bsrgi  (New England Biolabs)


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    Name:
    BsrGI
    Description:
    BsrGI 5 000 units
    Catalog Number:
    r0575l
    Price:
    277
    Size:
    5 000 units
    Category:
    Restriction Enzymes
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    Structured Review

    New England Biolabs bsrgi
    BsrGI
    BsrGI 5 000 units
    https://www.bioz.com/result/bsrgi/product/New England Biolabs
    Average 90 stars, based on 3 article reviews
    Price from $9.99 to $1999.99
    bsrgi - by Bioz Stars, 2020-01
    90/100 stars

    Images

    1) Product Images from "Asparagine requirement in Plasmodium berghei as a target to prevent malaria transmission and liver infections"

    Article Title: Asparagine requirement in Plasmodium berghei as a target to prevent malaria transmission and liver infections

    Journal: Nature Communications

    doi: 10.1038/ncomms9775

    Generation of AS knockout in P. berghei . ( a ) Double cross-over recombination strategy followed to generate Pb ASKO. ( b ) PCR analysis for genomic DNA isolated from Pb WT and Pb ASKO parasites. Lane 1 and 3: PCR with Pb AS-specific primers; Lane 2 and 4: PCR with Pb GAPDH-specific primers (control); Lane M: 1 kb DNA ladder. ( c ) RT-PCR analysis for RNA isolated from Pb WT and Pb ASKO parasites. Lane 1 and 3: RT-PCR with Pb AS-specific primers; Lane 2 and 4: RT-PCR with Pb GAPDH specific primers (control); Lane M: 1 kb DNA ladder. ( d ) Southern analysis for genomic DNA isolated from Pb WT and Pb ASKO parasites. Transgenic plasmid (TP) was also included as control to rule out the presence of any episomes. Genomic DNA preparations and TP were digested with BsrGI and XbaI followed by hybridization with 5′-UTR specific probe. ( e ) Northern analysis for RNA isolated from Pb WT and Pb ASKO parasites indicating the absence of AS mRNA (1.75 kb) in Pb ASKO. For control, GAPDH (1.01 kb; lower panel) was used (full blots are shown in Supplementary Fig. 6 ). ( f ) Western blot analysis indicating the absence of AS (67 kDa) in Pb ASKO parasite lysate ( Supplementary Fig. 6 ). Parasite lysates of Pb WT and Pb ASKO containing 100 μg of total protein were used. For control, hsp60 (60 kDa) was used (lower panel). ( g ) Enzyme assays for Pb WT and Pb ASKO parasites using [U- 14 C]-aspartate. 200 μg of total protein was used per assay.
    Figure Legend Snippet: Generation of AS knockout in P. berghei . ( a ) Double cross-over recombination strategy followed to generate Pb ASKO. ( b ) PCR analysis for genomic DNA isolated from Pb WT and Pb ASKO parasites. Lane 1 and 3: PCR with Pb AS-specific primers; Lane 2 and 4: PCR with Pb GAPDH-specific primers (control); Lane M: 1 kb DNA ladder. ( c ) RT-PCR analysis for RNA isolated from Pb WT and Pb ASKO parasites. Lane 1 and 3: RT-PCR with Pb AS-specific primers; Lane 2 and 4: RT-PCR with Pb GAPDH specific primers (control); Lane M: 1 kb DNA ladder. ( d ) Southern analysis for genomic DNA isolated from Pb WT and Pb ASKO parasites. Transgenic plasmid (TP) was also included as control to rule out the presence of any episomes. Genomic DNA preparations and TP were digested with BsrGI and XbaI followed by hybridization with 5′-UTR specific probe. ( e ) Northern analysis for RNA isolated from Pb WT and Pb ASKO parasites indicating the absence of AS mRNA (1.75 kb) in Pb ASKO. For control, GAPDH (1.01 kb; lower panel) was used (full blots are shown in Supplementary Fig. 6 ). ( f ) Western blot analysis indicating the absence of AS (67 kDa) in Pb ASKO parasite lysate ( Supplementary Fig. 6 ). Parasite lysates of Pb WT and Pb ASKO containing 100 μg of total protein were used. For control, hsp60 (60 kDa) was used (lower panel). ( g ) Enzyme assays for Pb WT and Pb ASKO parasites using [U- 14 C]-aspartate. 200 μg of total protein was used per assay.

    Techniques Used: Knock-Out, Polymerase Chain Reaction, Isolation, Reverse Transcription Polymerase Chain Reaction, Transgenic Assay, Plasmid Preparation, Hybridization, Northern Blot, Western Blot

    Related Articles

    Clone Assay:

    Article Title: High-Throughput System for the Presentation of Secreted and Surface-Exposed Proteins from Gram-Positive Bacteria in Functional Metagenomics Studies
    Article Snippet: The amplification products were cloned in pDONR223 (Invitrogen) by site-specific recombination between the att B1.1 and att B2.1 sites and the att P1 and att P2 sites, respectively, present on the plasmid, in vitro, using BP clonase II (Invitrogen) according to the recommendations of the supplier, followed by transformation of OneShotR TOP10 chemically competent E. coli (Invitrogen) with selection for spectinomycin (this E. coli strain allowing the counter selection of pDONR223 without insert). .. The presence of inserts in pDONR223 and pDG148-GW was verified by digestion with Bsr GI and Eco RI (New England Biolabs), respectively, and PCR amplifications using the pDONR223-specific primers 5′-CCCAGTCACGACGTTGTAAAACG and 5′-GTAACATCAGAGATTTTGAGACAC .

    Article Title: Directional cDNA library construction assisted by the in vitro recombination reaction
    Article Snippet: The resultant plasmid, attB-pBluescript, was used as a template for preparation of a DNA fragment containing multiple cloning sites (MCS). .. MCS, Tet and βGal thus obtained were digested with Bsr GI (New England BioLabs), which cleaves at the TGTACA sequence within the common 15 bp core region of each att sequence, leaving four-base 5′ overhangs.

    Article Title: Haplotype-specific modulation of a SOX10/CREB response element at the Charcot–Marie–Tooth disease type 4C locus SH3TC2
    Article Snippet: .. Successful cloning of each genomic segment upstream of the luciferase reporter gene was assessed by genotyping with Bsr GI (New England Biolabs). .. Site-directed mutagenesis reactions were performed using the QuikChange II Mutagenesis Kit (Agilent Technologies) and appropriate mutation-bearing primers ( ).

    Article Title: Highly Significant Antiviral Activity of HIV-1 LTR-Specific Tre-Recombinase in Humanized Mice
    Article Snippet: The respective genomic sites were cloned into the recombination reporter plasmid pEVO-Tre-target . .. Plasmid DNA was isolated from overnight cultures and digested with BsrG I and Xba I (NEB), resulting in different fragment sizes for recombined versus non-recombined substrate on agarose gels.

