phusion hot start polymerase new england biolabs  (New England Biolabs)


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  • 99
    Name:
    Phusion Hot Start Flex DNA Polymerase
    Description:
    Phusion Hot Start Flex DNA Polymerase 500 units
    Catalog Number:
    m0535l
    Price:
    554
    Size:
    500 units
    Category:
    Thermostable DNA Polymerases
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    Structured Review

    New England Biolabs phusion hot start polymerase new england biolabs
    Phusion Hot Start Flex DNA Polymerase
    Phusion Hot Start Flex DNA Polymerase 500 units
    https://www.bioz.com/result/phusion hot start polymerase new england biolabs/product/New England Biolabs
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    phusion hot start polymerase new england biolabs - by Bioz Stars, 2020-08
    99/100 stars

    Related Products / Commonly Used Together

    dmso
    phusion hf buffer
    pcr reactions

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    Related Articles

    Clone Assay:

    Article Title: Recombinase-mediated cassette exchange (RMCE) system for functional genomics studies in Mycoplasma mycoides
    Article Snippet: .. Transformation-Associated Recombination (TAR) cloning of the 100 kb DNA segment A 7.5 kilobase (kb) TAR cloning vector was generated by PCR amplification of pRC60 using two primers (5′- ACTAATAATAAAACATTTATATACTTAATGAATAAATATAATTAG TACCGTTCGTATAATGTATGC-3′ and 5′-ATTTTAAAATTTATGTAATTTATTAATTTTTATCTTTATAATATA TACCGTTCGTATATGGTTTCT-3′) and the Phusion Hot Start High-Fidelity DNA polymerase with HF buffer (New England Biolabs; NEB) according to the manufacturer’s instructions with modifications. ..

    Amplification:

    Article Title: Epigenetic and transcriptional determinants of the human breast
    Article Snippet: .. The resulting product was PCR amplified using the 3′ PCR primer (5′-CAAGCAGAAGACGGCATACGAGAT-3′) and an indexed 5′ PCR primer (5′-AATGATACGGCGACCACCGACAGNNNNNNGTTCAGAGTTCTACAGTCCGA-3′), Phusion Hot Start High Fidelity DNA polymerase (NEB Canada, cat. F-540 l), buffer, dNTPs and dimethylsulphoxide (DMSO). ..

    Article Title: Recombinase-mediated cassette exchange (RMCE) system for functional genomics studies in Mycoplasma mycoides
    Article Snippet: .. Transformation-Associated Recombination (TAR) cloning of the 100 kb DNA segment A 7.5 kilobase (kb) TAR cloning vector was generated by PCR amplification of pRC60 using two primers (5′- ACTAATAATAAAACATTTATATACTTAATGAATAAATATAATTAG TACCGTTCGTATAATGTATGC-3′ and 5′-ATTTTAAAATTTATGTAATTTATTAATTTTTATCTTTATAATATA TACCGTTCGTATATGGTTTCT-3′) and the Phusion Hot Start High-Fidelity DNA polymerase with HF buffer (New England Biolabs; NEB) according to the manufacturer’s instructions with modifications. ..

    Article Title: Solid-phase cloning for high-throughput assembly of single and multiple DNA parts
    Article Snippet: .. DNA parts were amplified with Phusion® Hot-Start Flex (2 U/μl, New England Biolabs, Ipswich, MA, USA) by the following PCR program: 98°C for 30 s, 30 cycles of 98°C 8 s, 25 s annealing with temperature depending on primer melting temperature and 72°C for 20 s/kb, before ending with 72°C for 7 min followed by 4°C hold. ..

    Article Title: A Flexible Approach for Highly Multiplexed Candidate Gene Targeted Resequencing
    Article Snippet: .. After a brief purification using the Spin-20 columns (Princeton Separations), the captured DNA pool was amplified by PCR (98°C for 30 seconds followed by 36–37 cycles at 98°C for 10 seconds, 65°C for 30 seconds, and 73°C for 30 seconds) using Phusion Hot Start High-Fidelity DNA polymerase (New England BioLabs) and non-target specific common primers that are homologous to the vector oligonucleotide. .. After purification using the Fermentas PCR Purification kit, 0.5–1 µg PCR products per sequencing library were ligated to each other using T4 DNA ligase (New England BioLabs).

