gene assemblies was quantified on a Qubit 2.0 fluorometer (Thermo Fisher Scientific) using a Qubit dsDNA HS Assay Kit (Thermo Fisher Scientific). .. The four samples were combined to a give an equimolar mix (assuming a molecular weight for each assembly based on the mean promoter length) with a total DNA content of 2.6 μg in 45 μl dH2 O. DNA underwent end repair using NEBNext FFPE DNA Repair Mix (M6630, New England Biolabs) according to the manufacturer's instructions. .. The repaired DNA was recovered using Agincourt AMPure XP beads (A63880, Beckman Coulter), washed twice in 200 μl 70% ethanol and eluted in 46 μl dH2 O. DNA was then dA-tailed using NEBNext Ultra II End Repair/dA-Tailing Module (E7546, New England Biolabs) according to the manufacturer's instructions and recovered using Agincourt AMPure XP beads as before, eluting in 30 μl dH2 O.

    Article Title: De novo whole-genome assembly of a wild type yeast isolate using nanopore sequencing
    Article Snippet: In short, high molecular weight DNA was sheared with a g-TUBE (Covaris) to an average fragment length of 20 kbp. .. The sheared DNA was repaired using the FFPE Repair Mix, according to the manufacturer’s instructions (New England Biolabs).

    Article Title: Rapid de novo assembly of the European eel genome from nanopore sequencing reads
    Article Snippet: Briefly, high molecular weight DNA was sheared with a g-TUBE (Covaris) to an average fragment length of 20 kbp. .. The sheared DNA was repaired using the FFPE repair mix according to the manufacturer’s instructions (New England Biolabs, Ipswich, USA).

    Mutagenesis:

    Article Title: Artifactual mutations resulting from DNA lesions limit detection levels in ultrasensitive sequencing applications
    Article Snippet: Fpg treatment completely eliminated G- > T/C- > A and G- > GT/C- > CA transversions on the oxidized HSI_vector and the mutation frequency was significantly reduced (P value = 1.8 × 10− 5 , Fisher’s exact test). .. In order to overcome this issue, instead of Fpg, DNA repair mixes could be used, such as available for formalin fixed, paraffin embedded (FFPE) samples (e.g. NEBNext FFPE DNA Repair Mix from NEB) or forensic samples (e.g. PreCR Repair Mix ), which include Fpg and other enzymes for the repair of generated abasic sites generating an amplifiable DNA template.

    Isolation:

    Article Title: Biosynthesis of the antibiotic nonribosomal peptide penicillin in baker's yeast
    Article Snippet: Restriction digest products ranging from 5,616 to 6,117 bp were isolated by agarose gel electrophoresis and purified using a QIAquick gel extraction kit (Qiagen). .. The four samples were combined to a give an equimolar mix (assuming a molecular weight for each assembly based on the mean promoter length) with a total DNA content of 2.6 μg in 45 μl dH2 O. DNA underwent end repair using NEBNext FFPE DNA Repair Mix (M6630, New England Biolabs) according to the manufacturer's instructions.

    Purification:

    Article Title: Nanopore sequencing of complex genomic rearrangements in yeast reveals mechanisms of repeat-mediated double-strand break repair
    Article Snippet: Using the ONT (Oxford Nanopore Technologies) Ligation Sequencing Kit 1D (SQK-LSK108) in combination with the Native Barcoding Kit 1D (EXP-NBD103), 1.5 µg of purified DNA was used to construct barcoded sequencing libraries. .. All procedures recommended in the ONT-provided protocol were followed, including nick repair (NEBNext FFPE Repair mix, New England Biolabs).

    Article Title: Biosynthesis of the antibiotic nonribosomal peptide penicillin in baker's yeast
    Article Snippet: Restriction digest products ranging from 2,062 to 2,229, 2,549 to 2,716 and 1,885 to 2,052 bp for the pcbC , pclA and penDE assemblies, respectively, were purified. .. The four samples were combined to a give an equimolar mix (assuming a molecular weight for each assembly based on the mean promoter length) with a total DNA content of 2.6 μg in 45 μl dH2 O. DNA underwent end repair using NEBNext FFPE DNA Repair Mix (M6630, New England Biolabs) according to the manufacturer's instructions.