    Article Title: Dimerization is required for GARS-mediated neurotoxicity in dominant CMT disease
    Article Snippet: .. Resulting clones were purified and digested with Bsr GI (New England Biolabs, Ipswich, MA, USA) to confirm recombination. .. The Δ grs1 haploid yeast strain [harboring a pRS316 maintenance vector to express wild-type GRS1 and URA3 ( ) was transformed with wild-type or mutant GRS1 in a LEU2 -bearing pRS315 vector and selected on medium lacking uracil and leucine (Teknova, Hollister, CA, USA).

    Article Title: A novel AARS mutation in a family with dominant myeloneuropathy
    Article Snippet: .. Resulting clones were purified and digested with Bsr GI (New England Biolabs, Ipswich, MA) to confirm recombination. .. The Δ ala1 haploid yeast strain (harboring a pRS316 maintenance vector to express wild-type ALA1 and URA3 ) was transformed with wild-type or mutant ALA1 in a LEU2 -bearing pRS315 vector and selected on medium lacking uracil and leucine (Teknova, Hollister, CA).

    Article Title: Haplotype-specific modulation of a SOX10/CREB response element at the Charcot–Marie–Tooth disease type 4C locus SH3TC2
    Article Snippet: Subsequent to PCR amplification and purification, each genomic segment was cloned into the pDONR221 vector using BP Clonase (Invitrogen). .. Resulting constructs were genotyped by digestion with Bsr GI (New England Biolabs) and subjected to DNA sequence analysis to ensure sequence specificity.

    Amplification:

    Article Title: High-Throughput System for the Presentation of Secreted and Surface-Exposed Proteins from Gram-Positive Bacteria in Functional Metagenomics Studies
    Article Snippet: The amplification products were cloned in pDONR223 (Invitrogen) by site-specific recombination between the att B1.1 and att B2.1 sites and the att P1 and att P2 sites, respectively, present on the plasmid, in vitro, using BP clonase II (Invitrogen) according to the recommendations of the supplier, followed by transformation of OneShotR TOP10 chemically competent E. coli (Invitrogen) with selection for spectinomycin (this E. coli strain allowing the counter selection of pDONR223 without insert). .. The presence of inserts in pDONR223 and pDG148-GW was verified by digestion with Bsr GI and Eco RI (New England Biolabs), respectively, and PCR amplifications using the pDONR223-specific primers 5′-CCCAGTCACGACGTTGTAAAACG and 5′-GTAACATCAGAGATTTTGAGACAC .

    Article Title: Equilibrium of sortase A dimerization on Staphylococcus aureus cell surface mediates its cell wall sorting activity
    Article Snippet: .. The amplified 350 bp fragment, which targets the intron to the SrtA sequence, was digested with Hind III and BsrG I (New England Biolabs, Beverly, MA) and inserted into pNL9164 to make pNL9164-srtA. .. The plasmid pNL9164-srtA was then transformed into S. aureus strain RN4220 (general gift from Dr. Alan Lambowitz).

    Article Title: Directional cDNA library construction assisted by the in vitro recombination reaction
    Article Snippet: DNA fragments containing the tetracycline-resistance gene (Tet) and beta-galactosidase gene (βGal) were amplified from control plasmids included in the field test kit provided by Life Technologies using the same set of PCR primers. .. MCS, Tet and βGal thus obtained were digested with Bsr GI (New England BioLabs), which cleaves at the TGTACA sequence within the common 15 bp core region of each att sequence, leaving four-base 5′ overhangs.

    Article Title: Haplotype-specific modulation of a SOX10/CREB response element at the Charcot–Marie–Tooth disease type 4C locus SH3TC2
    Article Snippet: Subsequent to PCR amplification and purification, each genomic segment was cloned into the pDONR221 vector using BP Clonase (Invitrogen). .. Successful cloning of each genomic segment upstream of the luciferase reporter gene was assessed by genotyping with Bsr GI (New England Biolabs).

    Positive Control:

    Article Title: Highly Significant Antiviral Activity of HIV-1 LTR-Specific Tre-Recombinase in Humanized Mice
    Article Snippet: Plasmid DNA was isolated from overnight cultures and digested with BsrG I and Xba I (NEB), resulting in different fragment sizes for recombined versus non-recombined substrate on agarose gels. .. Recombination on the Tre target loxLTR served as positive control.

    Synthesized:

    Article Title: Integration of RNAi and RNA-seq Reveals the Immune Responses of Epinephelus coioides to sigX Gene of Pseudomonas plecoglossicida
    Article Snippet: Five short hairpin RNA sequences targeting the sigX gene were designed and synthesized by Shanghai Generay Biotech Co., Ltd. (Shanghai, China) (Table S1 in Supplementary Material). .. After linearizing pCM130/tac vectors with the restriction enzymes Nsi I and Bsr GI (New England Biolabs, USA), the oligonucleotides were annealed and ligated to the linearized pCM130/tac vectors using T4 DNA ligase (New England Biolabs) following the manufacturer’s recommendations.

    Article Title: Dual RNA-Seq Unveils Pseudomonas plecoglossicida htpG Gene Functions During Host-Pathogen Interactions With Epinephelus coioides
    Article Snippet: Five short hairpin RNA sequences targeting htpG, rplF , and flgD were designed and synthesized by Shanghai Generay Biotech Co., Ltd. (Shanghai, China) ( ). .. After linearizing pCM130/tac vectors with the restriction enzymes Nsi I and Bsr GI (New England Biolabs, U.S.A), the oligonucleotides were annealed and ligated to the linearized pCM130/tac vectors using T4 DNA ligase (New England Biolabs) based on the manufacturer's recommendations.

    Construct:

    Article Title: Integration of RNAi and RNA-seq Reveals the Immune Responses of Epinephelus coioides to sigX Gene of Pseudomonas plecoglossicida
    Article Snippet: RNAi strain was constructed according to methods described by Choi and Schweizer ( ) and Darsigny et al. ( ). .. After linearizing pCM130/tac vectors with the restriction enzymes Nsi I and Bsr GI (New England Biolabs, USA), the oligonucleotides were annealed and ligated to the linearized pCM130/tac vectors using T4 DNA ligase (New England Biolabs) following the manufacturer’s recommendations.

    Article Title: Haplotype-specific modulation of a SOX10/CREB response element at the Charcot–Marie–Tooth disease type 4C locus SH3TC2
    Article Snippet: Each resulting pDONR221 construct was recombined with a destination vector harboring a minimal promoter directing a luciferase reporter gene (pE1B-luciferase) ( ) using LR Clonase (Invitrogen). .. Successful cloning of each genomic segment upstream of the luciferase reporter gene was assessed by genotyping with Bsr GI (New England Biolabs).

