thermopol ii reaction buffer  (New England Biolabs)


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    Name:
    ThermoPol II Mg free Reaction Buffer Pack
    Description:
    ThermoPol II Mg free Reaction Buffer Pack 6 0 ml
    Catalog Number:
    b9005s
    Price:
    22
    Size:
    6 0 ml
    Category:
    DNA Analysis
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    New England Biolabs thermopol ii reaction buffer
    ThermoPol II Mg free Reaction Buffer Pack
    ThermoPol II Mg free Reaction Buffer Pack 6 0 ml
    https://www.bioz.com/result/thermopol ii reaction buffer/product/New England Biolabs
    Average 90 stars, based on 3 article reviews
    Price from $9.99 to $1999.99
    thermopol ii reaction buffer - by Bioz Stars, 2020-02
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    Centrifugation:

    Article Title: Aerial Dissemination of Clostridium difficile spores
    Article Snippet: Strains were cultured in Brain Heart Infusion Broth supplemented with 0.5% yeast extract and 0.5 mg of haemin, incubated anaerobically at 37°C for 24 h. Cells from 1.5 mL of culture were pelleted by centrifugation. .. A 25 μl reaction mix contained 1× Thermopol II Buffer (New England Biolabs), 200 mM of each dNTP, 2.25 mM of MgSO4 , 50 pmol of each primer (Sigma-Genosys), 2.5 units of Taq polymerase (New England Biolabs) and 2.5 μl of DNA template.

    Amplification:

    Article Title: Mapping and Validation of the Major Sex-Determining Region in Nile Tilapia (Oreochromis niloticus L.) Using RAD Sequencing
    Article Snippet: The RAD specific P1 and P2 paired-end adapters and library amplification PCR primer sequences used in this study are detailed in Baxter et al. . .. Individual specific P1 adapters, each with a unique 5 bp barcode , were ligated to the Sbf I digested DNA at 22°C for 60 minutes by adding 1.8/0.6 µL 100 nmol/L P1 adapter, 0.45/0.15 µL 100 mmol/L rATP (Promega), 0.75/0.25 µL 10× Reaction Buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes made up to 45/15 µL with nuclease-free water for each parental/offspring sample.

    Article Title: Genome-wide association and genomic prediction of resistance to viral nervous necrosis in European sea bass (Dicentrarchus labrax) using RAD sequencing
    Article Snippet: Individual specific P1 adapters, each with a unique 5–7 bp barcode, were ligated to the Sbf I-digested DNA at 20 °C for 60 min by adding 1.8/0.6 µL of 100 nmol/L P1 adapter, 0.45/0.15 µL of 100 mmol/L rATP (Promega), 0.75/0.25 µL 10 × reaction buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes for each parental/offspring sample were completed to 45/15 µL with nuclease-free water. .. Shearing (Covaris S2 sonication) and initial size selection (300 to 600 bp) by agarose gel separation was followed by gel purification, end repair, dA overhang addition, P2 (individual specific adapters) paired-end adapter ligation, library amplification, as in the original RAD protocol [ , ].

    Article Title: Accuracy of Genomic Evaluations of Juvenile Growth Rate in Common Carp (Cyprinus carpio) Using Genotyping by Sequencing
    Article Snippet: The RAD-specific P1 and P2 paired-end adapters and library amplification PCR primer sequences used in this study are detailed in . .. The reactions (12 μL final volumes) were then heat inactivated at 65°C for 20 min. Individual-specific P1 adapters, each with a unique 5 bp barcode, were ligated to the Sbf I-digested DNA at 20°C for 60 min by adding 1.8/0.6 μL 100 nmol/L P1 adapter, 0.45/0.15 μL 100 mmol/L rATP (Promega), 0.75/0.25 μL 10× Reaction Buffer 2 (NEB), 0.36/0.12 μL T4 ligase (NEB, 2 M U/mL), and reaction volumes made up to 45/15 μL with nuclease-free water for each parental/offspring sample.

    Article Title: 5-Aza-2?-deoxycytidine Sensitizes Busulfan-resistant Myeloid Leukemia Cells By Regulating Expression of Genes Involved in Cell Cycle Checkpoint and Apoptosis
    Article Snippet: Methylation-specific PCR (MSP) was performed using 30 ng bisulfite-modified DNA, 0.5 μM of each primer , 1 mM each of dATP, dCTP, dGTP and dTTP, 5 mM MgSO4 , and Taq DNA polymerase with ThermoPol II buffer (New England Biolabs, Ipswich, MA). .. The amplification method included initial heating at 95°C for 3 min, followed by 30 cycles of 95°C for 30 s, 57°C or 65°C for 30 s depending on the primers (see ), and 72°C for 33 s, followed by extension at 72°C for 10 min. MSP products were visualized on a 2.5% agarose gel and DNA band intensities were analyzed using Quant One software (Bio-Rad, Hercules, CA).

    Article Title: Admixture in Humans of Two Divergent Plasmodium knowlesi Populations Associated with Different Macaque Host Species
    Article Snippet: PCR and genotyping protocols Genotyping of each microsatellite locus was performed using a hemi-nested protocol with a fluorescent dye-labelled inner primer during the second round PCR amplification (primers listed in ). .. Both first and second round PCR amplifications were conducted in individual tubes or wells for each locus, in 11 μl reaction volume containing 0.2 mM each dNTP (Bioline, UK), 2 mM MgSO4 , 1X ThermoPol II reaction buffer (NEB, UK), 0.275 U Taq DNA polymerase (NEB, UK), 0.1 μM of each forward and reverse primer, and 1 μl sample DNA template.

    SYBR Green Assay:

    Article Title: A mechanism for ramified rolling circle amplification
    Article Snippet: RAM reactions RAM reaction time-courses were done in RAM buffer 1: 1× New England Biolabs (NEB) Thermopol II buffer, 2 mM MgSO4, 0.2 mM each dNTP; 2% DMSO; 0.75 uM forward primer Cpr4PrFwd80_20; 1 uM Cpr4PrRvs09_19; 50 molecules circularized template Cpr4Stchy_ch154 per reaction; Bst DNA polymerase, large fragment (NEB), 0.13 units/ul. .. Real-time RAM reactions were done in RAM buffer 2: [1× New England Biolabs (NEB) Thermopol II buffer, 2 mM MgSO4, 0.2 mM each dNTP; 5% DMSO; 40 mM NaCl; 0.6× SYBR-Green (Molecular Probes), 1 nM fluorescein (BioRad), 0.75 uM forward primer Cpr2Fwd47_20; 1 uM Cpr3Rvs50_19; circularized template Cpr3Stchy_ch381 in the range of 0 (zero) to 107 molecules per reaction; Bst DNA polymerase, large fragment (NEB), 0.13 units/ul.

    Article Title: A mechanism for ramified rolling circle amplification
    Article Snippet: .. Real-time RAM reactions were done in RAM buffer 2: [1× New England Biolabs (NEB) Thermopol II buffer, 2 mM MgSO4, 0.2 mM each dNTP; 5% DMSO; 40 mM NaCl; 0.6× SYBR-Green (Molecular Probes), 1 nM fluorescein (BioRad), 0.75 uM forward primer Cpr2Fwd47_20; 1 uM Cpr3Rvs50_19; circularized template Cpr3Stchy_ch381 in the range of 0 (zero) to 107 molecules per reaction; Bst DNA polymerase, large fragment (NEB), 0.13 units/ul. ..