    Article Title: De Novo Assembly of a New Solanum pennellii Accession Using Nanopore Sequencing
    Article Snippet: Subsequently, up to 5 μg of DNA was used for NEBNext FFPE DNA Repair (New England Biolabs) in a total volume of 155 μL including 16.3 μL NEBNext FFPE DNA Repair Buffer and 5 μL NEBNext FFPE DNA Repair Mix. .. For NEBNext Ultra II End Repair/dA-Tailing treatment (New England Biolabs), 100 μL of FFPE repaired DNA, together with 14.0 μL NEBNext Ultra II End Prep Reaction Buffer and 6 μL NEBNext Ultra II End Prep Enzyme Mix, was incubated for 30 min at 20°C followed by 20 min at 65°C and 4°C until further processing.

    Article Title: High contiguity Arabidopsis thaliana genome assembly with a single nanopore flow cell
    Article Snippet: The purified DNA was then prepared for sequencing following the protocol in the genomic sequencing kit SQK-LSK108 (ONT, Oxford, UK). .. Briefly, approximately 2 μg of purified DNA, without exogenous shearing or size selection, was repaired with NEBNext FFPE Repair Mix for 60 min at 20 °C. .. The DNA was purified with 0.5× Ampure XP beads (Beckman Coulter).

    Agarose Gel Electrophoresis:

    Article Title: Biosynthesis of the antibiotic nonribosomal peptide penicillin in baker's yeast
    Article Snippet: Restriction digest products ranging from 5,616 to 6,117 bp were isolated by agarose gel electrophoresis and purified using a QIAquick gel extraction kit (Qiagen). .. The four samples were combined to a give an equimolar mix (assuming a molecular weight for each assembly based on the mean promoter length) with a total DNA content of 2.6 μg in 45 μl dH2 O. DNA underwent end repair using NEBNext FFPE DNA Repair Mix (M6630, New England Biolabs) according to the manufacturer's instructions.

    Plasmid Preparation:

    Article Title: Biosynthesis of the antibiotic nonribosomal peptide penicillin in baker's yeast
    Article Snippet: To enrich for the penicillin pathway assembly DNA and remove assembly vector backbone DNA, the multigene assembly library was digested with EcoRI and AlwNI. .. The four samples were combined to a give an equimolar mix (assuming a molecular weight for each assembly based on the mean promoter length) with a total DNA content of 2.6 μg in 45 μl dH2 O. DNA underwent end repair using NEBNext FFPE DNA Repair Mix (M6630, New England Biolabs) according to the manufacturer's instructions.

    Article Title: Artifactual mutations resulting from DNA lesions limit detection levels in ultrasensitive sequencing applications
    Article Snippet: To address this, we analyzed in total ∼9500 smPCRs, each one representing an amplifiable human genome and ∼400 smPCR reactions of one oxidized control plasmid (HSI_vector without synthetic insert). .. In order to overcome this issue, instead of Fpg, DNA repair mixes could be used, such as available for formalin fixed, paraffin embedded (FFPE) samples (e.g. NEBNext FFPE DNA Repair Mix from NEB) or forensic samples (e.g. PreCR Repair Mix ), which include Fpg and other enzymes for the repair of generated abasic sites generating an amplifiable DNA template.

    Software:

    Article Title: Antimicrobial resistance prediction and phylogenetic analysis of Neisseria gonorrhoeae isolates using the Oxford Nanopore MinION sequencer
    Article Snippet: Sequencing libraries were prepared according to the 2D library preparation protocol with the SQK-LSK208 2D ligation kit (Oxford Nanopore Technologies), including the DNA repair step with the NEBNext FFPE DNA repair module (New England Biolabs, Ipswich, MA, USA). .. The sequencing libraries were purified using MyOne C1 beads (Thermo Fisher Scientific), and 6 μl of sequencing library were loaded onto a R9.4 SpotON flow cell and sequenced with the MinION Mk 1B sequencing device (Oxford Nanopore Technologies) for 24 hours, including a top-up with additional 6 µl of DNA library after the first 6 hours of the sequencing run.

    Selection:

    Article Title: De Novo Assembly of a New Solanum pennellii Accession Using Nanopore Sequencing
    Article Snippet: Enrichment for long fragments was achieved by BluePippin size selection (Sage Science). .. Subsequently, up to 5 μg of DNA was used for NEBNext FFPE DNA Repair (New England Biolabs) in a total volume of 155 μL including 16.3 μL NEBNext FFPE DNA Repair Buffer and 5 μL NEBNext FFPE DNA Repair Mix.