    Article Title: Pullulanase Is Necessary for the Efficient Intracellular Growth of Francisella tularensis
    Article Snippet: Construction of a ΔpulB mutant of F . tularensis subsp . tularensis SCHU P9 A pulB gene knockout mutant of SCHU P9 (ΔpulB ) was constructed using a mutagenesis system based on the group II intron of the ltrB gene of Lactobacillus lactis designed to function in F . tularensis and plasmid pKEK1140 (GenBank accession number: EU499313), a kind gift from Dr. Karl E. Klose (South Texas Center for Emerging Infectious Diseases and Department of Biology, University of Texas, San Antonio). .. The PCR product treated with Xho I (New England Biolabs, Beverly, MA, USA) and Bsr GI (New England Biolabs) restriction enzymes was ligated into the pKEK1140 plasmid digested with the same enzymes.

    Article Title: Dual RNA-Seq Unveils Pseudomonas plecoglossicida htpG Gene Functions During Host-Pathogen Interactions With Epinephelus coioides
    Article Snippet: Construction of P. plecoglossicida RNAi Strain RNAi strain was constructed according to previously described methods ( ). .. After linearizing pCM130/tac vectors with the restriction enzymes Nsi I and Bsr GI (New England Biolabs, U.S.A), the oligonucleotides were annealed and ligated to the linearized pCM130/tac vectors using T4 DNA ligase (New England Biolabs) based on the manufacturer's recommendations.

    Article Title: Haplotype-specific modulation of a SOX10/CREB response element at the Charcot–Marie–Tooth disease type 4C locus SH3TC2
    Article Snippet: .. Resulting constructs were genotyped by digestion with Bsr GI (New England Biolabs) and subjected to DNA sequence analysis to ensure sequence specificity. .. Each resulting pDONR221 construct was recombined with a destination vector harboring a minimal promoter directing a luciferase reporter gene (pE1B-luciferase) ( ) using LR Clonase (Invitrogen).

    Electrophoresis:

    Article Title: C7 deficiency in an Irish family: a deletion defect which is predominant in the Irish
    Article Snippet: The restriction enzymes Dde I, Apo I, Rsa I and Bsr GI were obtained from New England Biolabs (St Albans, UK). .. Digests of C7 exon 9 with Dde I and C7 exon 13 with Bsr GI were examined following electrophoresis on 2% agarose/TBE mini-gels.

    Incubation:

    Article Title: BRCA2 controls DNA:RNA hybrid level at DSBs by mediating RNase H2 recruitment
    Article Snippet: Briefly, DNA was extracted gently with phenol:chloroform:isoamyl alcohol (Sigma-Aldrich cat. no. P2069) and digested with Hind III, Eco RI, Bsr GI, Xba I, and Ssp I (NEB). .. In the meantime, serum-free medium containing the S9.6 antibody (kind gift from D. Piccini and M. Foiani) was mixed with protein A and protein G Dynabeads (Invitrogen) and incubated on a rotating wheel overnight at 4 °C.

    Article Title: BRCA2 controls DNA:RNA hybrid level at DSBs by mediating RNase H2 recruitment
    Article Snippet: Briefly, DNA was extracted gently with phenol:chloroform:isoamyl alcohol (Sigma-Aldrich cat. no. P2069) and digested with Hind III, Eco RI, Bsr GI, Xba I, and Ssp I (NEB). .. In the meantime, serum-free medium containing the S9.6 antibody (kind gift from D. Piccini and M. Foiani) was mixed with protein A and protein G Dynabeads (Invitrogen) and incubated on a rotating wheel overnight at 4 °C.

    Article Title: Dimerization is required for GARS-mediated neurotoxicity in dominant CMT disease
    Article Snippet: Resulting clones were purified and digested with Bsr GI (New England Biolabs, Ipswich, MA, USA) to confirm recombination. .. Each colony was grown to saturation in the selection medium for 48 h. Next, 10 μl of undiluted and diluted (1:10 and 1:100) samples from each culture were spotted on plates containing 0.1% 5-FOA complete medium or SD -leu -ura growth medium (Teknova) and incubated at 30°C for 72 h. Yeast cell growth was determined by visual inspection.

    Article Title: A novel AARS mutation in a family with dominant myeloneuropathy
    Article Snippet: Resulting clones were purified and digested with Bsr GI (New England Biolabs, Ipswich, MA) to confirm recombination. .. Next, 10 μL of undiluted and diluted (1:10 and 1:100) samples from each culture were spotted on plates containing 0.1% 5-FOA complete medium or SD -leu -ura growth medium (Teknova) and incubated at 30°C for 72 hours.

    Luciferase:

    Article Title: Haplotype-specific modulation of a SOX10/CREB response element at the Charcot–Marie–Tooth disease type 4C locus SH3TC2
    Article Snippet: .. Successful cloning of each genomic segment upstream of the luciferase reporter gene was assessed by genotyping with Bsr GI (New England Biolabs). .. Site-directed mutagenesis reactions were performed using the QuikChange II Mutagenesis Kit (Agilent Technologies) and appropriate mutation-bearing primers ( ).

    Article Title: Haplotype-specific modulation of a SOX10/CREB response element at the Charcot–Marie–Tooth disease type 4C locus SH3TC2
    Article Snippet: Resulting constructs were genotyped by digestion with Bsr GI (New England Biolabs) and subjected to DNA sequence analysis to ensure sequence specificity. .. Each resulting pDONR221 construct was recombined with a destination vector harboring a minimal promoter directing a luciferase reporter gene (pE1B-luciferase) ( ) using LR Clonase (Invitrogen).

    Cell Culture:

    Article Title: Highly Significant Antiviral Activity of HIV-1 LTR-Specific Tre-Recombinase in Humanized Mice
    Article Snippet: Plasmid DNA was isolated from overnight cultures and digested with BsrG I and Xba I (NEB), resulting in different fragment sizes for recombined versus non-recombined substrate on agarose gels. .. In eukaryotic cell culture, HeLa cells were cotransfected with the reporter plasmids and the expression plasmid pIRESneo-Tre .

    Expressing:

    Article Title: Integration of RNAi and RNA-seq Reveals the Immune Responses of Epinephelus coioides to sigX Gene of Pseudomonas plecoglossicida
    Article Snippet: After linearizing pCM130/tac vectors with the restriction enzymes Nsi I and Bsr GI (New England Biolabs, USA), the oligonucleotides were annealed and ligated to the linearized pCM130/tac vectors using T4 DNA ligase (New England Biolabs) following the manufacturer’s recommendations. .. Finally, the expression level of sigX of each RNAi strain was detected by qRT-PCR.

    Article Title: Haplotype-specific modulation of a SOX10/CREB response element at the Charcot–Marie–Tooth disease type 4C locus SH3TC2
    Article Snippet: Paragraph title: Expression vector construction ... Successful cloning of each genomic segment upstream of the luciferase reporter gene was assessed by genotyping with Bsr GI (New England Biolabs).