    Article Title: Generalized stochastic profiling of transcriptional regulatory heterogeneities in tissues, tumors, and cultured cells
    Article Snippet: Terminal transferase (Roche #03333574001, 400 U μl−1 ) 10× ThermoPol buffer (New England Biolabs #B9005S) 100 mM MgSO4 (New England Biolabs #B1003S) BSA (Roche #10711454001) Roche Taq polymerase (Roche #04728858001, 5 U μl−1 ) AL1 primer (200 nmol synthesis from IDT or MWG): ATTGGATCCAGGCCGCTCTGGACAAAATATGAATTCTTTTTTTTTTTTTTTTTTTTT TTT ▲ CRITICAL Long desalted primers will have varying purities depending on the manufacturer. .. 100× SYBR Green I (Invitrogen #S7563 diluted 100-fold in DMSO to 100×) 50 mM aminoallyl-dUTP (Ambion #1103015) High-Fidelity polymerase (Roche #11732650001, 3.5 U μl−1 ) 10× High-Fidelity PCR buffer without Mg2+ (included with Roche #11759175001) PureLink PCR Purification Kit (Invitrogen #46-6056) NaOAc (Calbiochem #567418) 20 mg/ml glycogen (Invitrogen #10814-010) NaHCO3 (Acros #21712500) Alexa Fluor 555 reactive dye decapack (Invitrogen # ) GEX hybridization buffer (included with Illumina #BD-103-0204 or equivalent)

    Incubation:

    Article Title: Aerial Dissemination of Clostridium difficile spores
    Article Snippet: Strains were cultured in Brain Heart Infusion Broth supplemented with 0.5% yeast extract and 0.5 mg of haemin, incubated anaerobically at 37°C for 24 h. Cells from 1.5 mL of culture were pelleted by centrifugation. .. A 25 μl reaction mix contained 1× Thermopol II Buffer (New England Biolabs), 200 mM of each dNTP, 2.25 mM of MgSO4 , 50 pmol of each primer (Sigma-Genosys), 2.5 units of Taq polymerase (New England Biolabs) and 2.5 μl of DNA template.

    Modification:

    Article Title: 5-Aza-2?-deoxycytidine Sensitizes Busulfan-resistant Myeloid Leukemia Cells By Regulating Expression of Genes Involved in Cell Cycle Checkpoint and Apoptosis
    Article Snippet: Bisulfite modification of the genomic DNA and its purification was performed using a MethylDetector kit (Active Motif, Carlsbad, CA). .. Methylation-specific PCR (MSP) was performed using 30 ng bisulfite-modified DNA, 0.5 μM of each primer , 1 mM each of dATP, dCTP, dGTP and dTTP, 5 mM MgSO4 , and Taq DNA polymerase with ThermoPol II buffer (New England Biolabs, Ipswich, MA).

    Hybridization:

    Article Title: Generalized stochastic profiling of transcriptional regulatory heterogeneities in tissues, tumors, and cultured cells
    Article Snippet: Terminal transferase (Roche #03333574001, 400 U μl−1 ) 10× ThermoPol buffer (New England Biolabs #B9005S) 100 mM MgSO4 (New England Biolabs #B1003S) BSA (Roche #10711454001) Roche Taq polymerase (Roche #04728858001, 5 U μl−1 ) AL1 primer (200 nmol synthesis from IDT or MWG): ATTGGATCCAGGCCGCTCTGGACAAAATATGAATTCTTTTTTTTTTTTTTTTTTTTT TTT ▲ CRITICAL Long desalted primers will have varying purities depending on the manufacturer. .. 100× SYBR Green I (Invitrogen #S7563 diluted 100-fold in DMSO to 100×) 50 mM aminoallyl-dUTP (Ambion #1103015) High-Fidelity polymerase (Roche #11732650001, 3.5 U μl−1 ) 10× High-Fidelity PCR buffer without Mg2+ (included with Roche #11759175001) PureLink PCR Purification Kit (Invitrogen #46-6056) NaOAc (Calbiochem #567418) 20 mg/ml glycogen (Invitrogen #10814-010) NaHCO3 (Acros #21712500) Alexa Fluor 555 reactive dye decapack (Invitrogen # ) GEX hybridization buffer (included with Illumina #BD-103-0204 or equivalent)

    Southern Blot:

    Article Title: Susceptibility to liver fibrosis in mice expressing a connective tissue growth factor transgene in hepatocytes
    Article Snippet: Paragraph title: Southern Blot ... Genomic DNA (5 μg) from mice tails was digested in a 40 μl final volume with 20U Stu I in Reaction buffer II (New England Biolabs, Ipswich, MA).

    Ligation:

    Article Title: Mapping and Validation of the Major Sex-Determining Region in Nile Tilapia (Oreochromis niloticus L.) Using RAD Sequencing
    Article Snippet: Individual specific P1 adapters, each with a unique 5 bp barcode , were ligated to the Sbf I digested DNA at 22°C for 60 minutes by adding 1.8/0.6 µL 100 nmol/L P1 adapter, 0.45/0.15 µL 100 mmol/L rATP (Promega), 0.75/0.25 µL 10× Reaction Buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes made up to 45/15 µL with nuclease-free water for each parental/offspring sample. .. Following heat inactivation at 65°C for 20 minutes, the ligation reactions were slowly cooled to room temperature (over 1 hour) then combined in appropriate multiplex pools ( ).

    Article Title: Genome-wide association and genomic prediction of resistance to viral nervous necrosis in European sea bass (Dicentrarchus labrax) using RAD sequencing
    Article Snippet: Individual specific P1 adapters, each with a unique 5–7 bp barcode, were ligated to the Sbf I-digested DNA at 20 °C for 60 min by adding 1.8/0.6 µL of 100 nmol/L P1 adapter, 0.45/0.15 µL of 100 mmol/L rATP (Promega), 0.75/0.25 µL 10 × reaction buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes for each parental/offspring sample were completed to 45/15 µL with nuclease-free water. .. Following heat inactivation at 65 °C for 20 min, ligation reactions were slowly cooled down to room temperature (over 1 h), then combined in appropriate multiplex pools.

    Article Title: Accuracy of Genomic Evaluations of Juvenile Growth Rate in Common Carp (Cyprinus carpio) Using Genotyping by Sequencing
    Article Snippet: The reactions (12 μL final volumes) were then heat inactivated at 65°C for 20 min. Individual-specific P1 adapters, each with a unique 5 bp barcode, were ligated to the Sbf I-digested DNA at 20°C for 60 min by adding 1.8/0.6 μL 100 nmol/L P1 adapter, 0.45/0.15 μL 100 mmol/L rATP (Promega), 0.75/0.25 μL 10× Reaction Buffer 2 (NEB), 0.36/0.12 μL T4 ligase (NEB, 2 M U/mL), and reaction volumes made up to 45/15 μL with nuclease-free water for each parental/offspring sample. .. Following heat inactivation at 65°C for 20 min, the ligation reactions were slowly cooled to room temperature (over 1 h) then combined in appropriate multiplex pools.

    Cell Culture:

    Article Title: Aerial Dissemination of Clostridium difficile spores
    Article Snippet: Strains were cultured in Brain Heart Infusion Broth supplemented with 0.5% yeast extract and 0.5 mg of haemin, incubated anaerobically at 37°C for 24 h. Cells from 1.5 mL of culture were pelleted by centrifugation. .. A 25 μl reaction mix contained 1× Thermopol II Buffer (New England Biolabs), 200 mM of each dNTP, 2.25 mM of MgSO4 , 50 pmol of each primer (Sigma-Genosys), 2.5 units of Taq polymerase (New England Biolabs) and 2.5 μl of DNA template.

    Sequencing:

    Article Title: Mapping and Validation of the Major Sex-Determining Region in Nile Tilapia (Oreochromis niloticus L.) Using RAD Sequencing
    Article Snippet: Paragraph title: RAD Library Preparation and Sequencing ... Individual specific P1 adapters, each with a unique 5 bp barcode , were ligated to the Sbf I digested DNA at 22°C for 60 minutes by adding 1.8/0.6 µL 100 nmol/L P1 adapter, 0.45/0.15 µL 100 mmol/L rATP (Promega), 0.75/0.25 µL 10× Reaction Buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes made up to 45/15 µL with nuclease-free water for each parental/offspring sample.

    Article Title: Genome-wide association and genomic prediction of resistance to viral nervous necrosis in European sea bass (Dicentrarchus labrax) using RAD sequencing
    Article Snippet: Paragraph title: RAD library preparation and sequencing ... Individual specific P1 adapters, each with a unique 5–7 bp barcode, were ligated to the Sbf I-digested DNA at 20 °C for 60 min by adding 1.8/0.6 µL of 100 nmol/L P1 adapter, 0.45/0.15 µL of 100 mmol/L rATP (Promega), 0.75/0.25 µL 10 × reaction buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes for each parental/offspring sample were completed to 45/15 µL with nuclease-free water.