    Article Title: High contiguity Arabidopsis thaliana genome assembly with a single nanopore flow cell
    Article Snippet: The purified DNA was then prepared for sequencing following the protocol in the genomic sequencing kit SQK-LSK108 (ONT, Oxford, UK). .. Briefly, approximately 2 μg of purified DNA, without exogenous shearing or size selection, was repaired with NEBNext FFPE Repair Mix for 60 min at 20 °C. .. The DNA was purified with 0.5× Ampure XP beads (Beckman Coulter).

    Sample Prep:

    Article Title: Comparison of bacterial genome assembly software for MinION data and their applicability to medical microbiology
    Article Snippet: DNA was extracted using the QiaAMP DNA Mini kit (Qiagen), and quantified using the Qubit fluorimeter (Life Technologies). .. Sample preparation was carried out using the Genomic DNA Sequencing Kit SQK-MAP-006 (Oxford Nanopore Technologies) following the manufacturers instructions, including the optional NEBNext FFPE DNA repair step (NEB). .. A 6 µl aliquot of pre-sequencing mix was combined with 4 µl Fuel Mix (Oxford Nanopore), 75 µl running buffer (Oxford Nanopore) and 66 µl water and added to the flow cell.

    Ethanol Precipitation:

    Article Title: High contiguity Arabidopsis thaliana genome assembly with a single nanopore flow cell
    Article Snippet: Briefly, approximately 2 μg of purified DNA, without exogenous shearing or size selection, was repaired with NEBNext FFPE Repair Mix for 60 min at 20 °C. .. Briefly, approximately 2 μg of purified DNA, without exogenous shearing or size selection, was repaired with NEBNext FFPE Repair Mix for 60 min at 20 °C.

    Produced:

    Article Title: Hybrid genome assembly and annotation of Paenibacillus pasadenensis strain R16 reveals insights on endophytic life style and antifungal activity
    Article Snippet: DNA libraries for Oxford Nanopore sequencing were produced starting from 1.8 μg DNA that was randomly sheared to an average length of ~15Kbps using g-TUBEs (Covaris) centrifuged at 5000 rpm (Centrifuge 5424, Eppendorf) for 60 seconds. .. DNA nicks were repaired using the FFPE DNA repair mix (New England Biolabs, NEB) and End-repair and dA-tailing were performed using the Ultra II End Prep Module (New Englands Biolabs, NEB Ipswich, USA) according to manufacturer’s instructions.

    Marker:

    Article Title: De Novo Assembly of a New Solanum pennellii Accession Using Nanopore Sequencing
    Article Snippet: Approximately 35 μL per lane was run together with an S1 marker reference lane on a 0.75% Agarose Cassette (Biozyme) using the high pass protocol and a collection window of 12 to 80 kb or 15 to 80 kb. .. Subsequently, up to 5 μg of DNA was used for NEBNext FFPE DNA Repair (New England Biolabs) in a total volume of 155 μL including 16.3 μL NEBNext FFPE DNA Repair Buffer and 5 μL NEBNext FFPE DNA Repair Mix.

    Gel Extraction:

    Article Title: Biosynthesis of the antibiotic nonribosomal peptide penicillin in baker's yeast
    Article Snippet: Restriction digest products ranging from 5,616 to 6,117 bp were isolated by agarose gel electrophoresis and purified using a QIAquick gel extraction kit (Qiagen). .. The four samples were combined to a give an equimolar mix (assuming a molecular weight for each assembly based on the mean promoter length) with a total DNA content of 2.6 μg in 45 μl dH2 O. DNA underwent end repair using NEBNext FFPE DNA Repair Mix (M6630, New England Biolabs) according to the manufacturer's instructions.

    Nanopore Sequencing:

    Article Title: De novo yeast genome assemblies from MinION, PacBio and MiSeq platforms
    Article Snippet: Paragraph title: Library preparations for Oxford Nanopore sequencing ... Sequencing libraries were prepared according to the SQK-MAP006 or SQK-NSK007 Sequencing Kit protocol, including the NEBNext FFPE DNA repair step.