    Article Title: Highly Significant Antiviral Activity of HIV-1 LTR-Specific Tre-Recombinase in Humanized Mice
    Article Snippet: In E. coli , recombinase expression was induced with L-arabinose (Sigma-Aldrich) at 1 mg/ml. .. Plasmid DNA was isolated from overnight cultures and digested with BsrG I and Xba I (NEB), resulting in different fragment sizes for recombined versus non-recombined substrate on agarose gels.

    Article Title: Dual RNA-Seq Unveils Pseudomonas plecoglossicida htpG Gene Functions During Host-Pathogen Interactions With Epinephelus coioides
    Article Snippet: After linearizing pCM130/tac vectors with the restriction enzymes Nsi I and Bsr GI (New England Biolabs, U.S.A), the oligonucleotides were annealed and ligated to the linearized pCM130/tac vectors using T4 DNA ligase (New England Biolabs) based on the manufacturer's recommendations. .. Finally, the expression level of the target gene of each RNAi strain was evaluated by qRT-PCR.

    Transformation Assay:

    Article Title: High-Throughput System for the Presentation of Secreted and Surface-Exposed Proteins from Gram-Positive Bacteria in Functional Metagenomics Studies
    Article Snippet: After each transformation, transformant colonies were pooled and plasmids extracted using a QIAprep Spin Miniprep Kit (Qiagen). .. The presence of inserts in pDONR223 and pDG148-GW was verified by digestion with Bsr GI and Eco RI (New England Biolabs), respectively, and PCR amplifications using the pDONR223-specific primers 5′-CCCAGTCACGACGTTGTAAAACG and 5′-GTAACATCAGAGATTTTGAGACAC .

    Article Title: Integration of RNAi and RNA-seq Reveals the Immune Responses of Epinephelus coioides to sigX Gene of Pseudomonas plecoglossicida
    Article Snippet: After linearizing pCM130/tac vectors with the restriction enzymes Nsi I and Bsr GI (New England Biolabs, USA), the oligonucleotides were annealed and ligated to the linearized pCM130/tac vectors using T4 DNA ligase (New England Biolabs) following the manufacturer’s recommendations. .. The recombinant pCM130/tac vectors were transformed into the competent E. coli DH5a cells by heat shock and then were extracted and electroporated into P. plecoglossicida as described previously ( ).

    Article Title: Equilibrium of sortase A dimerization on Staphylococcus aureus cell surface mediates its cell wall sorting activity
    Article Snippet: The amplified 350 bp fragment, which targets the intron to the SrtA sequence, was digested with Hind III and BsrG I (New England Biolabs, Beverly, MA) and inserted into pNL9164 to make pNL9164-srtA. .. The plasmid pNL9164-srtA was then transformed into S. aureus strain RN4220 (general gift from Dr. Alan Lambowitz).

    Article Title: Pullulanase Is Necessary for the Efficient Intracellular Growth of Francisella tularensis
    Article Snippet: The PCR product treated with Xho I (New England Biolabs, Beverly, MA, USA) and Bsr GI (New England Biolabs) restriction enzymes was ligated into the pKEK1140 plasmid digested with the same enzymes. .. After ligation, competent cells of E . coli DH5α strains (Competent high DH5α, Toyobo, Tokyo, Japan) were transformed with the above plasmid.

    Article Title: Dimerization is required for GARS-mediated neurotoxicity in dominant CMT disease
    Article Snippet: Resulting clones were purified and digested with Bsr GI (New England Biolabs, Ipswich, MA, USA) to confirm recombination. .. The Δ grs1 haploid yeast strain [harboring a pRS316 maintenance vector to express wild-type GRS1 and URA3 ( ) was transformed with wild-type or mutant GRS1 in a LEU2 -bearing pRS315 vector and selected on medium lacking uracil and leucine (Teknova, Hollister, CA, USA).

    Article Title: A novel AARS mutation in a family with dominant myeloneuropathy
    Article Snippet: Resulting clones were purified and digested with Bsr GI (New England Biolabs, Ipswich, MA) to confirm recombination. .. The Δ ala1 haploid yeast strain (harboring a pRS316 maintenance vector to express wild-type ALA1 and URA3 ) was transformed with wild-type or mutant ALA1 in a LEU2 -bearing pRS315 vector and selected on medium lacking uracil and leucine (Teknova, Hollister, CA).

    Article Title: Dual RNA-Seq Unveils Pseudomonas plecoglossicida htpG Gene Functions During Host-Pathogen Interactions With Epinephelus coioides
    Article Snippet: After linearizing pCM130/tac vectors with the restriction enzymes Nsi I and Bsr GI (New England Biolabs, U.S.A), the oligonucleotides were annealed and ligated to the linearized pCM130/tac vectors using T4 DNA ligase (New England Biolabs) based on the manufacturer's recommendations. .. The recombinant pCM130/tac vectors were transformed into competent E. coli DH5a cells by heat shock, and then extracted and electroporated into P. plecoglossicida .

    Transfection:

    Article Title: Highly Significant Antiviral Activity of HIV-1 LTR-Specific Tre-Recombinase in Humanized Mice
    Article Snippet: Plasmid DNA was isolated from overnight cultures and digested with BsrG I and Xba I (NEB), resulting in different fragment sizes for recombined versus non-recombined substrate on agarose gels. .. DNA was isolated from the cells 48 h post transfection and analyzed for recombination by polymerase chain reaction using the primers F: 5′- GACAATAACCCTGATAAATGC-3′ , and R: 5′-CCTTAAACGCCTGGTGCTAC-3′ .

    Ligation:

    Article Title: Pullulanase Is Necessary for the Efficient Intracellular Growth of Francisella tularensis
    Article Snippet: The PCR product treated with Xho I (New England Biolabs, Beverly, MA, USA) and Bsr GI (New England Biolabs) restriction enzymes was ligated into the pKEK1140 plasmid digested with the same enzymes. .. After ligation, competent cells of E . coli DH5α strains (Competent high DH5α, Toyobo, Tokyo, Japan) were transformed with the above plasmid.

    Introduce:

    Article Title: Haplotype-specific modulation of a SOX10/CREB response element at the Charcot–Marie–Tooth disease type 4C locus SH3TC2
    Article Snippet: Successful cloning of each genomic segment upstream of the luciferase reporter gene was assessed by genotyping with Bsr GI (New England Biolabs). .. Mutagenesis primers were designed to delete or mutatespecific transcription factor consensus sequences or to introduce specific alleles of SNPs.

    Generated:

    Article Title: Equilibrium of sortase A dimerization on Staphylococcus aureus cell surface mediates its cell wall sorting activity
    Article Snippet: A SrtA knockout strain ( srtA − ) served as the host for assays was generated by using the TargeTron™ Gene Knockout System (Sigma-Aldrich, St. Louis, MO). .. The amplified 350 bp fragment, which targets the intron to the SrtA sequence, was digested with Hind III and BsrG I (New England Biolabs, Beverly, MA) and inserted into pNL9164 to make pNL9164-srtA.