    Article Title: Dimeric CRISPR RNA-Guided FokI-dCas9 Nucleases Directed by Truncated gRNAs for Highly Specific Genome Editing
    Article Snippet: .. Genomic sites were amplified for deep sequencing with Phusion Hot-start FLEX DNA polymerase (NEB) and primers listed in and the touchdown PCR protocol as described above. .. Amplicons of 200–350 bp were purified using Ampure XP beads (Beckman Coulter Genomics) according to manufacturer's instructions.

    In Vitro:

    Article Title: A strategy of gene overexpression based on tandem repetitive promoters in Escherichia coli
    Article Snippet: .. Then, fragment 1, 2 and 3 were assembled together in vitro under the action of T5 exonuclease (Epicentre), Phusion Hot Start DNA Polymerase (New England Biolabs (NEB)) and Taq DNA ligase (NEB) at 50°C for 15 min. .. The resulting constructs containing different promoters were then transformed into competent cells and were firstly screened based on the fluorescence signal and PCR detection.

    Purification:

    Article Title: A Flexible Approach for Highly Multiplexed Candidate Gene Targeted Resequencing
    Article Snippet: .. After a brief purification using the Spin-20 columns (Princeton Separations), the captured DNA pool was amplified by PCR (98°C for 30 seconds followed by 36–37 cycles at 98°C for 10 seconds, 65°C for 30 seconds, and 73°C for 30 seconds) using Phusion Hot Start High-Fidelity DNA polymerase (New England BioLabs) and non-target specific common primers that are homologous to the vector oligonucleotide. .. After purification using the Fermentas PCR Purification kit, 0.5–1 µg PCR products per sequencing library were ligated to each other using T4 DNA ligase (New England BioLabs).

    Polymerase Chain Reaction:

    Article Title: Systematic characterization of glutathione S-transferases in common marmosets.
    Article Snippet: .. The common marmoset is an important primate species used in drug metabolism studies. ..

    Article Title: Epitope-tagged protein-based artificial microRNA (ETPamir) screens for optimized gene silencing in plants
    Article Snippet: .. 4-morpholineethanesulfonic acid (MES, Sigma, cat. no. M3671) Mannitol (MP Biomedicals, cat. no. 102248) MgCl2 (Sigma, cat. no. M9272) KCl (Sigma, cat. no. P3911) NaCl (Sigma, cat. no. S9888) CaCl2 (Sigma, cat. no. C7902) CsCl (American Bioanalytical, cat. no. AB00300) Polyethylene glycol 4000 (PEG4000, Sigma, cat. no. 81240) Tween-20 (Sigma, cat. no. P7949) Phusion DNA polymerase (NEB, cat. no. M0535) PVDF membrane (Immobilon, cat. no. IPVH304F0) Nonfat dry milk (Santa Cruz Biotechology, cat. no. sc-2325) Bovine calf serum (HyClone, cat. no. SH30072.03) HA HRP-conjugated antibody (Roche, cat. no. 12013819001) SuperSignal West Pico chemiluminescent kit (Thermo Scientific, cat. no. 34080) SuperSignal West Femto chemiluminescent kit (Thermo Scientific, cat. no. 34095) 10% precast polyacrylamide gel (Bio-Rad, cat. no. 456-1034) Terrific broth (American Bioanalytical, cat. no. AB01966) Custom oligo primers for PCR generating amiRNA precursors (Oligo synthesis service provider) pHBT-HA constitutive expression vector (Available from the authors upon request) pHSP-HA heat shock inducible expression vector (Available from the authors upon request) pHBT-ath-miR319a constitutive expression plasmid (Available from the authors upon request) .. CL2 clinical centrifuge (Thermo Scientific, cat. no. 004260F) Mini-PROTEAN Tetra system (Bio-Rad, cat. no. 165-8006) Trans-Blot SD semi-dry electrophoretic transfer cell (Bio-Rad, cat. no. 170-3940) Fisher Scientific Isotemp heating block (Fisher Scientific, cat. no. 11-715-305Q) 2-ml round-bottom microcentrifuge tubes (USA Scientific, cat. no. 1620-2700) 1.5-ml microcentrifuge tubes (USA Scientific, cat. no. 1615-5500) 6-well culture plates (Falcon, cat. no. 3046) 1000-ml storage bottle with 0.22 μm vacuum filter (Corning, cat. no. 430517)