    Article Title: Accuracy of Genomic Evaluations of Juvenile Growth Rate in Common Carp (Cyprinus carpio) Using Genotyping by Sequencing
    Article Snippet: Paragraph title: RAD Library Preparation and Sequencing ... The reactions (12 μL final volumes) were then heat inactivated at 65°C for 20 min. Individual-specific P1 adapters, each with a unique 5 bp barcode, were ligated to the Sbf I-digested DNA at 20°C for 60 min by adding 1.8/0.6 μL 100 nmol/L P1 adapter, 0.45/0.15 μL 100 mmol/L rATP (Promega), 0.75/0.25 μL 10× Reaction Buffer 2 (NEB), 0.36/0.12 μL T4 ligase (NEB, 2 M U/mL), and reaction volumes made up to 45/15 μL with nuclease-free water for each parental/offspring sample.

    Sonication:

    Article Title: Mapping and Validation of the Major Sex-Determining Region in Nile Tilapia (Oreochromis niloticus L.) Using RAD Sequencing
    Article Snippet: Individual specific P1 adapters, each with a unique 5 bp barcode , were ligated to the Sbf I digested DNA at 22°C for 60 minutes by adding 1.8/0.6 µL 100 nmol/L P1 adapter, 0.45/0.15 µL 100 mmol/L rATP (Promega), 0.75/0.25 µL 10× Reaction Buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes made up to 45/15 µL with nuclease-free water for each parental/offspring sample. .. Shearing (Covaris S2 sonication) and initial size selection (250–550 bp) by agarose gel separation was followed by gel purification, end repair, dA overhang addition, P2 paired-end adapter ligation, library amplification, exactly as in the original RAD protocol , .

    Article Title: Genome-wide association and genomic prediction of resistance to viral nervous necrosis in European sea bass (Dicentrarchus labrax) using RAD sequencing
    Article Snippet: Individual specific P1 adapters, each with a unique 5–7 bp barcode, were ligated to the Sbf I-digested DNA at 20 °C for 60 min by adding 1.8/0.6 µL of 100 nmol/L P1 adapter, 0.45/0.15 µL of 100 mmol/L rATP (Promega), 0.75/0.25 µL 10 × reaction buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes for each parental/offspring sample were completed to 45/15 µL with nuclease-free water. .. Shearing (Covaris S2 sonication) and initial size selection (300 to 600 bp) by agarose gel separation was followed by gel purification, end repair, dA overhang addition, P2 (individual specific adapters) paired-end adapter ligation, library amplification, as in the original RAD protocol [ , ].

    Gel Purification:

    Article Title: Mapping and Validation of the Major Sex-Determining Region in Nile Tilapia (Oreochromis niloticus L.) Using RAD Sequencing
    Article Snippet: Individual specific P1 adapters, each with a unique 5 bp barcode , were ligated to the Sbf I digested DNA at 22°C for 60 minutes by adding 1.8/0.6 µL 100 nmol/L P1 adapter, 0.45/0.15 µL 100 mmol/L rATP (Promega), 0.75/0.25 µL 10× Reaction Buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes made up to 45/15 µL with nuclease-free water for each parental/offspring sample. .. Shearing (Covaris S2 sonication) and initial size selection (250–550 bp) by agarose gel separation was followed by gel purification, end repair, dA overhang addition, P2 paired-end adapter ligation, library amplification, exactly as in the original RAD protocol , .

    Article Title: Genome-wide association and genomic prediction of resistance to viral nervous necrosis in European sea bass (Dicentrarchus labrax) using RAD sequencing
    Article Snippet: Individual specific P1 adapters, each with a unique 5–7 bp barcode, were ligated to the Sbf I-digested DNA at 20 °C for 60 min by adding 1.8/0.6 µL of 100 nmol/L P1 adapter, 0.45/0.15 µL of 100 mmol/L rATP (Promega), 0.75/0.25 µL 10 × reaction buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes for each parental/offspring sample were completed to 45/15 µL with nuclease-free water. .. Shearing (Covaris S2 sonication) and initial size selection (300 to 600 bp) by agarose gel separation was followed by gel purification, end repair, dA overhang addition, P2 (individual specific adapters) paired-end adapter ligation, library amplification, as in the original RAD protocol [ , ].

    Article Title: Accuracy of Genomic Evaluations of Juvenile Growth Rate in Common Carp (Cyprinus carpio) Using Genotyping by Sequencing
    Article Snippet: The reactions (12 μL final volumes) were then heat inactivated at 65°C for 20 min. Individual-specific P1 adapters, each with a unique 5 bp barcode, were ligated to the Sbf I-digested DNA at 20°C for 60 min by adding 1.8/0.6 μL 100 nmol/L P1 adapter, 0.45/0.15 μL 100 mmol/L rATP (Promega), 0.75/0.25 μL 10× Reaction Buffer 2 (NEB), 0.36/0.12 μL T4 ligase (NEB, 2 M U/mL), and reaction volumes made up to 45/15 μL with nuclease-free water for each parental/offspring sample. .. Shearing and initial size selection (300–600 bp) by agarose gel separation were followed by gel purification, end repair, dA overhang addition, P2 (individual-specific adapters) paired-end adapter ligation, library amplification, as described in the original RAD protocol ( ; ).

    DNA Extraction:

    Article Title: Aerial Dissemination of Clostridium difficile spores
    Article Snippet: C. difficile NCTC 11209 was used as a control for the DNA extraction and purification process. .. A 25 μl reaction mix contained 1× Thermopol II Buffer (New England Biolabs), 200 mM of each dNTP, 2.25 mM of MgSO4 , 50 pmol of each primer (Sigma-Genosys), 2.5 units of Taq polymerase (New England Biolabs) and 2.5 μl of DNA template.

    Article Title: Genome-wide association and genomic prediction of resistance to viral nervous necrosis in European sea bass (Dicentrarchus labrax) using RAD sequencing
    Article Snippet: DNA was extracted from fin samples of the challenged fish using the REALPure genomic DNA extraction kit (Durviz S.L.) and treated with RNase. .. Individual specific P1 adapters, each with a unique 5–7 bp barcode, were ligated to the Sbf I-digested DNA at 20 °C for 60 min by adding 1.8/0.6 µL of 100 nmol/L P1 adapter, 0.45/0.15 µL of 100 mmol/L rATP (Promega), 0.75/0.25 µL 10 × reaction buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes for each parental/offspring sample were completed to 45/15 µL with nuclease-free water.

    Article Title: Accuracy of Genomic Evaluations of Juvenile Growth Rate in Common Carp (Cyprinus carpio) Using Genotyping by Sequencing
    Article Snippet: Genomic DNA was extracted from fin samples using the REALPure genomic DNA extraction kit (Durviz S.L.) and treated with RNase. .. The reactions (12 μL final volumes) were then heat inactivated at 65°C for 20 min. Individual-specific P1 adapters, each with a unique 5 bp barcode, were ligated to the Sbf I-digested DNA at 20°C for 60 min by adding 1.8/0.6 μL 100 nmol/L P1 adapter, 0.45/0.15 μL 100 mmol/L rATP (Promega), 0.75/0.25 μL 10× Reaction Buffer 2 (NEB), 0.36/0.12 μL T4 ligase (NEB, 2 M U/mL), and reaction volumes made up to 45/15 μL with nuclease-free water for each parental/offspring sample.

    Nucleic Acid Electrophoresis:

    Article Title: Mapping and Validation of the Major Sex-Determining Region in Nile Tilapia (Oreochromis niloticus L.) Using RAD Sequencing
    Article Snippet: Individual specific P1 adapters, each with a unique 5 bp barcode , were ligated to the Sbf I digested DNA at 22°C for 60 minutes by adding 1.8/0.6 µL 100 nmol/L P1 adapter, 0.45/0.15 µL 100 mmol/L rATP (Promega), 0.75/0.25 µL 10× Reaction Buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes made up to 45/15 µL with nuclease-free water for each parental/offspring sample. .. A total of 150 µL of each amplified library (16–18 PCR cycles, library dependent) was size selected (c. 350–650 bp) by gel electrophoresis .