    Polymerase Chain Reaction:

    Article Title: High-Throughput System for the Presentation of Secreted and Surface-Exposed Proteins from Gram-Positive Bacteria in Functional Metagenomics Studies
    Article Snippet: .. The presence of inserts in pDONR223 and pDG148-GW was verified by digestion with Bsr GI and Eco RI (New England Biolabs), respectively, and PCR amplifications using the pDONR223-specific primers 5′-CCCAGTCACGACGTTGTAAAACG and 5′-GTAACATCAGAGATTTTGAGACAC . .. The plasmid pool obtained after LR recombination and transformation was used to transform chemically competent E. coli TG1, in order to generate multimeric plasmids suitable for transformation of competent B. subtilis .

    Article Title: Equilibrium of sortase A dimerization on Staphylococcus aureus cell surface mediates its cell wall sorting activity
    Article Snippet: The primers designated as IBS (5′-AAAAAAGCTTATAATTATCCTTACATATCGATAATGTGCGCCCAGATAGGGTG-3′), EBS1d (5′-CAGATTGTACAAATGTGGTGATAACAGATAAGTCGATAATTATAACTTACCTTTCTTTGT-3′), EBS2 (5′-TGAACGCAAGTTTCTAATTTCGGTTATATGTCGATAGAGGAAAGTGTCT-3′) and EBS universal were used to perform the PCR reaction and amplify a 350 bp fragment from the template pNL9164. .. The amplified 350 bp fragment, which targets the intron to the SrtA sequence, was digested with Hind III and BsrG I (New England Biolabs, Beverly, MA) and inserted into pNL9164 to make pNL9164-srtA.

    Article Title: Directional cDNA library construction assisted by the in vitro recombination reaction
    Article Snippet: DNA fragments containing the tetracycline-resistance gene (Tet) and beta-galactosidase gene (βGal) were amplified from control plasmids included in the field test kit provided by Life Technologies using the same set of PCR primers. .. MCS, Tet and βGal thus obtained were digested with Bsr GI (New England BioLabs), which cleaves at the TGTACA sequence within the common 15 bp core region of each att sequence, leaving four-base 5′ overhangs.

    Article Title: C7 deficiency in an Irish family: a deletion defect which is predominant in the Irish
    Article Snippet: Genetic markers in PCR products were examined by restriction digests. .. The restriction enzymes Dde I, Apo I, Rsa I and Bsr GI were obtained from New England Biolabs (St Albans, UK).

    Article Title: Haplotype-specific modulation of a SOX10/CREB response element at the Charcot–Marie–Tooth disease type 4C locus SH3TC2
    Article Snippet: Subsequent to PCR amplification and purification, each genomic segment was cloned into the pDONR221 vector using BP Clonase (Invitrogen). .. Successful cloning of each genomic segment upstream of the luciferase reporter gene was assessed by genotyping with Bsr GI (New England Biolabs).

    Article Title: Highly Significant Antiviral Activity of HIV-1 LTR-Specific Tre-Recombinase in Humanized Mice
    Article Snippet: Plasmid DNA was isolated from overnight cultures and digested with BsrG I and Xba I (NEB), resulting in different fragment sizes for recombined versus non-recombined substrate on agarose gels. .. DNA was isolated from the cells 48 h post transfection and analyzed for recombination by polymerase chain reaction using the primers F: 5′- GACAATAACCCTGATAAATGC-3′ , and R: 5′-CCTTAAACGCCTGGTGCTAC-3′ .

    Article Title: Pullulanase Is Necessary for the Efficient Intracellular Growth of Francisella tularensis
    Article Snippet: .. The PCR product treated with Xho I (New England Biolabs, Beverly, MA, USA) and Bsr GI (New England Biolabs) restriction enzymes was ligated into the pKEK1140 plasmid digested with the same enzymes. .. After ligation, competent cells of E . coli DH5α strains (Competent high DH5α, Toyobo, Tokyo, Japan) were transformed with the above plasmid.

    Recombinant:

    Article Title: Integration of RNAi and RNA-seq Reveals the Immune Responses of Epinephelus coioides to sigX Gene of Pseudomonas plecoglossicida
    Article Snippet: After linearizing pCM130/tac vectors with the restriction enzymes Nsi I and Bsr GI (New England Biolabs, USA), the oligonucleotides were annealed and ligated to the linearized pCM130/tac vectors using T4 DNA ligase (New England Biolabs) following the manufacturer’s recommendations. .. The recombinant pCM130/tac vectors were transformed into the competent E. coli DH5a cells by heat shock and then were extracted and electroporated into P. plecoglossicida as described previously ( ).

    Article Title: The G-rich Repeats in FMR1 and C9orf72 Loci Are Hotspots for Local Unpairing of DNA
    Article Snippet: .. DNA was fragmented using Hin dIII, Eco RI, Bsr GI, Xba I, and Ssp I, and pretreated, or not, with recombinant RNase H (#B0297S; NEB) overnight. .. Then, 4 µg of digested DNA was immunoprecipitated with 10 µg of S9.6 antibody (kindly provided by Clinton E. Leysath, National Institutes of Health, or commercially available from Kerafast) overnight.

    Article Title: Dual RNA-Seq Unveils Pseudomonas plecoglossicida htpG Gene Functions During Host-Pathogen Interactions With Epinephelus coioides
    Article Snippet: After linearizing pCM130/tac vectors with the restriction enzymes Nsi I and Bsr GI (New England Biolabs, U.S.A), the oligonucleotides were annealed and ligated to the linearized pCM130/tac vectors using T4 DNA ligase (New England Biolabs) based on the manufacturer's recommendations. .. The recombinant pCM130/tac vectors were transformed into competent E. coli DH5a cells by heat shock, and then extracted and electroporated into P. plecoglossicida .

    Gene Knockout:

    Article Title: Equilibrium of sortase A dimerization on Staphylococcus aureus cell surface mediates its cell wall sorting activity
    Article Snippet: TargeTron™ Gene Knockout System uses site-specific group II intron insertions to permanently disrupt genes on the bacterial genomic DNA. .. The amplified 350 bp fragment, which targets the intron to the SrtA sequence, was digested with Hind III and BsrG I (New England Biolabs, Beverly, MA) and inserted into pNL9164 to make pNL9164-srtA.