    Article Title: Epigenetic and transcriptional determinants of the human breast
    Article Snippet: .. The resulting product was PCR amplified using the 3′ PCR primer (5′-CAAGCAGAAGACGGCATACGAGAT-3′) and an indexed 5′ PCR primer (5′-AATGATACGGCGACCACCGACAGNNNNNNGTTCAGAGTTCTACAGTCCGA-3′), Phusion Hot Start High Fidelity DNA polymerase (NEB Canada, cat. F-540 l), buffer, dNTPs and dimethylsulphoxide (DMSO). ..

    Article Title: Recombinase-mediated cassette exchange (RMCE) system for functional genomics studies in Mycoplasma mycoides
    Article Snippet: .. Transformation-Associated Recombination (TAR) cloning of the 100 kb DNA segment A 7.5 kilobase (kb) TAR cloning vector was generated by PCR amplification of pRC60 using two primers (5′- ACTAATAATAAAACATTTATATACTTAATGAATAAATATAATTAG TACCGTTCGTATAATGTATGC-3′ and 5′-ATTTTAAAATTTATGTAATTTATTAATTTTTATCTTTATAATATA TACCGTTCGTATATGGTTTCT-3′) and the Phusion Hot Start High-Fidelity DNA polymerase with HF buffer (New England Biolabs; NEB) according to the manufacturer’s instructions with modifications. ..

    Article Title: Solid-phase cloning for high-throughput assembly of single and multiple DNA parts
    Article Snippet: .. DNA parts were amplified with Phusion® Hot-Start Flex (2 U/μl, New England Biolabs, Ipswich, MA, USA) by the following PCR program: 98°C for 30 s, 30 cycles of 98°C 8 s, 25 s annealing with temperature depending on primer melting temperature and 72°C for 20 s/kb, before ending with 72°C for 7 min followed by 4°C hold. ..

    Article Title: A Flexible Approach for Highly Multiplexed Candidate Gene Targeted Resequencing
    Article Snippet: .. After a brief purification using the Spin-20 columns (Princeton Separations), the captured DNA pool was amplified by PCR (98°C for 30 seconds followed by 36–37 cycles at 98°C for 10 seconds, 65°C for 30 seconds, and 73°C for 30 seconds) using Phusion Hot Start High-Fidelity DNA polymerase (New England BioLabs) and non-target specific common primers that are homologous to the vector oligonucleotide. .. After purification using the Fermentas PCR Purification kit, 0.5–1 µg PCR products per sequencing library were ligated to each other using T4 DNA ligase (New England BioLabs).

    Generated:

    Article Title: Recombinase-mediated cassette exchange (RMCE) system for functional genomics studies in Mycoplasma mycoides
    Article Snippet: .. Transformation-Associated Recombination (TAR) cloning of the 100 kb DNA segment A 7.5 kilobase (kb) TAR cloning vector was generated by PCR amplification of pRC60 using two primers (5′- ACTAATAATAAAACATTTATATACTTAATGAATAAATATAATTAG TACCGTTCGTATAATGTATGC-3′ and 5′-ATTTTAAAATTTATGTAATTTATTAATTTTTATCTTTATAATATA TACCGTTCGTATATGGTTTCT-3′) and the Phusion Hot Start High-Fidelity DNA polymerase with HF buffer (New England Biolabs; NEB) according to the manufacturer’s instructions with modifications. ..