    Article Title: Genome-wide association and genomic prediction of resistance to viral nervous necrosis in European sea bass (Dicentrarchus labrax) using RAD sequencing
    Article Snippet: Individual specific P1 adapters, each with a unique 5–7 bp barcode, were ligated to the Sbf I-digested DNA at 20 °C for 60 min by adding 1.8/0.6 µL of 100 nmol/L P1 adapter, 0.45/0.15 µL of 100 mmol/L rATP (Promega), 0.75/0.25 µL 10 × reaction buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes for each parental/offspring sample were completed to 45/15 µL with nuclease-free water. .. A volume of 150 µL of each amplified library (16 to 18 PCR cycles, library-dependent) was size-selected (400 to 700 bp) by gel electrophoresis [ ].

    Methylation:

    Article Title: 5-Aza-2?-deoxycytidine Sensitizes Busulfan-resistant Myeloid Leukemia Cells By Regulating Expression of Genes Involved in Cell Cycle Checkpoint and Apoptosis
    Article Snippet: .. Methylation-specific PCR (MSP) was performed using 30 ng bisulfite-modified DNA, 0.5 μM of each primer , 1 mM each of dATP, dCTP, dGTP and dTTP, 5 mM MgSO4 , and Taq DNA polymerase with ThermoPol II buffer (New England Biolabs, Ipswich, MA). .. The amplification method included initial heating at 95°C for 3 min, followed by 30 cycles of 95°C for 30 s, 57°C or 65°C for 30 s depending on the primers (see ), and 72°C for 33 s, followed by extension at 72°C for 10 min. MSP products were visualized on a 2.5% agarose gel and DNA band intensities were analyzed using Quant One software (Bio-Rad, Hercules, CA).

    Isolation:

    Article Title: 5-Aza-2?-deoxycytidine Sensitizes Busulfan-resistant Myeloid Leukemia Cells By Regulating Expression of Genes Involved in Cell Cycle Checkpoint and Apoptosis
    Article Snippet: Genomic DNA was isolated from the cell lines using a Wizard Genomic DNA Purification kit (Promega, Madison, WI). .. Methylation-specific PCR (MSP) was performed using 30 ng bisulfite-modified DNA, 0.5 μM of each primer , 1 mM each of dATP, dCTP, dGTP and dTTP, 5 mM MgSO4 , and Taq DNA polymerase with ThermoPol II buffer (New England Biolabs, Ipswich, MA).

    Size-exclusion Chromatography:

    Article Title: Admixture in Humans of Two Divergent Plasmodium knowlesi Populations Associated with Different Macaque Host Species
    Article Snippet: Both first and second round PCR amplifications were conducted in individual tubes or wells for each locus, in 11 μl reaction volume containing 0.2 mM each dNTP (Bioline, UK), 2 mM MgSO4 , 1X ThermoPol II reaction buffer (NEB, UK), 0.275 U Taq DNA polymerase (NEB, UK), 0.1 μM of each forward and reverse primer, and 1 μl sample DNA template. .. The PCR cycling conditions were as follows: initial denaturation at 94°C for 2 min, followed by 28 cycles of 94°C for 30 sec, annealing at 56°C for 30 sec and elongation at 68°C for 30 sec, with a final elongation step at 68°C for 1 min.

    Labeling:

    Article Title: Susceptibility to liver fibrosis in mice expressing a connective tissue growth factor transgene in hepatocytes
    Article Snippet: Genomic DNA (5 μg) from mice tails was digested in a 40 μl final volume with 20U Stu I in Reaction buffer II (New England Biolabs, Ipswich, MA). .. The membrane was probed with a previously described 1.1 kb human CCN2 cDNA ( ) that was labeled using Radprime labeling kit (Invitrogen).

    Purification:

    Article Title: Aerial Dissemination of Clostridium difficile spores
    Article Snippet: C. difficile NCTC 11209 was used as a control for the DNA extraction and purification process. .. A 25 μl reaction mix contained 1× Thermopol II Buffer (New England Biolabs), 200 mM of each dNTP, 2.25 mM of MgSO4 , 50 pmol of each primer (Sigma-Genosys), 2.5 units of Taq polymerase (New England Biolabs) and 2.5 μl of DNA template.

    Article Title: 5-Aza-2?-deoxycytidine Sensitizes Busulfan-resistant Myeloid Leukemia Cells By Regulating Expression of Genes Involved in Cell Cycle Checkpoint and Apoptosis
    Article Snippet: Bisulfite modification of the genomic DNA and its purification was performed using a MethylDetector kit (Active Motif, Carlsbad, CA). .. Methylation-specific PCR (MSP) was performed using 30 ng bisulfite-modified DNA, 0.5 μM of each primer , 1 mM each of dATP, dCTP, dGTP and dTTP, 5 mM MgSO4 , and Taq DNA polymerase with ThermoPol II buffer (New England Biolabs, Ipswich, MA).

    Article Title: Generalized stochastic profiling of transcriptional regulatory heterogeneities in tissues, tumors, and cultured cells
    Article Snippet: Terminal transferase (Roche #03333574001, 400 U μl−1 ) 10× ThermoPol buffer (New England Biolabs #B9005S) 100 mM MgSO4 (New England Biolabs #B1003S) BSA (Roche #10711454001) Roche Taq polymerase (Roche #04728858001, 5 U μl−1 ) AL1 primer (200 nmol synthesis from IDT or MWG): ATTGGATCCAGGCCGCTCTGGACAAAATATGAATTCTTTTTTTTTTTTTTTTTTTTT TTT ▲ CRITICAL Long desalted primers will have varying purities depending on the manufacturer. .. 100× SYBR Green I (Invitrogen #S7563 diluted 100-fold in DMSO to 100×) 50 mM aminoallyl-dUTP (Ambion #1103015) High-Fidelity polymerase (Roche #11732650001, 3.5 U μl−1 ) 10× High-Fidelity PCR buffer without Mg2+ (included with Roche #11759175001) PureLink PCR Purification Kit (Invitrogen #46-6056) NaOAc (Calbiochem #567418) 20 mg/ml glycogen (Invitrogen #10814-010) NaHCO3 (Acros #21712500) Alexa Fluor 555 reactive dye decapack (Invitrogen # ) GEX hybridization buffer (included with Illumina #BD-103-0204 or equivalent)

    Polymerase Chain Reaction:

    Article Title: Aerial Dissemination of Clostridium difficile spores
    Article Snippet: PCR ribotyping was performed based on a method described previously [ ] using primers 5'-CTGGGGTGAAGTCGTAACAAGG-3' and 5'-GCGCCCTTTGTAGCTTGACC-3'. .. A 25 μl reaction mix contained 1× Thermopol II Buffer (New England Biolabs), 200 mM of each dNTP, 2.25 mM of MgSO4 , 50 pmol of each primer (Sigma-Genosys), 2.5 units of Taq polymerase (New England Biolabs) and 2.5 μl of DNA template.

    Article Title: Optogenetic Control of Motor Coordination by Gi/o Protein-coupled Vertebrate Rhodopsin in Cerebellar Purkinje Cells *
    Article Snippet: .. The 50-ml final PCR reaction contained 1 ml of gDNA, 1 ml of each primer, 1 ml of dNTP mix (10 m m each of dATP, dTTP, dCTP, dGTP; New England Biolabs (NEB)), 5 ml of 10× Thermopol II reaction buffer (NEB), 5 ml of dimethyl sulfoxide, 0.5 ml of Taq Polymerase (NEB), and 35.5 ml of distilled H2 O. PCR reactions were run on an Eppendorf thermocycler using the following conditions: 92 °C for 30 s, 60 °C for 45 s, and 72 °C for 1 min run for 40 cycles or 95 °C for 30 s, 55 °C for 1 min and 72 °C for 1 min 30 s for 40 cycles to detect vRh-GFP or Cre-Recombinase, respectively. ..

