N3041 Search Results


  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 99
    New England Biolabs puc19 vector
    Data analysis of spike-in controls from MethylC-seq and 4mC-TAB-seq in the context of C. kristjanssonii genomic DNA. ( A ) Composition of <t>pUC19</t> DNA, lambda DNA, and C. kristjanssonii genomic DNA. ( B ) The percentage of detected as cytosine reads on 4mC sites in untreated and Tet-treated samples. ( C ) The detected as cytosine reads percentage on unmodified cytosine sites (non-CpG context) and 5mC sites (CpG context) in untreated and Tet-treated samples. ( D ) Quantification of 4mC and 5mC in C. kristjanssonii genomic DNA, determined by LC-MS/MS and deep-sequencing respectively. Error bars indicate mean ± SD, n = 4.
    Puc19 Vector, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 1188 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/puc19 vector/product/New England Biolabs
    Average 99 stars, based on 1188 article reviews
    Price from $9.99 to $1999.99
    puc19 vector - by Bioz Stars, 2020-09
    99/100 stars
      Buy from Supplier

    Image Search Results


    Data analysis of spike-in controls from MethylC-seq and 4mC-TAB-seq in the context of C. kristjanssonii genomic DNA. ( A ) Composition of pUC19 DNA, lambda DNA, and C. kristjanssonii genomic DNA. ( B ) The percentage of detected as cytosine reads on 4mC sites in untreated and Tet-treated samples. ( C ) The detected as cytosine reads percentage on unmodified cytosine sites (non-CpG context) and 5mC sites (CpG context) in untreated and Tet-treated samples. ( D ) Quantification of 4mC and 5mC in C. kristjanssonii genomic DNA, determined by LC-MS/MS and deep-sequencing respectively. Error bars indicate mean ± SD, n = 4.

    Journal: Nucleic Acids Research

    Article Title: Base-resolution detection of N4-methylcytosine in genomic DNA using 4mC-Tet-assisted-bisulfite- sequencing

    doi: 10.1093/nar/gkv738

    Figure Lengend Snippet: Data analysis of spike-in controls from MethylC-seq and 4mC-TAB-seq in the context of C. kristjanssonii genomic DNA. ( A ) Composition of pUC19 DNA, lambda DNA, and C. kristjanssonii genomic DNA. ( B ) The percentage of detected as cytosine reads on 4mC sites in untreated and Tet-treated samples. ( C ) The detected as cytosine reads percentage on unmodified cytosine sites (non-CpG context) and 5mC sites (CpG context) in untreated and Tet-treated samples. ( D ) Quantification of 4mC and 5mC in C. kristjanssonii genomic DNA, determined by LC-MS/MS and deep-sequencing respectively. Error bars indicate mean ± SD, n = 4.

    Article Snippet: To generate the 4mC spike-in control, 0.5 ng of pUC19 vector (NEB) was PCR amplified as follows in a 50 μl reaction: 2.5 U RedTaq polymerase (Sigma), 5 μl 10× reaction buffer, 1 μl 4mdCTP (Trilink)/dATP/dGTP/dTTP cocktail (10 mM each), 1 μl 10 mM forward primer (5′-GCGGTAATACGGTTATCCAC), 1 μl 10 mM reverse primer (5′-TGCTGATAAATCTGGAGCCG).

    Techniques: Lambda DNA Preparation, Liquid Chromatography with Mass Spectroscopy, Mass Spectrometry, Sequencing

    In vivo activities of Streptomyces SerRS1 and SerRS2. s erS1 , s erS2 , E. coli s erS , and the s erS2 ( H270G ) mutant were cloned into pUC19 (pUC) vector under the control of the glnS ). (A) The plasmids were transformed into an E. coli temperature-sensitive

    Journal: Antimicrobial Agents and Chemotherapy

    Article Title: Characterization of Two Seryl-tRNA Synthetases in Albomycin-Producing Streptomyces sp. Strain ATCC 700974 ▿

    doi: 10.1128/AAC.00782-09

    Figure Lengend Snippet: In vivo activities of Streptomyces SerRS1 and SerRS2. s erS1 , s erS2 , E. coli s erS , and the s erS2 ( H270G ) mutant were cloned into pUC19 (pUC) vector under the control of the glnS ). (A) The plasmids were transformed into an E. coli temperature-sensitive

    Article Snippet: The host strain was an E. coli mutant (K28) that is ineffective at making its own serS gene product at high temperature ( , ). serS1 and serS2 were separately cloned into a high-copy vector, pUC19, under the control of a constitutive glnS ′ promoter, which is a glnS mutant promoter used for aaRS expression in E. coli , and the resulting plasmids were transformed into E. coli K28.