    Article Title: Pullulanase Is Necessary for the Efficient Intracellular Growth of Francisella tularensis
    Article Snippet: Construction of a ΔpulB mutant of F . tularensis subsp . tularensis SCHU P9 A pulB gene knockout mutant of SCHU P9 (ΔpulB ) was constructed using a mutagenesis system based on the group II intron of the ltrB gene of Lactobacillus lactis designed to function in F . tularensis and plasmid pKEK1140 (GenBank accession number: EU499313), a kind gift from Dr. Karl E. Klose (South Texas Center for Emerging Infectious Diseases and Department of Biology, University of Texas, San Antonio). .. The PCR product treated with Xho I (New England Biolabs, Beverly, MA, USA) and Bsr GI (New England Biolabs) restriction enzymes was ligated into the pKEK1140 plasmid digested with the same enzymes.

    Mutagenesis:

    Article Title: Haplotype-specific modulation of a SOX10/CREB response element at the Charcot–Marie–Tooth disease type 4C locus SH3TC2
    Article Snippet: Successful cloning of each genomic segment upstream of the luciferase reporter gene was assessed by genotyping with Bsr GI (New England Biolabs). .. Site-directed mutagenesis reactions were performed using the QuikChange II Mutagenesis Kit (Agilent Technologies) and appropriate mutation-bearing primers ( ).

    Article Title: Pullulanase Is Necessary for the Efficient Intracellular Growth of Francisella tularensis
    Article Snippet: Paragraph title: Construction of a ΔpulB mutant of F . tularensis subsp . tularensis SCHU P9 ... The PCR product treated with Xho I (New England Biolabs, Beverly, MA, USA) and Bsr GI (New England Biolabs) restriction enzymes was ligated into the pKEK1140 plasmid digested with the same enzymes.

    Article Title: Dimerization is required for GARS-mediated neurotoxicity in dominant CMT disease
    Article Snippet: Plasmids were isolated from individual clones and sequenced to confirm mutagenesis and exclude polymerase errors. .. Resulting clones were purified and digested with Bsr GI (New England Biolabs, Ipswich, MA, USA) to confirm recombination.

    Article Title: A novel AARS mutation in a family with dominant myeloneuropathy
    Article Snippet: Plasmids were isolated from individual clones and sequenced to confirm mutagenesis and exclude polymerase errors. .. Resulting clones were purified and digested with Bsr GI (New England Biolabs, Ipswich, MA) to confirm recombination.

    Isolation:

    Article Title: Highly Significant Antiviral Activity of HIV-1 LTR-Specific Tre-Recombinase in Humanized Mice
    Article Snippet: .. Plasmid DNA was isolated from overnight cultures and digested with BsrG I and Xba I (NEB), resulting in different fragment sizes for recombined versus non-recombined substrate on agarose gels. .. Recombination on the Tre target loxLTR served as positive control.

    Article Title: Dimerization is required for GARS-mediated neurotoxicity in dominant CMT disease
    Article Snippet: Plasmids were isolated from individual clones and sequenced to confirm mutagenesis and exclude polymerase errors. .. Resulting clones were purified and digested with Bsr GI (New England Biolabs, Ipswich, MA, USA) to confirm recombination.

    Article Title: A novel AARS mutation in a family with dominant myeloneuropathy
    Article Snippet: Plasmids were isolated from individual clones and sequenced to confirm mutagenesis and exclude polymerase errors. .. Resulting clones were purified and digested with Bsr GI (New England Biolabs, Ipswich, MA) to confirm recombination.

    Purification:

    Article Title: BRCA2 controls DNA:RNA hybrid level at DSBs by mediating RNase H2 recruitment
    Article Snippet: Briefly, DNA was extracted gently with phenol:chloroform:isoamyl alcohol (Sigma-Aldrich cat. no. P2069) and digested with Hind III, Eco RI, Bsr GI, Xba I, and Ssp I (NEB). .. After purification from restriction enzymes, half of the DNA was treated overnight with RNase H (NEB).

    Article Title: BRCA2 controls DNA:RNA hybrid level at DSBs by mediating RNase H2 recruitment
    Article Snippet: Briefly, DNA was extracted gently with phenol:chloroform:isoamyl alcohol (Sigma-Aldrich cat. no. P2069) and digested with Hind III, Eco RI, Bsr GI, Xba I, and Ssp I (NEB). .. After purification from restriction enzymes, half of the DNA was treated overnight with RNase H (NEB).

    Article Title: Directional cDNA library construction assisted by the in vitro recombination reaction
    Article Snippet: MCS, Tet and βGal thus obtained were digested with Bsr GI (New England BioLabs), which cleaves at the TGTACA sequence within the common 15 bp core region of each att sequence, leaving four-base 5′ overhangs. .. The products with or without Bsr GI digestion were finally purified using the Concert Rapid PCR Purification System (Life Technologies) and then used for LC or RC, respectively.

    Article Title: Haplotype-specific modulation of a SOX10/CREB response element at the Charcot–Marie–Tooth disease type 4C locus SH3TC2
    Article Snippet: Subsequent to PCR amplification and purification, each genomic segment was cloned into the pDONR221 vector using BP Clonase (Invitrogen). .. Successful cloning of each genomic segment upstream of the luciferase reporter gene was assessed by genotyping with Bsr GI (New England Biolabs).

    Article Title: Pullulanase Is Necessary for the Efficient Intracellular Growth of Francisella tularensis
    Article Snippet: The PCR product treated with Xho I (New England Biolabs, Beverly, MA, USA) and Bsr GI (New England Biolabs) restriction enzymes was ligated into the pKEK1140 plasmid digested with the same enzymes. .. Transformants were selected on LB agar plates containing 50 μg/ml kanamycin, and the plasmid DNAs were purified using NucleoBond PC 100 columns (Macherey-Nagel GmbH & Co., Doren, Germany).

    Article Title: Dimerization is required for GARS-mediated neurotoxicity in dominant CMT disease
    Article Snippet: .. Resulting clones were purified and digested with Bsr GI (New England Biolabs, Ipswich, MA, USA) to confirm recombination. .. The Δ grs1 haploid yeast strain [harboring a pRS316 maintenance vector to express wild-type GRS1 and URA3 ( ) was transformed with wild-type or mutant GRS1 in a LEU2 -bearing pRS315 vector and selected on medium lacking uracil and leucine (Teknova, Hollister, CA, USA).

    Article Title: A novel AARS mutation in a family with dominant myeloneuropathy
    Article Snippet: .. Resulting clones were purified and digested with Bsr GI (New England Biolabs, Ipswich, MA) to confirm recombination. .. The Δ ala1 haploid yeast strain (harboring a pRS316 maintenance vector to express wild-type ALA1 and URA3 ) was transformed with wild-type or mutant ALA1 in a LEU2 -bearing pRS315 vector and selected on medium lacking uracil and leucine (Teknova, Hollister, CA).