    Article Title: Mapping and Validation of the Major Sex-Determining Region in Nile Tilapia (Oreochromis niloticus L.) Using RAD Sequencing
    Article Snippet: The RAD specific P1 and P2 paired-end adapters and library amplification PCR primer sequences used in this study are detailed in Baxter et al. . .. Individual specific P1 adapters, each with a unique 5 bp barcode , were ligated to the Sbf I digested DNA at 22°C for 60 minutes by adding 1.8/0.6 µL 100 nmol/L P1 adapter, 0.45/0.15 µL 100 mmol/L rATP (Promega), 0.75/0.25 µL 10× Reaction Buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes made up to 45/15 µL with nuclease-free water for each parental/offspring sample.

    Article Title: Visible Genotype Sensor Array
    Article Snippet: .. Typically, fifty microliters of a reaction mixture contained 1 x Mg-free ThermoPol II reaction buffer (New England Biolabs, Beverly, MA), 2.5 units of HotStar Taq DNA polymerase, 10 μM biotin-modified dUTP (Fermentas, Hanover, MD), 10 μM each of normal nucleotide (dATP, dCTP and dGTP), 4mM MgCl2 , and 1 μL of PCR product mixture. ..

    Article Title: Genome-wide association and genomic prediction of resistance to viral nervous necrosis in European sea bass (Dicentrarchus labrax) using RAD sequencing
    Article Snippet: Individual specific P1 adapters, each with a unique 5–7 bp barcode, were ligated to the Sbf I-digested DNA at 20 °C for 60 min by adding 1.8/0.6 µL of 100 nmol/L P1 adapter, 0.45/0.15 µL of 100 mmol/L rATP (Promega), 0.75/0.25 µL 10 × reaction buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes for each parental/offspring sample were completed to 45/15 µL with nuclease-free water. .. A volume of 150 µL of each amplified library (16 to 18 PCR cycles, library-dependent) was size-selected (400 to 700 bp) by gel electrophoresis [ ].

    Article Title: Aerial Dissemination of Clostridium difficile spores
    Article Snippet: REP primers REP1R-I, 5'-IIIICGICGICATCIGGC-3' and REP2-I, 5'-ICGICTTATCIGGCCTAC-3' were used in a PCR reaction based on a previously described method for typing C. difficile [ ] with minor modifications. .. A 25 μL reaction mix contained 1× Thermopol II Buffer (New England Biolabs), 200 μM of each dNTP, 2.5 mM of MgSO4 , 25pmol of each primer (Sigma-Genosys), 2 units Taq polymerase (New England Biolabs), 100 μg/mL bovine serum albumin (Promega) and DNA template (60 ng).

    Article Title: Osteosarcoma is characterised by reduced expression of markers of osteoclastogenesis and antigen presentation compared with normal bone
    Article Snippet: .. For PCR, 2 μ g of non-malignant bone or OS tumour biopsy RNA was reverse transcribed with BioScript (Bioline, Sydney, Australia); PCR was performed using Taq DNA polymerase with ThermoPol II buffer (New England Biolabs, Ipswich, MA, USA) at an annealing temperature of 53–55°C for 30 cycles on a ThermoHybaid PxE0.2 (Thermo Scientific, Waltham, MA, USA). ..

    Article Title: Accuracy of Genomic Evaluations of Juvenile Growth Rate in Common Carp (Cyprinus carpio) Using Genotyping by Sequencing
    Article Snippet: The RAD-specific P1 and P2 paired-end adapters and library amplification PCR primer sequences used in this study are detailed in . .. The reactions (12 μL final volumes) were then heat inactivated at 65°C for 20 min. Individual-specific P1 adapters, each with a unique 5 bp barcode, were ligated to the Sbf I-digested DNA at 20°C for 60 min by adding 1.8/0.6 μL 100 nmol/L P1 adapter, 0.45/0.15 μL 100 mmol/L rATP (Promega), 0.75/0.25 μL 10× Reaction Buffer 2 (NEB), 0.36/0.12 μL T4 ligase (NEB, 2 M U/mL), and reaction volumes made up to 45/15 μL with nuclease-free water for each parental/offspring sample.

    Article Title: 5-Aza-2?-deoxycytidine Sensitizes Busulfan-resistant Myeloid Leukemia Cells By Regulating Expression of Genes Involved in Cell Cycle Checkpoint and Apoptosis
    Article Snippet: .. Methylation-specific PCR (MSP) was performed using 30 ng bisulfite-modified DNA, 0.5 μM of each primer , 1 mM each of dATP, dCTP, dGTP and dTTP, 5 mM MgSO4 , and Taq DNA polymerase with ThermoPol II buffer (New England Biolabs, Ipswich, MA). .. The amplification method included initial heating at 95°C for 3 min, followed by 30 cycles of 95°C for 30 s, 57°C or 65°C for 30 s depending on the primers (see ), and 72°C for 33 s, followed by extension at 72°C for 10 min. MSP products were visualized on a 2.5% agarose gel and DNA band intensities were analyzed using Quant One software (Bio-Rad, Hercules, CA).

    Article Title: Generalized stochastic profiling of transcriptional regulatory heterogeneities in tissues, tumors, and cultured cells
    Article Snippet: Terminal transferase (Roche #03333574001, 400 U μl−1 ) 10× ThermoPol buffer (New England Biolabs #B9005S) 100 mM MgSO4 (New England Biolabs #B1003S) BSA (Roche #10711454001) Roche Taq polymerase (Roche #04728858001, 5 U μl−1 ) AL1 primer (200 nmol synthesis from IDT or MWG): ATTGGATCCAGGCCGCTCTGGACAAAATATGAATTCTTTTTTTTTTTTTTTTTTTTT TTT ▲ CRITICAL Long desalted primers will have varying purities depending on the manufacturer. .. 100× SYBR Green I (Invitrogen #S7563 diluted 100-fold in DMSO to 100×) 50 mM aminoallyl-dUTP (Ambion #1103015) High-Fidelity polymerase (Roche #11732650001, 3.5 U μl−1 ) 10× High-Fidelity PCR buffer without Mg2+ (included with Roche #11759175001) PureLink PCR Purification Kit (Invitrogen #46-6056) NaOAc (Calbiochem #567418) 20 mg/ml glycogen (Invitrogen #10814-010) NaHCO3 (Acros #21712500) Alexa Fluor 555 reactive dye decapack (Invitrogen # ) GEX hybridization buffer (included with Illumina #BD-103-0204 or equivalent)

    Article Title: Admixture in Humans of Two Divergent Plasmodium knowlesi Populations Associated with Different Macaque Host Species
    Article Snippet: .. Both first and second round PCR amplifications were conducted in individual tubes or wells for each locus, in 11 μl reaction volume containing 0.2 mM each dNTP (Bioline, UK), 2 mM MgSO4 , 1X ThermoPol II reaction buffer (NEB, UK), 0.275 U Taq DNA polymerase (NEB, UK), 0.1 μM of each forward and reverse primer, and 1 μl sample DNA template. .. The PCR cycling conditions were as follows: initial denaturation at 94°C for 2 min, followed by 28 cycles of 94°C for 30 sec, annealing at 56°C for 30 sec and elongation at 68°C for 30 sec, with a final elongation step at 68°C for 1 min.

    Lysis:

    Article Title: Optogenetic Control of Motor Coordination by Gi/o Protein-coupled Vertebrate Rhodopsin in Cerebellar Purkinje Cells *
    Article Snippet: Twenty microliters of proteinase K (20 mg/ml, Roche Diagnostics) was added to the lysis buffer, and the mixture was shaken overnight at 55 °C. .. The 50-ml final PCR reaction contained 1 ml of gDNA, 1 ml of each primer, 1 ml of dNTP mix (10 m m each of dATP, dTTP, dCTP, dGTP; New England Biolabs (NEB)), 5 ml of 10× Thermopol II reaction buffer (NEB), 5 ml of dimethyl sulfoxide, 0.5 ml of Taq Polymerase (NEB), and 35.5 ml of distilled H2 O. PCR reactions were run on an Eppendorf thermocycler using the following conditions: 92 °C for 30 s, 60 °C for 45 s, and 72 °C for 1 min run for 40 cycles or 95 °C for 30 s, 55 °C for 1 min and 72 °C for 1 min 30 s for 40 cycles to detect vRh-GFP or Cre-Recombinase, respectively.