    Techniques: In Vivo, Mutagenesis, Clone Assay, Plasmid Preparation, Transformation Assay

    Strategy for converting hSIRPA -BAC DNA into a piggyBac transposon. ( A ) Diagram illustrating the strategy used for retrofitting hSIRPA-BAC DNA (RP11-887J4) with piggyBac TIR elements. 5′ TIR (green) and 3′ TIR (orange) elements were sub-cloned into pUC19 vector backbone with spectinomycin resistance gene (purple), and 50 bp homology arm sequences (red) used for replacing the chloramphenicol resistance gene in the BAC vector backbone via recombineering technology. The diagram also indicates that the genomic DNA insert in the RP11-887J4 BAC is 176,233 bps, covering the SIRPA genic region, on chromosome 20 between 1,842,086-2,018,318. ( B ) The green arrows indicate the primer pairs used to verify hSIRPA-BAC retrofitting after the recombineering process. ( C ) A schematic diagram describing the transpositioning strategy of hSIRPA-BAC retrofitted with TIR elements mediated by piggyBac transposase. Illustration (i) shows the retrofitted BAC DNA. Illustrations (ii) and (iii) show the process by which the piggyBac transposase proteins bind to the TIR sequences, initiating nicking of the DNA strands, allowing 3′ hydroxyl group at both ends of the transposon to hydrophilic attack the flanking TTAA sequence and freeing the BAC from the spectinomycin resistance gene by forming hairpin structure at the TIR ends. Once the BAC DNA is released from spectinomycin resistance gene, illustration (iv) shows repairing of the linearized BAC DNA by ligating into the complementary TTAA overhangs in the genomic DNA through the mediation of the piggyBac transposase proteins.

    Journal: Scientific Reports

    Article Title: Comparative Analysis of piggyBac, CRISPR/Cas9 and TALEN Mediated BAC Transgenesis in the Zygote for the Generation of Humanized SIRPA Rats

    doi: 10.1038/srep31455

    Figure Lengend Snippet: Strategy for converting hSIRPA -BAC DNA into a piggyBac transposon. ( A ) Diagram illustrating the strategy used for retrofitting hSIRPA-BAC DNA (RP11-887J4) with piggyBac TIR elements. 5′ TIR (green) and 3′ TIR (orange) elements were sub-cloned into pUC19 vector backbone with spectinomycin resistance gene (purple), and 50 bp homology arm sequences (red) used for replacing the chloramphenicol resistance gene in the BAC vector backbone via recombineering technology. The diagram also indicates that the genomic DNA insert in the RP11-887J4 BAC is 176,233 bps, covering the SIRPA genic region, on chromosome 20 between 1,842,086-2,018,318. ( B ) The green arrows indicate the primer pairs used to verify hSIRPA-BAC retrofitting after the recombineering process. ( C ) A schematic diagram describing the transpositioning strategy of hSIRPA-BAC retrofitted with TIR elements mediated by piggyBac transposase. Illustration (i) shows the retrofitted BAC DNA. Illustrations (ii) and (iii) show the process by which the piggyBac transposase proteins bind to the TIR sequences, initiating nicking of the DNA strands, allowing 3′ hydroxyl group at both ends of the transposon to hydrophilic attack the flanking TTAA sequence and freeing the BAC from the spectinomycin resistance gene by forming hairpin structure at the TIR ends. Once the BAC DNA is released from spectinomycin resistance gene, illustration (iv) shows repairing of the linearized BAC DNA by ligating into the complementary TTAA overhangs in the genomic DNA through the mediation of the piggyBac transposase proteins.

    Article Snippet: The PCR amplified fragments were subcloned into a pUC19 vector backbone with neomycin resistance gene driven by PGK and EM7 as a selection cassette using the Gibson Assembly kit (NEB).

    Techniques: BAC Assay, Clone Assay, Plasmid Preparation, Sequencing

    GlpG specifically targets orphan HybA and does not influence hydrogenase‐2 activity Bacterial two‐hybrid analysis with HybA and/or HybB fused chromosomally with the T25 and T18 domains of B. pertussis CyaA, respectively, in the absence of endogenous CyaA. Bacteria were grown on LB agar containing 20 μg/ml X‐gal at 37°C for 14 h in the presence/absence of O 2 . Scale bar, 1 cm. Western blot analysis to detect HybA cleavage in S. sonnei Δ rhom7 by wild‐type (+) or inactive (S 201 A) GlpG expressed chromosomally (native) or from pUC19 (plasmid) with (+)/without (−) HybB. HybA was expressed from its native locus or a plasmid (pHybA). Quantification of the ratio of cleaved/uncleaved HybA in strains with (+) or without (−) HybB. Mean ± S.D. of three experiments. * P