    Expressing:

    Article Title: Epitope-tagged protein-based artificial microRNA (ETPamir) screens for optimized gene silencing in plants
    Article Snippet: .. 4-morpholineethanesulfonic acid (MES, Sigma, cat. no. M3671) Mannitol (MP Biomedicals, cat. no. 102248) MgCl2 (Sigma, cat. no. M9272) KCl (Sigma, cat. no. P3911) NaCl (Sigma, cat. no. S9888) CaCl2 (Sigma, cat. no. C7902) CsCl (American Bioanalytical, cat. no. AB00300) Polyethylene glycol 4000 (PEG4000, Sigma, cat. no. 81240) Tween-20 (Sigma, cat. no. P7949) Phusion DNA polymerase (NEB, cat. no. M0535) PVDF membrane (Immobilon, cat. no. IPVH304F0) Nonfat dry milk (Santa Cruz Biotechology, cat. no. sc-2325) Bovine calf serum (HyClone, cat. no. SH30072.03) HA HRP-conjugated antibody (Roche, cat. no. 12013819001) SuperSignal West Pico chemiluminescent kit (Thermo Scientific, cat. no. 34080) SuperSignal West Femto chemiluminescent kit (Thermo Scientific, cat. no. 34095) 10% precast polyacrylamide gel (Bio-Rad, cat. no. 456-1034) Terrific broth (American Bioanalytical, cat. no. AB01966) Custom oligo primers for PCR generating amiRNA precursors (Oligo synthesis service provider) pHBT-HA constitutive expression vector (Available from the authors upon request) pHSP-HA heat shock inducible expression vector (Available from the authors upon request) pHBT-ath-miR319a constitutive expression plasmid (Available from the authors upon request) .. CL2 clinical centrifuge (Thermo Scientific, cat. no. 004260F) Mini-PROTEAN Tetra system (Bio-Rad, cat. no. 165-8006) Trans-Blot SD semi-dry electrophoretic transfer cell (Bio-Rad, cat. no. 170-3940) Fisher Scientific Isotemp heating block (Fisher Scientific, cat. no. 11-715-305Q) 2-ml round-bottom microcentrifuge tubes (USA Scientific, cat. no. 1620-2700) 1.5-ml microcentrifuge tubes (USA Scientific, cat. no. 1615-5500) 6-well culture plates (Falcon, cat. no. 3046) 1000-ml storage bottle with 0.22 μm vacuum filter (Corning, cat. no. 430517)

    Touchdown PCR:

    Article Title: Dimeric CRISPR RNA-Guided FokI-dCas9 Nucleases Directed by Truncated gRNAs for Highly Specific Genome Editing
    Article Snippet: .. Genomic sites were amplified for deep sequencing with Phusion Hot-start FLEX DNA polymerase (NEB) and primers listed in and the touchdown PCR protocol as described above. .. Amplicons of 200–350 bp were purified using Ampure XP beads (Beckman Coulter Genomics) according to manufacturer's instructions.

    Sequencing:

    Article Title: Dimeric CRISPR RNA-Guided FokI-dCas9 Nucleases Directed by Truncated gRNAs for Highly Specific Genome Editing
    Article Snippet: .. Genomic sites were amplified for deep sequencing with Phusion Hot-start FLEX DNA polymerase (NEB) and primers listed in and the touchdown PCR protocol as described above. .. Amplicons of 200–350 bp were purified using Ampure XP beads (Beckman Coulter Genomics) according to manufacturer's instructions.

    Transformation Assay:

    Article Title: Recombinase-mediated cassette exchange (RMCE) system for functional genomics studies in Mycoplasma mycoides
    Article Snippet: .. Transformation-Associated Recombination (TAR) cloning of the 100 kb DNA segment A 7.5 kilobase (kb) TAR cloning vector was generated by PCR amplification of pRC60 using two primers (5′- ACTAATAATAAAACATTTATATACTTAATGAATAAATATAATTAG TACCGTTCGTATAATGTATGC-3′ and 5′-ATTTTAAAATTTATGTAATTTATTAATTTTTATCTTTATAATATA TACCGTTCGTATATGGTTTCT-3′) and the Phusion Hot Start High-Fidelity DNA polymerase with HF buffer (New England Biolabs; NEB) according to the manufacturer’s instructions with modifications. ..