    Article Title: Nanopore sequencing of complex genomic rearrangements in yeast reveals mechanisms of repeat-mediated double-strand break repair
    Article Snippet: Paragraph title: Nanopore sequencing ... All procedures recommended in the ONT-provided protocol were followed, including nick repair (NEBNext FFPE Repair mix, New England Biolabs).

    Article Title: Biosynthesis of the antibiotic nonribosomal peptide penicillin in baker's yeast
    Article Snippet: Paragraph title: Nanopore sequencing of library construction ... The four samples were combined to a give an equimolar mix (assuming a molecular weight for each assembly based on the mean promoter length) with a total DNA content of 2.6 μg in 45 μl dH2 O. DNA underwent end repair using NEBNext FFPE DNA Repair Mix (M6630, New England Biolabs) according to the manufacturer's instructions.

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  • 79
    New England Biolabs dna repair procedure
    CN profiles for two breast pre-cancerous samples (P1 ( a , b ) and P2 ( c , d )). a , c MIP arrays in the range of 40–80 ng input of <t>DNA.</t> b , d LC <t>WGS</t> from 5 ng input of DNA. Each data point in ( b ) and ( d ) represents normalized read count ratios from a 50 kb window. Segments were removed from highly repetitive or problematic regions [ 8 ]
    Dna Repair Procedure, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 79/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/dna repair procedure/product/New England Biolabs
    Average 79 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    dna repair procedure - by Bioz Stars, 2019-10
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    78
    New England Biolabs dna repair endonucleases
    Acrylamide and glycidamide induces <t>DNA</t> damage in the male germ line. DNA damage was assessed in spermatocytes treated with acrylamide or glycidamide using the comet assay in the presence or absence of <t>FPG.</t> (A) Representative comet images from control, acrylamide (1 µM, 18 h), glycidamide (0.5 µM, 18 h) and H 2 O 2 (500 µM, 5 min) treated spermatocytes. (B) The average Tail DNA % was assessed for each sample and in the absence of FPG, a modest increase in Tail DNA % was observed in spermatocytes treated with glycidamide ( F 7,663 = 61.7, * p
    Dna Repair Endonucleases, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 78/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    New England Biolabs 3 end
    LITE-Seq analysis. a Germline tissue used in this study. Whole gonads were isolated from adult hermaphrodite (XX) and male (XO) worms. Female germline samples were generated by manual removal of the spermatheca. Developmentally staged samples were obtained by microdissection of the distal mitotic proliferation zone, the meiotic pachytene region, and cellularized oocytes. b LITE-Seq protocol. Extracts of total RNA spiked with ERCC RNA standards were reverse transcribed using an anchored hairpin polydT primer. cDNAs were PCR amplified using a biotinylated half-hairpin primer, fragmented, and subjected to <t>3′-end</t> capture with streptavidin beads. Samples were barcoded, pooled, and subjected to paired-end sequencing using the standard Illumina Read 1 primer and a custom Read 2 primer that initiates sequencing immediately upstream of the polyA tail. c Mapping and quantification of transcript 3′ ends. Paired-end reads were mapped to the C. elegans reference genome (WS250) and assigned to genes with overlapping exons or to the closest gene within 1500 nt upstream. Representative 3′ CPA sites (arrowheads) and isoform abundance were determined by clustering 3′ end reads within ±12 nucleotides of the position with the most supported reads
    3 End, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    CN profiles for two breast pre-cancerous samples (P1 ( a , b ) and P2 ( c , d )). a , c MIP arrays in the range of 40–80 ng input of DNA. b , d LC WGS from 5 ng input of DNA. Each data point in ( b ) and ( d ) represents normalized read count ratios from a 50 kb window. Segments were removed from highly repetitive or problematic regions [ 8 ]

    Journal: Genome Medicine

    Article Title: Copy number analysis by low coverage whole genome sequencing using ultra low-input DNA from formalin-fixed paraffin embedded tumor tissue

    doi: 10.1186/s13073-016-0375-z

    Figure Lengend Snippet: CN profiles for two breast pre-cancerous samples (P1 ( a , b ) and P2 ( c , d )). a , c MIP arrays in the range of 40–80 ng input of DNA. b , d LC WGS from 5 ng input of DNA. Each data point in ( b ) and ( d ) represents normalized read count ratios from a 50 kb window. Segments were removed from highly repetitive or problematic regions [ 8 ]

    Article Snippet: In parallel, we assessed whether a DNA repair procedure (NEB Next) could improve LC WGS CNA detection performance.