    Mouse Assay:

    Article Title: Optogenetic Control of Motor Coordination by Gi/o Protein-coupled Vertebrate Rhodopsin in Cerebellar Purkinje Cells *
    Article Snippet: Paragraph title: Generation and Screening of Transgenic Mice ... The 50-ml final PCR reaction contained 1 ml of gDNA, 1 ml of each primer, 1 ml of dNTP mix (10 m m each of dATP, dTTP, dCTP, dGTP; New England Biolabs (NEB)), 5 ml of 10× Thermopol II reaction buffer (NEB), 5 ml of dimethyl sulfoxide, 0.5 ml of Taq Polymerase (NEB), and 35.5 ml of distilled H2 O. PCR reactions were run on an Eppendorf thermocycler using the following conditions: 92 °C for 30 s, 60 °C for 45 s, and 72 °C for 1 min run for 40 cycles or 95 °C for 30 s, 55 °C for 1 min and 72 °C for 1 min 30 s for 40 cycles to detect vRh-GFP or Cre-Recombinase, respectively.

    Article Title: Susceptibility to liver fibrosis in mice expressing a connective tissue growth factor transgene in hepatocytes
    Article Snippet: .. Genomic DNA (5 μg) from mice tails was digested in a 40 μl final volume with 20U Stu I in Reaction buffer II (New England Biolabs, Ipswich, MA). .. DNA fragments were separated in a 0.7% agarose gel and transferred to Hybond N+ membrane (Amersham-Pharmacia, Piscataway, NJ).

    Plasmid Preparation:

    Article Title: Characterization of the Human SNM1A and SNM1B/Apollo DNA Repair Exonucleases *
    Article Snippet: .. First, plasmid (325 ng, supplemental Fig. S7C ) was either left untreated, gapped (nicked at five sites within a 52 base pair region) with Nb.BbvCI (5 units; New England Biolabs), or linearized with HindIII (10 units; New England Biolabs) in 10 μl for 90 min at 37 °C in 1× New England Biolabs reaction buffer 2. .. Second, reaction buffer (final composition 20 mm HEPES-KOH, pH 7.5, 50 mm KCl, 0.5 mm DTT, 10 mm MgCl2 , 0.05% Triton X, 0.1 mg/ml BSA, 5% glycerol) and, in competition experiments unlabeled 21-mer dsDNA, were added and hSNM1A or hSNM1B were allowed to digest the plasmid for 1 h. Products were analyzed on 1% agarose gels containing 1× Tris borate EDTA (130 V, 150 min).

    Software:

    Article Title: 5-Aza-2?-deoxycytidine Sensitizes Busulfan-resistant Myeloid Leukemia Cells By Regulating Expression of Genes Involved in Cell Cycle Checkpoint and Apoptosis
    Article Snippet: Methylation-specific PCR (MSP) was performed using 30 ng bisulfite-modified DNA, 0.5 μM of each primer , 1 mM each of dATP, dCTP, dGTP and dTTP, 5 mM MgSO4 , and Taq DNA polymerase with ThermoPol II buffer (New England Biolabs, Ipswich, MA). .. The amplification method included initial heating at 95°C for 3 min, followed by 30 cycles of 95°C for 30 s, 57°C or 65°C for 30 s depending on the primers (see ), and 72°C for 33 s, followed by extension at 72°C for 10 min. MSP products were visualized on a 2.5% agarose gel and DNA band intensities were analyzed using Quant One software (Bio-Rad, Hercules, CA).

    Article Title: Admixture in Humans of Two Divergent Plasmodium knowlesi Populations Associated with Different Macaque Host Species
    Article Snippet: Both first and second round PCR amplifications were conducted in individual tubes or wells for each locus, in 11 μl reaction volume containing 0.2 mM each dNTP (Bioline, UK), 2 mM MgSO4 , 1X ThermoPol II reaction buffer (NEB, UK), 0.275 U Taq DNA polymerase (NEB, UK), 0.1 μM of each forward and reverse primer, and 1 μl sample DNA template. .. GENEMAPPER version 4.0 software (Applied Biosystems, UK) was used for scoring of allele electrophoretic size, and quantification of peak heights.

    Multiplex Assay:

    Article Title: Mapping and Validation of the Major Sex-Determining Region in Nile Tilapia (Oreochromis niloticus L.) Using RAD Sequencing
    Article Snippet: Individual specific P1 adapters, each with a unique 5 bp barcode , were ligated to the Sbf I digested DNA at 22°C for 60 minutes by adding 1.8/0.6 µL 100 nmol/L P1 adapter, 0.45/0.15 µL 100 mmol/L rATP (Promega), 0.75/0.25 µL 10× Reaction Buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes made up to 45/15 µL with nuclease-free water for each parental/offspring sample. .. Following heat inactivation at 65°C for 20 minutes, the ligation reactions were slowly cooled to room temperature (over 1 hour) then combined in appropriate multiplex pools ( ).

    Article Title: Genome-wide association and genomic prediction of resistance to viral nervous necrosis in European sea bass (Dicentrarchus labrax) using RAD sequencing
    Article Snippet: Individual specific P1 adapters, each with a unique 5–7 bp barcode, were ligated to the Sbf I-digested DNA at 20 °C for 60 min by adding 1.8/0.6 µL of 100 nmol/L P1 adapter, 0.45/0.15 µL of 100 mmol/L rATP (Promega), 0.75/0.25 µL 10 × reaction buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes for each parental/offspring sample were completed to 45/15 µL with nuclease-free water. .. Following heat inactivation at 65 °C for 20 min, ligation reactions were slowly cooled down to room temperature (over 1 h), then combined in appropriate multiplex pools.

    Article Title: Accuracy of Genomic Evaluations of Juvenile Growth Rate in Common Carp (Cyprinus carpio) Using Genotyping by Sequencing
    Article Snippet: The reactions (12 μL final volumes) were then heat inactivated at 65°C for 20 min. Individual-specific P1 adapters, each with a unique 5 bp barcode, were ligated to the Sbf I-digested DNA at 20°C for 60 min by adding 1.8/0.6 μL 100 nmol/L P1 adapter, 0.45/0.15 μL 100 mmol/L rATP (Promega), 0.75/0.25 μL 10× Reaction Buffer 2 (NEB), 0.36/0.12 μL T4 ligase (NEB, 2 M U/mL), and reaction volumes made up to 45/15 μL with nuclease-free water for each parental/offspring sample. .. Following heat inactivation at 65°C for 20 min, the ligation reactions were slowly cooled to room temperature (over 1 h) then combined in appropriate multiplex pools.

    Selection:

    Article Title: Mapping and Validation of the Major Sex-Determining Region in Nile Tilapia (Oreochromis niloticus L.) Using RAD Sequencing
    Article Snippet: Individual specific P1 adapters, each with a unique 5 bp barcode , were ligated to the Sbf I digested DNA at 22°C for 60 minutes by adding 1.8/0.6 µL 100 nmol/L P1 adapter, 0.45/0.15 µL 100 mmol/L rATP (Promega), 0.75/0.25 µL 10× Reaction Buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes made up to 45/15 µL with nuclease-free water for each parental/offspring sample. .. Shearing (Covaris S2 sonication) and initial size selection (250–550 bp) by agarose gel separation was followed by gel purification, end repair, dA overhang addition, P2 paired-end adapter ligation, library amplification, exactly as in the original RAD protocol , .