    Oligo Synthesis:

    Article Title: Epitope-tagged protein-based artificial microRNA (ETPamir) screens for optimized gene silencing in plants
    Article Snippet: .. 4-morpholineethanesulfonic acid (MES, Sigma, cat. no. M3671) Mannitol (MP Biomedicals, cat. no. 102248) MgCl2 (Sigma, cat. no. M9272) KCl (Sigma, cat. no. P3911) NaCl (Sigma, cat. no. S9888) CaCl2 (Sigma, cat. no. C7902) CsCl (American Bioanalytical, cat. no. AB00300) Polyethylene glycol 4000 (PEG4000, Sigma, cat. no. 81240) Tween-20 (Sigma, cat. no. P7949) Phusion DNA polymerase (NEB, cat. no. M0535) PVDF membrane (Immobilon, cat. no. IPVH304F0) Nonfat dry milk (Santa Cruz Biotechology, cat. no. sc-2325) Bovine calf serum (HyClone, cat. no. SH30072.03) HA HRP-conjugated antibody (Roche, cat. no. 12013819001) SuperSignal West Pico chemiluminescent kit (Thermo Scientific, cat. no. 34080) SuperSignal West Femto chemiluminescent kit (Thermo Scientific, cat. no. 34095) 10% precast polyacrylamide gel (Bio-Rad, cat. no. 456-1034) Terrific broth (American Bioanalytical, cat. no. AB01966) Custom oligo primers for PCR generating amiRNA precursors (Oligo synthesis service provider) pHBT-HA constitutive expression vector (Available from the authors upon request) pHSP-HA heat shock inducible expression vector (Available from the authors upon request) pHBT-ath-miR319a constitutive expression plasmid (Available from the authors upon request) .. CL2 clinical centrifuge (Thermo Scientific, cat. no. 004260F) Mini-PROTEAN Tetra system (Bio-Rad, cat. no. 165-8006) Trans-Blot SD semi-dry electrophoretic transfer cell (Bio-Rad, cat. no. 170-3940) Fisher Scientific Isotemp heating block (Fisher Scientific, cat. no. 11-715-305Q) 2-ml round-bottom microcentrifuge tubes (USA Scientific, cat. no. 1620-2700) 1.5-ml microcentrifuge tubes (USA Scientific, cat. no. 1615-5500) 6-well culture plates (Falcon, cat. no. 3046) 1000-ml storage bottle with 0.22 μm vacuum filter (Corning, cat. no. 430517)

    Plasmid Preparation:

