bsteii hf  (New England Biolabs)


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  • 93
    Name:
    BstEII HF
    Description:
    BstEII HF 10 000 units
    Catalog Number:
    R3162L
    Price:
    257
    Category:
    Restriction Enzymes
    Size:
    10 000 units
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    New England Biolabs bsteii hf
    BstEII HF
    BstEII HF 10 000 units
    https://www.bioz.com/result/bsteii hf/product/New England Biolabs
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    bsteii hf - by Bioz Stars, 2021-06
    93/100 stars

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    Related Articles

    Plasmid Preparation:

    Article Title: Highly parallel genome variant engineering with CRISPR/Cas9
    Article Snippet: The amplified library was cloned into pLK88, a version of pLK78 modified to include the BstEII and SphI sites. pLK88 was isolated with a QIAGEN Plasmid Plus Maxiprep kit (Qiagen, Hilden, Germany) from 200 milliliters of Escherichia coli culture. .. 20 micrograms of plasmid was then digested by BstEII-HF and EagI-HF, treated with Shrimp Alkaline Phosphatase (New England Biolabs, Ipswich, MA), and purified with the Qiagen PCR purification kit. .. We tested two ligation reactions: 1 microgram of digested vector was ligated with either 100 nanograms or 800 nanograms of the digested insert, with 4 microliters of T4 DNA Ligase M0202M (New England Biolabs, Ipswich, MA), in an 800 or 200 microliter reaction, respectively, at room temperature for 10 minutes.

    Article Title: Tighter αC-helix–αL16-helix interactions seem to make p38α less prone to activation by autophosphorylation than Hog1
    Article Snippet: Plasmids constructions and mutagenesis procedures Site-directed mutagenesis was performed according to the manufacturer's instructions, using the PfuUltraII Fusion HotStart DNA Polymerase (Agilent # 600670) and pBluescriptIISK+ containing the relevant DNA as a template. .. Then, the DNA was cloned to the vector of interest, using the BstEII (NEB # R3162S) and BlpI (NEB # R0585S) restriction enzymes for HOG1 and the EcoRI (NEB #R3101S) restriction enzyme for p38α. .. All HOG1 molecules were expressed from the pES86 vector as N-terminal 3X-HemeAgluttinin (3XHA)-tagged proteins, as described previously [ ].

    Purification:

    Article Title: Highly parallel genome variant engineering with CRISPR/Cas9
    Article Snippet: The amplified library was cloned into pLK88, a version of pLK78 modified to include the BstEII and SphI sites. pLK88 was isolated with a QIAGEN Plasmid Plus Maxiprep kit (Qiagen, Hilden, Germany) from 200 milliliters of Escherichia coli culture. .. 20 micrograms of plasmid was then digested by BstEII-HF and EagI-HF, treated with Shrimp Alkaline Phosphatase (New England Biolabs, Ipswich, MA), and purified with the Qiagen PCR purification kit. .. We tested two ligation reactions: 1 microgram of digested vector was ligated with either 100 nanograms or 800 nanograms of the digested insert, with 4 microliters of T4 DNA Ligase M0202M (New England Biolabs, Ipswich, MA), in an 800 or 200 microliter reaction, respectively, at room temperature for 10 minutes.

    Article Title: Generation and characterization of CD1d‐specific single‐domain antibodies with distinct functional features
    Article Snippet: For this purpose total RNA was extracted from the peripheral blood lymphocyte, transcribed into cDNA, purified and used as a template for immunoglobulin heavy‐chain‐encoding gene amplification, as described previously. .. Agarose gel electrophoresed purified genes encoding heavy‐chain‐only immunoglobulin (~ 700 bp) were digested with Sfi I and Bst EII (catalogue nos R0123L, R3162L; New England Biolabs, Ipswich, MA) followed by cDNA VHH gene (~ 300–400 bp) extraction through agarose gel electrophoresis. .. Isolated VHH genes were subsequently ligated into the phagemid vector pUR8100 (a derivate of pHen1 with addition of an HC‐V cassette, to enable Sfi I‐ Bst EII cloning, conferring Amp‐resistance for selection, and encoding a C‐terminal Myc and His6 tag for detection (kind gift from Dr M. El Khattabi, QVQ, Utrecht, the Netherlands) and transformed into Escherichia coli TG1 for display on filamentous bacteriophage.