    Sequencing:

    Article Title: Equilibrium of sortase A dimerization on Staphylococcus aureus cell surface mediates its cell wall sorting activity
    Article Snippet: .. The amplified 350 bp fragment, which targets the intron to the SrtA sequence, was digested with Hind III and BsrG I (New England Biolabs, Beverly, MA) and inserted into pNL9164 to make pNL9164-srtA. .. The plasmid pNL9164-srtA was then transformed into S. aureus strain RN4220 (general gift from Dr. Alan Lambowitz).

    Article Title: Directional cDNA library construction assisted by the in vitro recombination reaction
    Article Snippet: .. MCS, Tet and βGal thus obtained were digested with Bsr GI (New England BioLabs), which cleaves at the TGTACA sequence within the common 15 bp core region of each att sequence, leaving four-base 5′ overhangs. .. The products with or without Bsr GI digestion were finally purified using the Concert Rapid PCR Purification System (Life Technologies) and then used for LC or RC, respectively.

    Article Title: Haplotype-specific modulation of a SOX10/CREB response element at the Charcot–Marie–Tooth disease type 4C locus SH3TC2
    Article Snippet: Resulting constructs were genotyped by digestion with Bsr GI (New England Biolabs) and subjected to DNA sequence analysis to ensure sequence specificity. .. Successful cloning of each genomic segment upstream of the luciferase reporter gene was assessed by genotyping with Bsr GI (New England Biolabs).

    Article Title: Haplotype-specific modulation of a SOX10/CREB response element at the Charcot–Marie–Tooth disease type 4C locus SH3TC2
    Article Snippet: .. Resulting constructs were genotyped by digestion with Bsr GI (New England Biolabs) and subjected to DNA sequence analysis to ensure sequence specificity. .. Each resulting pDONR221 construct was recombined with a destination vector harboring a minimal promoter directing a luciferase reporter gene (pE1B-luciferase) ( ) using LR Clonase (Invitrogen).

    Selection:

    Article Title: High-Throughput System for the Presentation of Secreted and Surface-Exposed Proteins from Gram-Positive Bacteria in Functional Metagenomics Studies
    Article Snippet: The resulting entry clones (25–50) were pooled and the plasmid inserts, now flanked by att L1 and att L2, were transferred to pDG148-GW by site-specific recombination between the att L1 and att L2 sites and the att R1 and att R2 sites, respectively, present on the latter plasmid, in vitro , using LR clonase II (Invitrogen), followed by transformation of E. coli TOP10 with selection for ampicilin. .. The presence of inserts in pDONR223 and pDG148-GW was verified by digestion with Bsr GI and Eco RI (New England Biolabs), respectively, and PCR amplifications using the pDONR223-specific primers 5′-CCCAGTCACGACGTTGTAAAACG and 5′-GTAACATCAGAGATTTTGAGACAC .

    Article Title: Dimerization is required for GARS-mediated neurotoxicity in dominant CMT disease
    Article Snippet: Resulting clones were purified and digested with Bsr GI (New England Biolabs, Ipswich, MA, USA) to confirm recombination. .. Each colony was grown to saturation in the selection medium for 48 h. Next, 10 μl of undiluted and diluted (1:10 and 1:100) samples from each culture were spotted on plates containing 0.1% 5-FOA complete medium or SD -leu -ura growth medium (Teknova) and incubated at 30°C for 72 h. Yeast cell growth was determined by visual inspection.

    Article Title: A novel AARS mutation in a family with dominant myeloneuropathy
    Article Snippet: Resulting clones were purified and digested with Bsr GI (New England Biolabs, Ipswich, MA) to confirm recombination. .. Each colony was grown to saturation in selection media for 48 hours.

    Quantitative RT-PCR:

    Article Title: Integration of RNAi and RNA-seq Reveals the Immune Responses of Epinephelus coioides to sigX Gene of Pseudomonas plecoglossicida
    Article Snippet: After linearizing pCM130/tac vectors with the restriction enzymes Nsi I and Bsr GI (New England Biolabs, USA), the oligonucleotides were annealed and ligated to the linearized pCM130/tac vectors using T4 DNA ligase (New England Biolabs) following the manufacturer’s recommendations. .. Finally, the expression level of sigX of each RNAi strain was detected by qRT-PCR.

    Article Title: Dual RNA-Seq Unveils Pseudomonas plecoglossicida htpG Gene Functions During Host-Pathogen Interactions With Epinephelus coioides
    Article Snippet: After linearizing pCM130/tac vectors with the restriction enzymes Nsi I and Bsr GI (New England Biolabs, U.S.A), the oligonucleotides were annealed and ligated to the linearized pCM130/tac vectors using T4 DNA ligase (New England Biolabs) based on the manufacturer's recommendations. .. Finally, the expression level of the target gene of each RNAi strain was evaluated by qRT-PCR.

    Plasmid Preparation:

    Article Title: High-Throughput System for the Presentation of Secreted and Surface-Exposed Proteins from Gram-Positive Bacteria in Functional Metagenomics Studies
    Article Snippet: The resulting entry clones (25–50) were pooled and the plasmid inserts, now flanked by att L1 and att L2, were transferred to pDG148-GW by site-specific recombination between the att L1 and att L2 sites and the att R1 and att R2 sites, respectively, present on the latter plasmid, in vitro , using LR clonase II (Invitrogen), followed by transformation of E. coli TOP10 with selection for ampicilin. .. The presence of inserts in pDONR223 and pDG148-GW was verified by digestion with Bsr GI and Eco RI (New England Biolabs), respectively, and PCR amplifications using the pDONR223-specific primers 5′-CCCAGTCACGACGTTGTAAAACG and 5′-GTAACATCAGAGATTTTGAGACAC .

    Article Title: Equilibrium of sortase A dimerization on Staphylococcus aureus cell surface mediates its cell wall sorting activity
    Article Snippet: The amplified 350 bp fragment, which targets the intron to the SrtA sequence, was digested with Hind III and BsrG I (New England Biolabs, Beverly, MA) and inserted into pNL9164 to make pNL9164-srtA. .. The plasmid pNL9164-srtA was then transformed into S. aureus strain RN4220 (general gift from Dr. Alan Lambowitz).

    Article Title: Directional cDNA library construction assisted by the in vitro recombination reaction
    Article Snippet: The resultant plasmid, attB-pBluescript, was used as a template for preparation of a DNA fragment containing multiple cloning sites (MCS). .. MCS, Tet and βGal thus obtained were digested with Bsr GI (New England BioLabs), which cleaves at the TGTACA sequence within the common 15 bp core region of each att sequence, leaving four-base 5′ overhangs.

    Article Title: Haplotype-specific modulation of a SOX10/CREB response element at the Charcot–Marie–Tooth disease type 4C locus SH3TC2
    Article Snippet: Paragraph title: Expression vector construction ... Successful cloning of each genomic segment upstream of the luciferase reporter gene was assessed by genotyping with Bsr GI (New England Biolabs).