    Article Title: Genome-wide association and genomic prediction of resistance to viral nervous necrosis in European sea bass (Dicentrarchus labrax) using RAD sequencing
    Article Snippet: Individual specific P1 adapters, each with a unique 5–7 bp barcode, were ligated to the Sbf I-digested DNA at 20 °C for 60 min by adding 1.8/0.6 µL of 100 nmol/L P1 adapter, 0.45/0.15 µL of 100 mmol/L rATP (Promega), 0.75/0.25 µL 10 × reaction buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes for each parental/offspring sample were completed to 45/15 µL with nuclease-free water. .. Shearing (Covaris S2 sonication) and initial size selection (300 to 600 bp) by agarose gel separation was followed by gel purification, end repair, dA overhang addition, P2 (individual specific adapters) paired-end adapter ligation, library amplification, as in the original RAD protocol [ , ].

    Article Title: Accuracy of Genomic Evaluations of Juvenile Growth Rate in Common Carp (Cyprinus carpio) Using Genotyping by Sequencing
    Article Snippet: The reactions (12 μL final volumes) were then heat inactivated at 65°C for 20 min. Individual-specific P1 adapters, each with a unique 5 bp barcode, were ligated to the Sbf I-digested DNA at 20°C for 60 min by adding 1.8/0.6 μL 100 nmol/L P1 adapter, 0.45/0.15 μL 100 mmol/L rATP (Promega), 0.75/0.25 μL 10× Reaction Buffer 2 (NEB), 0.36/0.12 μL T4 ligase (NEB, 2 M U/mL), and reaction volumes made up to 45/15 μL with nuclease-free water for each parental/offspring sample. .. Shearing and initial size selection (300–600 bp) by agarose gel separation were followed by gel purification, end repair, dA overhang addition, P2 (individual-specific adapters) paired-end adapter ligation, library amplification, as described in the original RAD protocol ( ; ).

    Agarose Gel Electrophoresis:

    Article Title: Aerial Dissemination of Clostridium difficile spores
    Article Snippet: A 25 μl reaction mix contained 1× Thermopol II Buffer (New England Biolabs), 200 mM of each dNTP, 2.25 mM of MgSO4 , 50 pmol of each primer (Sigma-Genosys), 2.5 units of Taq polymerase (New England Biolabs) and 2.5 μl of DNA template. .. Amplimers were resolved on a 2% agarose gel (Invitrogen).

    Article Title: Mapping and Validation of the Major Sex-Determining Region in Nile Tilapia (Oreochromis niloticus L.) Using RAD Sequencing
    Article Snippet: Each sample was quantified by spectrophotometry (Nanodrop) and quality assessed by agarose gel electrophoresis, and was finally diluted to a concentration of 50 ng/µL in 5 mmol/L Tris, pH 8.5. .. Individual specific P1 adapters, each with a unique 5 bp barcode , were ligated to the Sbf I digested DNA at 22°C for 60 minutes by adding 1.8/0.6 µL 100 nmol/L P1 adapter, 0.45/0.15 µL 100 mmol/L rATP (Promega), 0.75/0.25 µL 10× Reaction Buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes made up to 45/15 µL with nuclease-free water for each parental/offspring sample.

    Article Title: Susceptibility to liver fibrosis in mice expressing a connective tissue growth factor transgene in hepatocytes
    Article Snippet: Genomic DNA (5 μg) from mice tails was digested in a 40 μl final volume with 20U Stu I in Reaction buffer II (New England Biolabs, Ipswich, MA). .. DNA fragments were separated in a 0.7% agarose gel and transferred to Hybond N+ membrane (Amersham-Pharmacia, Piscataway, NJ).

    Article Title: Genome-wide association and genomic prediction of resistance to viral nervous necrosis in European sea bass (Dicentrarchus labrax) using RAD sequencing
    Article Snippet: Each sample was quantified by spectrophotometry (Nanodrop), quality-assessed by agarose gel electrophoresis, and finally diluted to a concentration of 20 ng/µL in 5 mmol/L Tris, pH 8.5 using a Qubit Fluorometer (Invitrogen). .. Individual specific P1 adapters, each with a unique 5–7 bp barcode, were ligated to the Sbf I-digested DNA at 20 °C for 60 min by adding 1.8/0.6 µL of 100 nmol/L P1 adapter, 0.45/0.15 µL of 100 mmol/L rATP (Promega), 0.75/0.25 µL 10 × reaction buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes for each parental/offspring sample were completed to 45/15 µL with nuclease-free water.

    Article Title: Accuracy of Genomic Evaluations of Juvenile Growth Rate in Common Carp (Cyprinus carpio) Using Genotyping by Sequencing
    Article Snippet: Each sample was quantified by spectrophotometry (Nanodrop), and its quality was assessed by agarose gel electrophoresis, before being diluted to a concentration of 20 ng/μL [measured by Qubit Fluorometer (Invitrogen)] in 5 mmol/L Tris, pH 8.5. .. The reactions (12 μL final volumes) were then heat inactivated at 65°C for 20 min. Individual-specific P1 adapters, each with a unique 5 bp barcode, were ligated to the Sbf I-digested DNA at 20°C for 60 min by adding 1.8/0.6 μL 100 nmol/L P1 adapter, 0.45/0.15 μL 100 mmol/L rATP (Promega), 0.75/0.25 μL 10× Reaction Buffer 2 (NEB), 0.36/0.12 μL T4 ligase (NEB, 2 M U/mL), and reaction volumes made up to 45/15 μL with nuclease-free water for each parental/offspring sample.

    Article Title: 5-Aza-2?-deoxycytidine Sensitizes Busulfan-resistant Myeloid Leukemia Cells By Regulating Expression of Genes Involved in Cell Cycle Checkpoint and Apoptosis
    Article Snippet: Methylation-specific PCR (MSP) was performed using 30 ng bisulfite-modified DNA, 0.5 μM of each primer , 1 mM each of dATP, dCTP, dGTP and dTTP, 5 mM MgSO4 , and Taq DNA polymerase with ThermoPol II buffer (New England Biolabs, Ipswich, MA). .. The amplification method included initial heating at 95°C for 3 min, followed by 30 cycles of 95°C for 30 s, 57°C or 65°C for 30 s depending on the primers (see ), and 72°C for 33 s, followed by extension at 72°C for 10 min. MSP products were visualized on a 2.5% agarose gel and DNA band intensities were analyzed using Quant One software (Bio-Rad, Hercules, CA).

    Transgenic Assay:

    Article Title: Optogenetic Control of Motor Coordination by Gi/o Protein-coupled Vertebrate Rhodopsin in Cerebellar Purkinje Cells *
    Article Snippet: Paragraph title: Generation and Screening of Transgenic Mice ... The 50-ml final PCR reaction contained 1 ml of gDNA, 1 ml of each primer, 1 ml of dNTP mix (10 m m each of dATP, dTTP, dCTP, dGTP; New England Biolabs (NEB)), 5 ml of 10× Thermopol II reaction buffer (NEB), 5 ml of dimethyl sulfoxide, 0.5 ml of Taq Polymerase (NEB), and 35.5 ml of distilled H2 O. PCR reactions were run on an Eppendorf thermocycler using the following conditions: 92 °C for 30 s, 60 °C for 45 s, and 72 °C for 1 min run for 40 cycles or 95 °C for 30 s, 55 °C for 1 min and 72 °C for 1 min 30 s for 40 cycles to detect vRh-GFP or Cre-Recombinase, respectively.

    Spectrophotometry:

    Article Title: Mapping and Validation of the Major Sex-Determining Region in Nile Tilapia (Oreochromis niloticus L.) Using RAD Sequencing
    Article Snippet: Each sample was quantified by spectrophotometry (Nanodrop) and quality assessed by agarose gel electrophoresis, and was finally diluted to a concentration of 50 ng/µL in 5 mmol/L Tris, pH 8.5. .. Individual specific P1 adapters, each with a unique 5 bp barcode , were ligated to the Sbf I digested DNA at 22°C for 60 minutes by adding 1.8/0.6 µL 100 nmol/L P1 adapter, 0.45/0.15 µL 100 mmol/L rATP (Promega), 0.75/0.25 µL 10× Reaction Buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes made up to 45/15 µL with nuclease-free water for each parental/offspring sample.