    Article Title: Characterizing Locus Specific Chromatin Structure and Dynamics with Correlative Conventional and Super Resolution imaging in living cells
    Article Snippet: A PCR was performed with eGFP using Thermofisher Platinum Taq High Fidelity to add BsteII and AflII restriction enzyme sites on the N-terminus and C-terminus of GFP resp ectively with the forward primer: AGTCAGCTAGGAGgtgacccaggagctcccaagaaaaagcgcaaggtaggtagttccgtgagcaagggcgaggagcta and reverse primer GCTGATCAGCGGTTTAAACttaagtttacttgtacagctcgtccatgccgag. .. The purified PCR product and CMV-dCas9-VP64 backbone were digested using NEB BsteII-HF and AflII-HF restriction enzymes, gel purified and ligated using the NEB T4 DNA ligase kit. .. Ligated plasmids were amplified with DH5α competent cells and purified using Qiagen Miniprep kit. mEos2 (obtained from Davidson Lab) was cloned into a CMV-MCP-YFP plasmid (from Mazhar Adli from Addgene) to replace MCP-YFP .

    Article Title: Highly parallel genome variant engineering with CRISPR/Cas9
    Article Snippet: The amplification primers were designed to introduce an EagI cut site into the 3’ end of the amplification product ( , primers named OLS Library Amplification F and R). .. Reactions were stopped during linear amplification, and the amplified library was then purified with the QIAquick PCR purification kit (Qiagen, Hilden, Germany), digested with BstEII-HF and EagI-HF (New England Biolab, Ipswich, MA), and purified again. .. The amplified library was cloned into pLK88, a version of pLK78 modified to include the BstEII and SphI sites. pLK88 was isolated with a QIAGEN Plasmid Plus Maxiprep kit (Qiagen, Hilden, Germany) from 200 milliliters of Escherichia coli culture.

    Polymerase Chain Reaction:

    Article Title: Highly parallel genome variant engineering with CRISPR/Cas9
    Article Snippet: The amplified library was cloned into pLK88, a version of pLK78 modified to include the BstEII and SphI sites. pLK88 was isolated with a QIAGEN Plasmid Plus Maxiprep kit (Qiagen, Hilden, Germany) from 200 milliliters of Escherichia coli culture. .. 20 micrograms of plasmid was then digested by BstEII-HF and EagI-HF, treated with Shrimp Alkaline Phosphatase (New England Biolabs, Ipswich, MA), and purified with the Qiagen PCR purification kit. .. We tested two ligation reactions: 1 microgram of digested vector was ligated with either 100 nanograms or 800 nanograms of the digested insert, with 4 microliters of T4 DNA Ligase M0202M (New England Biolabs, Ipswich, MA), in an 800 or 200 microliter reaction, respectively, at room temperature for 10 minutes.

    Article Title: Characterizing Locus Specific Chromatin Structure and Dynamics with Correlative Conventional and Super Resolution imaging in living cells
    Article Snippet: A PCR was performed with eGFP using Thermofisher Platinum Taq High Fidelity to add BsteII and AflII restriction enzyme sites on the N-terminus and C-terminus of GFP resp ectively with the forward primer: AGTCAGCTAGGAGgtgacccaggagctcccaagaaaaagcgcaaggtaggtagttccgtgagcaagggcgaggagcta and reverse primer GCTGATCAGCGGTTTAAACttaagtttacttgtacagctcgtccatgccgag. .. The purified PCR product and CMV-dCas9-VP64 backbone were digested using NEB BsteII-HF and AflII-HF restriction enzymes, gel purified and ligated using the NEB T4 DNA ligase kit. .. Ligated plasmids were amplified with DH5α competent cells and purified using Qiagen Miniprep kit. mEos2 (obtained from Davidson Lab) was cloned into a CMV-MCP-YFP plasmid (from Mazhar Adli from Addgene) to replace MCP-YFP .