    Journal: The EMBO Journal

    Article Title: Bacterial rhomboid proteases mediate quality control of orphan membrane proteins

    doi: 10.15252/embj.2019102922

    Figure Lengend Snippet: GlpG specifically targets orphan HybA and does not influence hydrogenase‐2 activity Bacterial two‐hybrid analysis with HybA and/or HybB fused chromosomally with the T25 and T18 domains of B. pertussis CyaA, respectively, in the absence of endogenous CyaA. Bacteria were grown on LB agar containing 20 μg/ml X‐gal at 37°C for 14 h in the presence/absence of O 2 . Scale bar, 1 cm. Western blot analysis to detect HybA cleavage in S. sonnei Δ rhom7 by wild‐type (+) or inactive (S 201 A) GlpG expressed chromosomally (native) or from pUC19 (plasmid) with (+)/without (−) HybB. HybA was expressed from its native locus or a plasmid (pHybA). Quantification of the ratio of cleaved/uncleaved HybA in strains with (+) or without (−) HybB. Mean ± S.D. of three experiments. * P

    Article Snippet: Specific point mutations or addition of a sequence encoding a 3xHA tag was introduced by Gibson cloning (New England Biolabs) with appropriate primers. pUC19‐based vectors (Yanisch‐Perron et al , ) were constructed by assembling appropriate glpG fragments into Sph I/Hind III‐linearised pUC19 by NEBuilder, and Gibson cloning was used to introduce specific point mutations or sequences encoding a triple‐HA tag. pLAC101 is a low‐copy‐number plasmid for expressing proteins under IPTG induction. pLAC101 contains the pSC101 origin (amplified with pGL1315/pGL1316 from pUA139) (Zaslaver et al , ), the kanamycin resistance gene and lacI with a cloning site amplified from pNCC1 (Wormann et al , ) with primers pGL897/pGL1314 and pGL894/pGL1314, respectively. pLAC101V5‐hybA /V5‐hybA P300A were generated by assembling appropriate fragments into Pac I/Kpn I‐linearised pLAC101.

    Techniques: Activity Assay, Western Blot, Plasmid Preparation

    Rhomboids prevent aggregation of orphan substrates in the inner membrane Western blot analysis probing the localisation and status of plasmid‐encoded N‐terminally V5‐tagged wild‐type (WT) or modified (P 300 A) HybA in S. sonnei Δ rhom7 Δ hybB with wild‐type (+) or inactive (SAHA) GlpG expressed from pUC19. Whole‐cell lysate (Whole cell.), Soluble (Sol.), Membrane (detergent‐solubilised, Mem.) and the Aggregate (Agg.) fractions are shown. HybA that is uncleaved, cleaved only by GlpG and further degraded post‐GlpG cleavage is marked by black, red and green arrows, respectively. Model of rhomboid‐mediated quality control by selectively targeting orphan components of multiprotein respiratory complexes. Source data are available online for this figure.

    Journal: The EMBO Journal

    Article Title: Bacterial rhomboid proteases mediate quality control of orphan membrane proteins

    doi: 10.15252/embj.2019102922

    Figure Lengend Snippet: Rhomboids prevent aggregation of orphan substrates in the inner membrane Western blot analysis probing the localisation and status of plasmid‐encoded N‐terminally V5‐tagged wild‐type (WT) or modified (P 300 A) HybA in S. sonnei Δ rhom7 Δ hybB with wild‐type (+) or inactive (SAHA) GlpG expressed from pUC19. Whole‐cell lysate (Whole cell.), Soluble (Sol.), Membrane (detergent‐solubilised, Mem.) and the Aggregate (Agg.) fractions are shown. HybA that is uncleaved, cleaved only by GlpG and further degraded post‐GlpG cleavage is marked by black, red and green arrows, respectively. Model of rhomboid‐mediated quality control by selectively targeting orphan components of multiprotein respiratory complexes. Source data are available online for this figure.

    Article Snippet: Specific point mutations or addition of a sequence encoding a 3xHA tag was introduced by Gibson cloning (New England Biolabs) with appropriate primers. pUC19‐based vectors (Yanisch‐Perron et al , ) were constructed by assembling appropriate glpG fragments into Sph I/Hind III‐linearised pUC19 by NEBuilder, and Gibson cloning was used to introduce specific point mutations or sequences encoding a triple‐HA tag. pLAC101 is a low‐copy‐number plasmid for expressing proteins under IPTG induction. pLAC101 contains the pSC101 origin (amplified with pGL1315/pGL1316 from pUA139) (Zaslaver et al , ), the kanamycin resistance gene and lacI with a cloning site amplified from pNCC1 (Wormann et al , ) with primers pGL897/pGL1314 and pGL894/pGL1314, respectively. pLAC101V5‐hybA /V5‐hybA P300A were generated by assembling appropriate fragments into Pac I/Kpn I‐linearised pLAC101.