    Article Title: Highly Significant Antiviral Activity of HIV-1 LTR-Specific Tre-Recombinase in Humanized Mice
    Article Snippet: .. Plasmid DNA was isolated from overnight cultures and digested with BsrG I and Xba I (NEB), resulting in different fragment sizes for recombined versus non-recombined substrate on agarose gels. .. Recombination on the Tre target loxLTR served as positive control.

    Article Title: Pullulanase Is Necessary for the Efficient Intracellular Growth of Francisella tularensis
    Article Snippet: .. The PCR product treated with Xho I (New England Biolabs, Beverly, MA, USA) and Bsr GI (New England Biolabs) restriction enzymes was ligated into the pKEK1140 plasmid digested with the same enzymes. .. After ligation, competent cells of E . coli DH5α strains (Competent high DH5α, Toyobo, Tokyo, Japan) were transformed with the above plasmid.

    Article Title: Dimerization is required for GARS-mediated neurotoxicity in dominant CMT disease
    Article Snippet: The T209K GRS1 /pDONR221 entry clone was recombined into a Gateway-compatible LEU2 -bearing pRS315 destination vector. .. Resulting clones were purified and digested with Bsr GI (New England Biolabs, Ipswich, MA, USA) to confirm recombination.

    Real-time Polymerase Chain Reaction:

    Article Title: The G-rich Repeats in FMR1 and C9orf72 Loci Are Hotspots for Local Unpairing of DNA
    Article Snippet: DNA was fragmented using Hin dIII, Eco RI, Bsr GI, Xba I, and Ssp I, and pretreated, or not, with recombinant RNase H (#B0297S; NEB) overnight. .. The pulled-down material (with and without RNase H treatment) and 1% input DNA were then subjected to quantitative PCR analysis.

    shRNA:

    Article Title: Integration of RNAi and RNA-seq Reveals the Immune Responses of Epinephelus coioides to sigX Gene of Pseudomonas plecoglossicida
    Article Snippet: Five short hairpin RNA sequences targeting the sigX gene were designed and synthesized by Shanghai Generay Biotech Co., Ltd. (Shanghai, China) (Table S1 in Supplementary Material). .. After linearizing pCM130/tac vectors with the restriction enzymes Nsi I and Bsr GI (New England Biolabs, USA), the oligonucleotides were annealed and ligated to the linearized pCM130/tac vectors using T4 DNA ligase (New England Biolabs) following the manufacturer’s recommendations.

    Article Title: Dual RNA-Seq Unveils Pseudomonas plecoglossicida htpG Gene Functions During Host-Pathogen Interactions With Epinephelus coioides
    Article Snippet: Five short hairpin RNA sequences targeting htpG, rplF , and flgD were designed and synthesized by Shanghai Generay Biotech Co., Ltd. (Shanghai, China) ( ). .. After linearizing pCM130/tac vectors with the restriction enzymes Nsi I and Bsr GI (New England Biolabs, U.S.A), the oligonucleotides were annealed and ligated to the linearized pCM130/tac vectors using T4 DNA ligase (New England Biolabs) based on the manufacturer's recommendations.

    Agarose Gel Electrophoresis:

    Article Title: Directional cDNA library construction assisted by the in vitro recombination reaction
    Article Snippet: MCS, Tet and βGal thus obtained were digested with Bsr GI (New England BioLabs), which cleaves at the TGTACA sequence within the common 15 bp core region of each att sequence, leaving four-base 5′ overhangs. .. The linearized vector portion of attR-pSP73 was purified on an agarose gel, recovered using CONCERT Gel Extraction System (Life Technologies) and quantified by absorption at 260 nm.

    In Vitro:

    Article Title: High-Throughput System for the Presentation of Secreted and Surface-Exposed Proteins from Gram-Positive Bacteria in Functional Metagenomics Studies
    Article Snippet: The resulting entry clones (25–50) were pooled and the plasmid inserts, now flanked by att L1 and att L2, were transferred to pDG148-GW by site-specific recombination between the att L1 and att L2 sites and the att R1 and att R2 sites, respectively, present on the latter plasmid, in vitro , using LR clonase II (Invitrogen), followed by transformation of E. coli TOP10 with selection for ampicilin. .. The presence of inserts in pDONR223 and pDG148-GW was verified by digestion with Bsr GI and Eco RI (New England Biolabs), respectively, and PCR amplifications using the pDONR223-specific primers 5′-CCCAGTCACGACGTTGTAAAACG and 5′-GTAACATCAGAGATTTTGAGACAC .

    Ethanol Precipitation:

    Article Title: The G-rich Repeats in FMR1 and C9orf72 Loci Are Hotspots for Local Unpairing of DNA
    Article Snippet: Briefly, total nucleic acids were extracted by SDS/Proteinase K treatment at 37°, followed by phenol-chloroform extraction and ethanol precipitation. .. DNA was fragmented using Hin dIII, Eco RI, Bsr GI, Xba I, and Ssp I, and pretreated, or not, with recombinant RNase H (#B0297S; NEB) overnight.

    Knock-Out:

    Article Title: Equilibrium of sortase A dimerization on Staphylococcus aureus cell surface mediates its cell wall sorting activity
    Article Snippet: Paragraph title: Construction of SrtA knockout S. aureus strain ... The amplified 350 bp fragment, which targets the intron to the SrtA sequence, was digested with Hind III and BsrG I (New England Biolabs, Beverly, MA) and inserted into pNL9164 to make pNL9164-srtA.

    Immunoprecipitation:

    Article Title: BRCA2 controls DNA:RNA hybrid level at DSBs by mediating RNase H2 recruitment
    Article Snippet: Paragraph title: DNA:RNA hybrids immunoprecipitation ... Briefly, DNA was extracted gently with phenol:chloroform:isoamyl alcohol (Sigma-Aldrich cat. no. P2069) and digested with Hind III, Eco RI, Bsr GI, Xba I, and Ssp I (NEB).

    Article Title: The G-rich Repeats in FMR1 and C9orf72 Loci Are Hotspots for Local Unpairing of DNA
    Article Snippet: Paragraph title: DNA:RNA immunoprecipitation assay ... DNA was fragmented using Hin dIII, Eco RI, Bsr GI, Xba I, and Ssp I, and pretreated, or not, with recombinant RNase H (#B0297S; NEB) overnight.

    Gel Extraction:

    Article Title: Directional cDNA library construction assisted by the in vitro recombination reaction
    Article Snippet: MCS, Tet and βGal thus obtained were digested with Bsr GI (New England BioLabs), which cleaves at the TGTACA sequence within the common 15 bp core region of each att sequence, leaving four-base 5′ overhangs. .. The linearized vector portion of attR-pSP73 was purified on an agarose gel, recovered using CONCERT Gel Extraction System (Life Technologies) and quantified by absorption at 260 nm.

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    New England Biolabs bsr gi
    Bsr Gi, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 90/100, based on 14 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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