    Article Title: Epitope-tagged protein-based artificial microRNA (ETPamir) screens for optimized gene silencing in plants
    Article Snippet: .. 4-morpholineethanesulfonic acid (MES, Sigma, cat. no. M3671) Mannitol (MP Biomedicals, cat. no. 102248) MgCl2 (Sigma, cat. no. M9272) KCl (Sigma, cat. no. P3911) NaCl (Sigma, cat. no. S9888) CaCl2 (Sigma, cat. no. C7902) CsCl (American Bioanalytical, cat. no. AB00300) Polyethylene glycol 4000 (PEG4000, Sigma, cat. no. 81240) Tween-20 (Sigma, cat. no. P7949) Phusion DNA polymerase (NEB, cat. no. M0535) PVDF membrane (Immobilon, cat. no. IPVH304F0) Nonfat dry milk (Santa Cruz Biotechology, cat. no. sc-2325) Bovine calf serum (HyClone, cat. no. SH30072.03) HA HRP-conjugated antibody (Roche, cat. no. 12013819001) SuperSignal West Pico chemiluminescent kit (Thermo Scientific, cat. no. 34080) SuperSignal West Femto chemiluminescent kit (Thermo Scientific, cat. no. 34095) 10% precast polyacrylamide gel (Bio-Rad, cat. no. 456-1034) Terrific broth (American Bioanalytical, cat. no. AB01966) Custom oligo primers for PCR generating amiRNA precursors (Oligo synthesis service provider) pHBT-HA constitutive expression vector (Available from the authors upon request) pHSP-HA heat shock inducible expression vector (Available from the authors upon request) pHBT-ath-miR319a constitutive expression plasmid (Available from the authors upon request) .. CL2 clinical centrifuge (Thermo Scientific, cat. no. 004260F) Mini-PROTEAN Tetra system (Bio-Rad, cat. no. 165-8006) Trans-Blot SD semi-dry electrophoretic transfer cell (Bio-Rad, cat. no. 170-3940) Fisher Scientific Isotemp heating block (Fisher Scientific, cat. no. 11-715-305Q) 2-ml round-bottom microcentrifuge tubes (USA Scientific, cat. no. 1620-2700) 1.5-ml microcentrifuge tubes (USA Scientific, cat. no. 1615-5500) 6-well culture plates (Falcon, cat. no. 3046) 1000-ml storage bottle with 0.22 μm vacuum filter (Corning, cat. no. 430517)

    Article Title: Recombinase-mediated cassette exchange (RMCE) system for functional genomics studies in Mycoplasma mycoides
    Article Snippet: .. Transformation-Associated Recombination (TAR) cloning of the 100 kb DNA segment A 7.5 kilobase (kb) TAR cloning vector was generated by PCR amplification of pRC60 using two primers (5′- ACTAATAATAAAACATTTATATACTTAATGAATAAATATAATTAG TACCGTTCGTATAATGTATGC-3′ and 5′-ATTTTAAAATTTATGTAATTTATTAATTTTTATCTTTATAATATA TACCGTTCGTATATGGTTTCT-3′) and the Phusion Hot Start High-Fidelity DNA polymerase with HF buffer (New England Biolabs; NEB) according to the manufacturer’s instructions with modifications. ..

    Article Title: A Flexible Approach for Highly Multiplexed Candidate Gene Targeted Resequencing
    Article Snippet: .. After a brief purification using the Spin-20 columns (Princeton Separations), the captured DNA pool was amplified by PCR (98°C for 30 seconds followed by 36–37 cycles at 98°C for 10 seconds, 65°C for 30 seconds, and 73°C for 30 seconds) using Phusion Hot Start High-Fidelity DNA polymerase (New England BioLabs) and non-target specific common primers that are homologous to the vector oligonucleotide. .. After purification using the Fermentas PCR Purification kit, 0.5–1 µg PCR products per sequencing library were ligated to each other using T4 DNA ligase (New England BioLabs).

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  • 99
    New England Biolabs phusion hot start flex
    Results from head-to-tail SPC. ( A ) Comparison of compatibility of thermostable polymerases with the head-to-tail method. The polymerases with best proof-reading capabilities, <t>Phusion</t> and Deep Vent, all failed to generate transformants with the standard head-to-tail protocol. ( B ) Activity was retained for protocols using Phusion after supplementation of the washing buffer with SDS. ( C ) Colony screens of inserted region representing a selection of assemblies of various lengths and number of inserts assembled by head-to-tail SPC. Final construct sizes spanned 2.9 to 7.2 kbps.
    Phusion Hot Start Flex, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 37 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/phusion hot start flex/product/New England Biolabs
    Average 99 stars, based on 37 article reviews
    Price from $9.99 to $1999.99
    phusion hot start flex - by Bioz Stars, 2020-08
    99/100 stars
      Buy from Supplier