    Techniques: Liquid Chromatography

    Experimental design testing a low-input method (NEBNext Ultra II) on copy number detection of two FFPE MCC samples by LC WGS as well as the effect of WGA with or without NEB DNA repair treatment

    Journal: Genome Medicine

    Article Title: Copy number analysis by low coverage whole genome sequencing using ultra low-input DNA from formalin-fixed paraffin embedded tumor tissue

    doi: 10.1186/s13073-016-0375-z

    Figure Lengend Snippet: Experimental design testing a low-input method (NEBNext Ultra II) on copy number detection of two FFPE MCC samples by LC WGS as well as the effect of WGA with or without NEB DNA repair treatment

    Article Snippet: In parallel, we assessed whether a DNA repair procedure (NEB Next) could improve LC WGS CNA detection performance.

    Techniques: Formalin-fixed Paraffin-Embedded, Liquid Chromatography, Whole Genome Amplification

    CN profiles for two breast tumor samples (LP S1 ( a , b ) and LP S4 ( c , d )). a , c Low coverage WGS from 5 ng input of DNA. b , d MIP arrays in the range of 80–100 ng input of DNA. Each data point in ( a ) and ( c ) represents normalized read count ratios from a 50 kb window. Segments were removed from highly repetitive or problematic regions [ 8 ]

    Journal: Genome Medicine

    Article Title: Copy number analysis by low coverage whole genome sequencing using ultra low-input DNA from formalin-fixed paraffin embedded tumor tissue

    doi: 10.1186/s13073-016-0375-z

    Figure Lengend Snippet: CN profiles for two breast tumor samples (LP S1 ( a , b ) and LP S4 ( c , d )). a , c Low coverage WGS from 5 ng input of DNA. b , d MIP arrays in the range of 80–100 ng input of DNA. Each data point in ( a ) and ( c ) represents normalized read count ratios from a 50 kb window. Segments were removed from highly repetitive or problematic regions [ 8 ]

    Article Snippet: In parallel, we assessed whether a DNA repair procedure (NEB Next) could improve LC WGS CNA detection performance.

    Techniques:

    Acrylamide and glycidamide induces DNA damage in the male germ line. DNA damage was assessed in spermatocytes treated with acrylamide or glycidamide using the comet assay in the presence or absence of FPG. (A) Representative comet images from control, acrylamide (1 µM, 18 h), glycidamide (0.5 µM, 18 h) and H 2 O 2 (500 µM, 5 min) treated spermatocytes. (B) The average Tail DNA % was assessed for each sample and in the absence of FPG, a modest increase in Tail DNA % was observed in spermatocytes treated with glycidamide ( F 7,663 = 61.7, * p

    Journal: PLoS ONE

    Article Title: Mouse Spermatocytes Express CYP2E1 and Respond to Acrylamide Exposure

    doi: 10.1371/journal.pone.0094904

    Figure Lengend Snippet: Acrylamide and glycidamide induces DNA damage in the male germ line. DNA damage was assessed in spermatocytes treated with acrylamide or glycidamide using the comet assay in the presence or absence of FPG. (A) Representative comet images from control, acrylamide (1 µM, 18 h), glycidamide (0.5 µM, 18 h) and H 2 O 2 (500 µM, 5 min) treated spermatocytes. (B) The average Tail DNA % was assessed for each sample and in the absence of FPG, a modest increase in Tail DNA % was observed in spermatocytes treated with glycidamide ( F 7,663 = 61.7, * p

    Article Snippet: DNA repair endonucleases, formamidopyrimidine-DNA glycosylase (FPG) and 8-oxoguanine DNA glycosylase (hOGG1) were purchased from New England Biolabs Inc. (Arundel, Qld).