    Article Title: Genome-wide association and genomic prediction of resistance to viral nervous necrosis in European sea bass (Dicentrarchus labrax) using RAD sequencing
    Article Snippet: Each sample was quantified by spectrophotometry (Nanodrop), quality-assessed by agarose gel electrophoresis, and finally diluted to a concentration of 20 ng/µL in 5 mmol/L Tris, pH 8.5 using a Qubit Fluorometer (Invitrogen). .. Individual specific P1 adapters, each with a unique 5–7 bp barcode, were ligated to the Sbf I-digested DNA at 20 °C for 60 min by adding 1.8/0.6 µL of 100 nmol/L P1 adapter, 0.45/0.15 µL of 100 mmol/L rATP (Promega), 0.75/0.25 µL 10 × reaction buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes for each parental/offspring sample were completed to 45/15 µL with nuclease-free water.

    Article Title: Accuracy of Genomic Evaluations of Juvenile Growth Rate in Common Carp (Cyprinus carpio) Using Genotyping by Sequencing
    Article Snippet: Each sample was quantified by spectrophotometry (Nanodrop), and its quality was assessed by agarose gel electrophoresis, before being diluted to a concentration of 20 ng/μL [measured by Qubit Fluorometer (Invitrogen)] in 5 mmol/L Tris, pH 8.5. .. The reactions (12 μL final volumes) were then heat inactivated at 65°C for 20 min. Individual-specific P1 adapters, each with a unique 5 bp barcode, were ligated to the Sbf I-digested DNA at 20°C for 60 min by adding 1.8/0.6 μL 100 nmol/L P1 adapter, 0.45/0.15 μL 100 mmol/L rATP (Promega), 0.75/0.25 μL 10× Reaction Buffer 2 (NEB), 0.36/0.12 μL T4 ligase (NEB, 2 M U/mL), and reaction volumes made up to 45/15 μL with nuclease-free water for each parental/offspring sample.

    Concentration Assay:

    Article Title: Mapping and Validation of the Major Sex-Determining Region in Nile Tilapia (Oreochromis niloticus L.) Using RAD Sequencing
    Article Snippet: Each sample (0.72 µg parental DNA/0.24 µg offspring DNA) was digested at 37°C for 40 minutes with Sbf I (recognising the CCTGCA|GG motif) high fidelity restriction enzyme (New England Biolabs; NEB) using 6 U Sbf I per µg genomic DNA in 1× Reaction Buffer 4 (NEB) at a final concentration of c. 1 µg DNA per 50 µL reaction volume. .. Individual specific P1 adapters, each with a unique 5 bp barcode , were ligated to the Sbf I digested DNA at 22°C for 60 minutes by adding 1.8/0.6 µL 100 nmol/L P1 adapter, 0.45/0.15 µL 100 mmol/L rATP (Promega), 0.75/0.25 µL 10× Reaction Buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes made up to 45/15 µL with nuclease-free water for each parental/offspring sample.

    Article Title: Characterization of the Human SNM1A and SNM1B/Apollo DNA Repair Exonucleases *
    Article Snippet: It is worth noting that at this substrate concentration the hSNM1B-catalyzed hydrolysis of dsDNA is not maximal, but an equivalent level of substrate was used for comparison purposes after a normalization experiment was carried out to determine the amounts of hSNM1A and hSNM1B that would result in an approximately equivalent level of hydrolysis ( supplemental Fig. S2A ). .. First, plasmid (325 ng, supplemental Fig. S7C ) was either left untreated, gapped (nicked at five sites within a 52 base pair region) with Nb.BbvCI (5 units; New England Biolabs), or linearized with HindIII (10 units; New England Biolabs) in 10 μl for 90 min at 37 °C in 1× New England Biolabs reaction buffer 2.

    Article Title: Genome-wide association and genomic prediction of resistance to viral nervous necrosis in European sea bass (Dicentrarchus labrax) using RAD sequencing
    Article Snippet: Briefly, each sample (0.72 µg parental DNA per 0.24 µg offspring DNA) was digested at 37 °C for 60 min with the high-fidelity restriction enzyme Sbf I (that recognises the CCTGCA|GG motif) − (New England Biolabs; NEB) using 6 U Sbf I per µg genomic DNA in 1 × Reaction Buffer 4 (NEB) at a final concentration of 1 µg DNA per 50 µL reaction volume. .. Individual specific P1 adapters, each with a unique 5–7 bp barcode, were ligated to the Sbf I-digested DNA at 20 °C for 60 min by adding 1.8/0.6 µL of 100 nmol/L P1 adapter, 0.45/0.15 µL of 100 mmol/L rATP (Promega), 0.75/0.25 µL 10 × reaction buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes for each parental/offspring sample were completed to 45/15 µL with nuclease-free water.

    Article Title: Accuracy of Genomic Evaluations of Juvenile Growth Rate in Common Carp (Cyprinus carpio) Using Genotyping by Sequencing
    Article Snippet: Each sample was quantified by spectrophotometry (Nanodrop), and its quality was assessed by agarose gel electrophoresis, before being diluted to a concentration of 20 ng/μL [measured by Qubit Fluorometer (Invitrogen)] in 5 mmol/L Tris, pH 8.5. .. The reactions (12 μL final volumes) were then heat inactivated at 65°C for 20 min. Individual-specific P1 adapters, each with a unique 5 bp barcode, were ligated to the Sbf I-digested DNA at 20°C for 60 min by adding 1.8/0.6 μL 100 nmol/L P1 adapter, 0.45/0.15 μL 100 mmol/L rATP (Promega), 0.75/0.25 μL 10× Reaction Buffer 2 (NEB), 0.36/0.12 μL T4 ligase (NEB, 2 M U/mL), and reaction volumes made up to 45/15 μL with nuclease-free water for each parental/offspring sample.

    DNA Purification:

    Article Title: 5-Aza-2?-deoxycytidine Sensitizes Busulfan-resistant Myeloid Leukemia Cells By Regulating Expression of Genes Involved in Cell Cycle Checkpoint and Apoptosis
    Article Snippet: Genomic DNA was isolated from the cell lines using a Wizard Genomic DNA Purification kit (Promega, Madison, WI). .. Methylation-specific PCR (MSP) was performed using 30 ng bisulfite-modified DNA, 0.5 μM of each primer , 1 mM each of dATP, dCTP, dGTP and dTTP, 5 mM MgSO4 , and Taq DNA polymerase with ThermoPol II buffer (New England Biolabs, Ipswich, MA).

    Staining:

    Article Title: Aerial Dissemination of Clostridium difficile spores
    Article Snippet: A 25 μl reaction mix contained 1× Thermopol II Buffer (New England Biolabs), 200 mM of each dNTP, 2.25 mM of MgSO4 , 50 pmol of each primer (Sigma-Genosys), 2.5 units of Taq polymerase (New England Biolabs) and 2.5 μl of DNA template. .. After ethidium bromide staining, banding patterns were compared visually.

    Fluorescence In Situ Hybridization:

    Article Title: Genome-wide association and genomic prediction of resistance to viral nervous necrosis in European sea bass (Dicentrarchus labrax) using RAD sequencing
    Article Snippet: DNA was extracted from fin samples of the challenged fish using the REALPure genomic DNA extraction kit (Durviz S.L.) and treated with RNase. .. Individual specific P1 adapters, each with a unique 5–7 bp barcode, were ligated to the Sbf I-digested DNA at 20 °C for 60 min by adding 1.8/0.6 µL of 100 nmol/L P1 adapter, 0.45/0.15 µL of 100 mmol/L rATP (Promega), 0.75/0.25 µL 10 × reaction buffer 2 (NEB), 0.36/0.12 µL T4 ligase (NEB, 2 M U/mL) and reaction volumes for each parental/offspring sample were completed to 45/15 µL with nuclease-free water.

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  • 90
    New England Biolabs thermopol ii reaction buffer
    Thermopol Ii Reaction Buffer, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 90/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/thermopol ii reaction buffer/product/New England Biolabs
    Average 90 stars, based on 4 article reviews
    Price from $9.99 to $1999.99
    thermopol ii reaction buffer - by Bioz Stars, 2020-02
    90/100 stars
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