    99
    New England Biolabs hot start phusion high fidelity dna polymerase
    Results from head-to-tail SPC. ( A ) Comparison of compatibility of thermostable polymerases with the head-to-tail method. The polymerases with best proof-reading capabilities, <t>Phusion</t> and Deep Vent, all failed to generate transformants with the standard head-to-tail protocol. ( B ) Activity was retained for protocols using Phusion after supplementation of the washing buffer with SDS. ( C ) Colony screens of inserted region representing a selection of assemblies of various lengths and number of inserts assembled by head-to-tail SPC. Final construct sizes spanned 2.9 to 7.2 kbps.
    Hot Start Phusion High Fidelity Dna Polymerase, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hot start phusion high fidelity dna polymerase/product/New England Biolabs
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    hot start phusion high fidelity dna polymerase - by Bioz Stars, 2020-08
    99/100 stars
      Buy from Supplier

    Image Search Results


    Results from head-to-tail SPC. ( A ) Comparison of compatibility of thermostable polymerases with the head-to-tail method. The polymerases with best proof-reading capabilities, Phusion and Deep Vent, all failed to generate transformants with the standard head-to-tail protocol. ( B ) Activity was retained for protocols using Phusion after supplementation of the washing buffer with SDS. ( C ) Colony screens of inserted region representing a selection of assemblies of various lengths and number of inserts assembled by head-to-tail SPC. Final construct sizes spanned 2.9 to 7.2 kbps.

    Journal: Nucleic Acids Research

    Article Title: Solid-phase cloning for high-throughput assembly of single and multiple DNA parts

    doi: 10.1093/nar/gkv036

    Figure Lengend Snippet: Results from head-to-tail SPC. ( A ) Comparison of compatibility of thermostable polymerases with the head-to-tail method. The polymerases with best proof-reading capabilities, Phusion and Deep Vent, all failed to generate transformants with the standard head-to-tail protocol. ( B ) Activity was retained for protocols using Phusion after supplementation of the washing buffer with SDS. ( C ) Colony screens of inserted region representing a selection of assemblies of various lengths and number of inserts assembled by head-to-tail SPC. Final construct sizes spanned 2.9 to 7.2 kbps.

    Article Snippet: DNA parts were amplified with Phusion® Hot-Start Flex (2 U/μl, New England Biolabs, Ipswich, MA, USA) by the following PCR program: 98°C for 30 s, 30 cycles of 98°C 8 s, 25 s annealing with temperature depending on primer melting temperature and 72°C for 20 s/kb, before ending with 72°C for 7 min followed by 4°C hold.

    Techniques: Activity Assay, Selection, Construct

    Construction outline of the MCP tac s promoter clusters . Fragment 5CP tac s with the flanking sequence was amplified by PCR with p5TG as the template. Fragment 1 was generated by digesting fragment 5CP tac s with BamH I. Fragment 2 was digested from fragment 5CP tac s with BamH I and Hind III. Fragment 3 was linearized from the plasmid p5TG with Hind III. Then, the three fragments were assembled together under the action of T5 exonuclease, Phusion DNA polymerase and Taq DNA ligase in the isothermal process.

    Journal: Microbial Cell Factories

    Article Title: A strategy of gene overexpression based on tandem repetitive promoters in Escherichia coli

    doi: 10.1186/1475-2859-11-19

    Figure Lengend Snippet: Construction outline of the MCP tac s promoter clusters . Fragment 5CP tac s with the flanking sequence was amplified by PCR with p5TG as the template. Fragment 1 was generated by digesting fragment 5CP tac s with BamH I. Fragment 2 was digested from fragment 5CP tac s with BamH I and Hind III. Fragment 3 was linearized from the plasmid p5TG with Hind III. Then, the three fragments were assembled together under the action of T5 exonuclease, Phusion DNA polymerase and Taq DNA ligase in the isothermal process.

    Article Snippet: Then, fragment 1, 2 and 3 were assembled together in vitro under the action of T5 exonuclease (Epicentre), Phusion Hot Start DNA Polymerase (New England Biolabs (NEB)) and Taq DNA ligase (NEB) at 50°C for 15 min.

    Techniques: Sequencing, Amplification, Polymerase Chain Reaction, Generated, Plasmid Preparation