    Techniques: Single Cell Gel Electrophoresis

    Acrylamide is metabolized in spermatocytes to glycidamide via CYP2E1. The Isolated spermatocytes were treated with acrylamide (1 µM, 18 h), glycidamide (0.5 µM, 18 h), resveratrol (0.1 µM, 18 h), or a combination of acrylamide (1 µM) and resveratrol (0.1 µM, 18 h) or glycidamide (0.5 µM) and resveratrol (0.1 µM, 18 h). DNA damage was assessed by the comet assay modified by the addition of the FPG or hOGG1 enzyme. Significant levels of DNA damage were detected in the acrylamide treated spermatocytes in the presence of FPG ( F 13,877 = 92.5, *** p

    Journal: PLoS ONE

    Article Title: Mouse Spermatocytes Express CYP2E1 and Respond to Acrylamide Exposure

    doi: 10.1371/journal.pone.0094904

    Figure Lengend Snippet: Acrylamide is metabolized in spermatocytes to glycidamide via CYP2E1. The Isolated spermatocytes were treated with acrylamide (1 µM, 18 h), glycidamide (0.5 µM, 18 h), resveratrol (0.1 µM, 18 h), or a combination of acrylamide (1 µM) and resveratrol (0.1 µM, 18 h) or glycidamide (0.5 µM) and resveratrol (0.1 µM, 18 h). DNA damage was assessed by the comet assay modified by the addition of the FPG or hOGG1 enzyme. Significant levels of DNA damage were detected in the acrylamide treated spermatocytes in the presence of FPG ( F 13,877 = 92.5, *** p

    Article Snippet: DNA repair endonucleases, formamidopyrimidine-DNA glycosylase (FPG) and 8-oxoguanine DNA glycosylase (hOGG1) were purchased from New England Biolabs Inc. (Arundel, Qld).

    Techniques: Isolation, Single Cell Gel Electrophoresis, Modification

    LITE-Seq analysis. a Germline tissue used in this study. Whole gonads were isolated from adult hermaphrodite (XX) and male (XO) worms. Female germline samples were generated by manual removal of the spermatheca. Developmentally staged samples were obtained by microdissection of the distal mitotic proliferation zone, the meiotic pachytene region, and cellularized oocytes. b LITE-Seq protocol. Extracts of total RNA spiked with ERCC RNA standards were reverse transcribed using an anchored hairpin polydT primer. cDNAs were PCR amplified using a biotinylated half-hairpin primer, fragmented, and subjected to 3′-end capture with streptavidin beads. Samples were barcoded, pooled, and subjected to paired-end sequencing using the standard Illumina Read 1 primer and a custom Read 2 primer that initiates sequencing immediately upstream of the polyA tail. c Mapping and quantification of transcript 3′ ends. Paired-end reads were mapped to the C. elegans reference genome (WS250) and assigned to genes with overlapping exons or to the closest gene within 1500 nt upstream. Representative 3′ CPA sites (arrowheads) and isoform abundance were determined by clustering 3′ end reads within ±12 nucleotides of the position with the most supported reads

    Journal: Genome Biology

    Article Title: Developmental dynamics of gene expression and alternative polyadenylation in the Caenorhabditis elegans germline

    doi: 10.1186/s13059-017-1369-x

    Figure Lengend Snippet: LITE-Seq analysis. a Germline tissue used in this study. Whole gonads were isolated from adult hermaphrodite (XX) and male (XO) worms. Female germline samples were generated by manual removal of the spermatheca. Developmentally staged samples were obtained by microdissection of the distal mitotic proliferation zone, the meiotic pachytene region, and cellularized oocytes. b LITE-Seq protocol. Extracts of total RNA spiked with ERCC RNA standards were reverse transcribed using an anchored hairpin polydT primer. cDNAs were PCR amplified using a biotinylated half-hairpin primer, fragmented, and subjected to 3′-end capture with streptavidin beads. Samples were barcoded, pooled, and subjected to paired-end sequencing using the standard Illumina Read 1 primer and a custom Read 2 primer that initiates sequencing immediately upstream of the polyA tail. c Mapping and quantification of transcript 3′ ends. Paired-end reads were mapped to the C. elegans reference genome (WS250) and assigned to genes with overlapping exons or to the closest gene within 1500 nt upstream. Representative 3′ CPA sites (arrowheads) and isoform abundance were determined by clustering 3′ end reads within ±12 nucleotides of the position with the most supported reads

    Article Snippet: While still attached to beads, the captured cDNA was end-repaired and an adenosine was added to the 3′ end (NEBNext Ultra End Repair/dA-Tailing module).

    Techniques: Isolation, Generated, Laser Capture Microdissection, Polymerase Chain Reaction, Amplification, Sequencing