    Article Title: Highly parallel genome variant engineering with CRISPR/Cas9
    Article Snippet: The amplification primers were designed to introduce an EagI cut site into the 3’ end of the amplification product ( , primers named OLS Library Amplification F and R). .. Reactions were stopped during linear amplification, and the amplified library was then purified with the QIAquick PCR purification kit (Qiagen, Hilden, Germany), digested with BstEII-HF and EagI-HF (New England Biolab, Ipswich, MA), and purified again. .. The amplified library was cloned into pLK88, a version of pLK78 modified to include the BstEII and SphI sites. pLK88 was isolated with a QIAGEN Plasmid Plus Maxiprep kit (Qiagen, Hilden, Germany) from 200 milliliters of Escherichia coli culture.

    Agarose Gel Electrophoresis:

    Article Title: Generation and characterization of CD1d‐specific single‐domain antibodies with distinct functional features
    Article Snippet: For this purpose total RNA was extracted from the peripheral blood lymphocyte, transcribed into cDNA, purified and used as a template for immunoglobulin heavy‐chain‐encoding gene amplification, as described previously. .. Agarose gel electrophoresed purified genes encoding heavy‐chain‐only immunoglobulin (~ 700 bp) were digested with Sfi I and Bst EII (catalogue nos R0123L, R3162L; New England Biolabs, Ipswich, MA) followed by cDNA VHH gene (~ 300–400 bp) extraction through agarose gel electrophoresis. .. Isolated VHH genes were subsequently ligated into the phagemid vector pUR8100 (a derivate of pHen1 with addition of an HC‐V cassette, to enable Sfi I‐ Bst EII cloning, conferring Amp‐resistance for selection, and encoding a C‐terminal Myc and His6 tag for detection (kind gift from Dr M. El Khattabi, QVQ, Utrecht, the Netherlands) and transformed into Escherichia coli TG1 for display on filamentous bacteriophage.

    Clone Assay:

    Article Title: Tighter αC-helix–αL16-helix interactions seem to make p38α less prone to activation by autophosphorylation than Hog1
    Article Snippet: Plasmids constructions and mutagenesis procedures Site-directed mutagenesis was performed according to the manufacturer's instructions, using the PfuUltraII Fusion HotStart DNA Polymerase (Agilent # 600670) and pBluescriptIISK+ containing the relevant DNA as a template. .. Then, the DNA was cloned to the vector of interest, using the BstEII (NEB # R3162S) and BlpI (NEB # R0585S) restriction enzymes for HOG1 and the EcoRI (NEB #R3101S) restriction enzyme for p38α. .. All HOG1 molecules were expressed from the pES86 vector as N-terminal 3X-HemeAgluttinin (3XHA)-tagged proteins, as described previously [ ].

    Amplification:

    Article Title: Highly parallel genome variant engineering with CRISPR/Cas9
    Article Snippet: The amplification primers were designed to introduce an EagI cut site into the 3’ end of the amplification product ( , primers named OLS Library Amplification F and R). .. Reactions were stopped during linear amplification, and the amplified library was then purified with the QIAquick PCR purification kit (Qiagen, Hilden, Germany), digested with BstEII-HF and EagI-HF (New England Biolab, Ipswich, MA), and purified again. .. The amplified library was cloned into pLK88, a version of pLK78 modified to include the BstEII and SphI sites. pLK88 was isolated with a QIAGEN Plasmid Plus Maxiprep kit (Qiagen, Hilden, Germany) from 200 milliliters of Escherichia coli culture.

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  • 93
    New England Biolabs bsteii hf
    Bsteii Hf, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/bsteii hf/product/New England Biolabs
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    bsteii hf - by Bioz Stars, 2021-06
    93/100 stars
      Buy from Supplier

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