    Techniques: Western Blot, Plasmid Preparation, Modification

    Rhomboid cleavage licenses further degradation of substrates Western blot analysis (probing with an anti‐V5 mAb) to detect degradation of N‐terminally V5‐tagged wild‐type (WT) or modified (P 300 A) HybA in S. sonnei Δ rhom7 chromosomally expressing wild‐type (+) or inactive (S 201 A) GlpG with (+) or without (−) HybB. Degradation of V5‐tagged HybA at times after blocking protein translation at T 0 in the presence (+) or absence (−) of HybB. Western blot analysis of N‐terminally V5‐tagged wild‐type (WT) or modified (P 259 A) FdoH in S. sonnei Δ rhom7 chromosomally expressing wild‐type (+) or inactive (S 201 A) GlpG with (+)/without (−) FdoI. Western blot analysis of N‐terminally V5‐tagged wild‐type (WT) or modified (P 259 A) FdnH in S. sonnei Δ rhom7 with wild‐type (+) or inactive (SAHA) Rhom7 expressed from pBAD33 with (+)/without (−) FdnI. Degradation of N‐terminally V5‐tagged wild‐type (WT) or modified (P 259 A) FdoH (E) or FdnH (F) in S. sonnei Δ rhom7 with wild‐type (+) or inactive (S 201 A, SAHA) GlpG expressed chromosomally (native) or from pUC19 (plasmid) without FdoI or FdnI (−), respectively, +/− exposure to 400 μM CuCl 2 for 30 min. Data information: Rhomboid substrates that are uncleaved, cleaved by GlpG or cleaved by Rhom7 are marked by black, red and blue arrows, respectively. Degradation products post‐rhomboid cleavage are marked by green arrows. RecA, loading control. Source data are available online for this figure.

    Journal: The EMBO Journal

    Article Title: Bacterial rhomboid proteases mediate quality control of orphan membrane proteins

    doi: 10.15252/embj.2019102922

    Figure Lengend Snippet: Rhomboid cleavage licenses further degradation of substrates Western blot analysis (probing with an anti‐V5 mAb) to detect degradation of N‐terminally V5‐tagged wild‐type (WT) or modified (P 300 A) HybA in S. sonnei Δ rhom7 chromosomally expressing wild‐type (+) or inactive (S 201 A) GlpG with (+) or without (−) HybB. Degradation of V5‐tagged HybA at times after blocking protein translation at T 0 in the presence (+) or absence (−) of HybB. Western blot analysis of N‐terminally V5‐tagged wild‐type (WT) or modified (P 259 A) FdoH in S. sonnei Δ rhom7 chromosomally expressing wild‐type (+) or inactive (S 201 A) GlpG with (+)/without (−) FdoI. Western blot analysis of N‐terminally V5‐tagged wild‐type (WT) or modified (P 259 A) FdnH in S. sonnei Δ rhom7 with wild‐type (+) or inactive (SAHA) Rhom7 expressed from pBAD33 with (+)/without (−) FdnI. Degradation of N‐terminally V5‐tagged wild‐type (WT) or modified (P 259 A) FdoH (E) or FdnH (F) in S. sonnei Δ rhom7 with wild‐type (+) or inactive (S 201 A, SAHA) GlpG expressed chromosomally (native) or from pUC19 (plasmid) without FdoI or FdnI (−), respectively, +/− exposure to 400 μM CuCl 2 for 30 min. Data information: Rhomboid substrates that are uncleaved, cleaved by GlpG or cleaved by Rhom7 are marked by black, red and blue arrows, respectively. Degradation products post‐rhomboid cleavage are marked by green arrows. RecA, loading control. Source data are available online for this figure.

    Article Snippet: Specific point mutations or addition of a sequence encoding a 3xHA tag was introduced by Gibson cloning (New England Biolabs) with appropriate primers. pUC19‐based vectors (Yanisch‐Perron et al , ) were constructed by assembling appropriate glpG fragments into Sph I/Hind III‐linearised pUC19 by NEBuilder, and Gibson cloning was used to introduce specific point mutations or sequences encoding a triple‐HA tag. pLAC101 is a low‐copy‐number plasmid for expressing proteins under IPTG induction. pLAC101 contains the pSC101 origin (amplified with pGL1315/pGL1316 from pUA139) (Zaslaver et al , ), the kanamycin resistance gene and lacI with a cloning site amplified from pNCC1 (Wormann et al , ) with primers pGL897/pGL1314 and pGL894/pGL1314, respectively. pLAC101V5‐hybA /V5‐hybA P300A were generated by assembling appropriate fragments into Pac I/Kpn I‐linearised pLAC101.

    Techniques: Western Blot, Modification, Expressing, Blocking Assay, Plasmid Preparation