smai  (New England Biolabs)


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  • 99
    Name:
    SmaI
    Description:
    SmaI 10 000 units
    Catalog Number:
    R0141L
    Price:
    256
    Size:
    10 000 units
    Category:
    Restriction Enzymes
    Score:
    85
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    Structured Review

    New England Biolabs smai
    SmaI
    SmaI 10 000 units
    https://www.bioz.com/result/smai/product/New England Biolabs
    Average 99 stars, based on 73 article reviews
    Price from $9.99 to $1999.99
    smai - by Bioz Stars, 2019-12
    99/100 stars

    Images

    1) Product Images from "Clonal Population Structure and Specific Genotypes of Multidrug-Resistant Campylobacter coli from Turkeys"

    Article Title: Clonal Population Structure and Specific Genotypes of Multidrug-Resistant Campylobacter coli from Turkeys

    Journal:

    doi: 10.1128/AEM.02346-06

    PFGE-based clustering of C. coli strains investigated in this study. All 68 strains were analyzed by PFGE with SmaI and KpnI, and cluster analysis of the patterns was performed by BioNumerics as described in Materials and Methods. Two strains (1004 A
    Figure Legend Snippet: PFGE-based clustering of C. coli strains investigated in this study. All 68 strains were analyzed by PFGE with SmaI and KpnI, and cluster analysis of the patterns was performed by BioNumerics as described in Materials and Methods. Two strains (1004 A

    Techniques Used:

    PFGE profiles of group I strains. Nineteen MDR strains were characterized by PFGE using SmaI and KpnI, and cluster analysis of the patterns was performed by BioNumerics as described in Materials and Methods. The line labeled “a” indicates
    Figure Legend Snippet: PFGE profiles of group I strains. Nineteen MDR strains were characterized by PFGE using SmaI and KpnI, and cluster analysis of the patterns was performed by BioNumerics as described in Materials and Methods. The line labeled “a” indicates

    Techniques Used: Labeling

    A conserved SmaI PFGE pattern (type a ) was disseminated among MDR strains of different STs.
    Figure Legend Snippet: A conserved SmaI PFGE pattern (type a ) was disseminated among MDR strains of different STs.

    Techniques Used:

    PFGE profiles of group II strains. Forty-two strains were characterized by PFGE using SmaI and KpnI, and cluster analysis of the patterns was performed by BioNumerics as described in Materials and Methods. Strains which were not resistant to all six antibiotics
    Figure Legend Snippet: PFGE profiles of group II strains. Forty-two strains were characterized by PFGE using SmaI and KpnI, and cluster analysis of the patterns was performed by BioNumerics as described in Materials and Methods. Strains which were not resistant to all six antibiotics

    Techniques Used:

    PFGE profiles of 13 strains with various STs but a conserved PFGE profile (type a ). Cluster analysis of the SmaI and KpnI patterns was performed by BioNumerics as described in Materials and Methods.
    Figure Legend Snippet: PFGE profiles of 13 strains with various STs but a conserved PFGE profile (type a ). Cluster analysis of the SmaI and KpnI patterns was performed by BioNumerics as described in Materials and Methods.

    Techniques Used:

    2) Product Images from "Staphylococcus spp., Streptococcus canis, and Arcanobacterium phocae of healthy Canadian farmed mink and mink with pododermatitis"

    Article Title: Staphylococcus spp., Streptococcus canis, and Arcanobacterium phocae of healthy Canadian farmed mink and mink with pododermatitis

    Journal:

    doi:

    Pulsed-field gel electrophoresis (PFGE)-based dendrogram of 13 SmaI digested S. canis genomic DNA from healthy and diseased mink from 2 unrelated farms in Canada and clinical infections in 4 unrelated dogs. Scale bar represents percent similarity based on Dice coefficients and UPGMA clustering.
    Figure Legend Snippet: Pulsed-field gel electrophoresis (PFGE)-based dendrogram of 13 SmaI digested S. canis genomic DNA from healthy and diseased mink from 2 unrelated farms in Canada and clinical infections in 4 unrelated dogs. Scale bar represents percent similarity based on Dice coefficients and UPGMA clustering.

    Techniques Used: Pulsed-Field Gel, Electrophoresis

    Pulsed-field gel electrophoresis (PFGE)-based dendrogram of 21 SmaI digested S. delphini genomic DNA from healthy and diseased mink from 2 unrelated farms in Canada. Scale bar represents percent similarity based on Dice coefficients and UPGMA clustering. Asterisk (*) indicates 2 isolates taken from the same individual mink.
    Figure Legend Snippet: Pulsed-field gel electrophoresis (PFGE)-based dendrogram of 21 SmaI digested S. delphini genomic DNA from healthy and diseased mink from 2 unrelated farms in Canada. Scale bar represents percent similarity based on Dice coefficients and UPGMA clustering. Asterisk (*) indicates 2 isolates taken from the same individual mink.

    Techniques Used: Pulsed-Field Gel, Electrophoresis

    3) Product Images from "Identification of fusB-Mediated Fusidic Acid Resistance Islands in Staphylococcus epidermidis Isolates"

    Article Title: Identification of fusB-Mediated Fusidic Acid Resistance Islands in Staphylococcus epidermidis Isolates

    Journal:

    doi: 10.1128/AAC.00592-11

    Dendrogram produced by BioNumerics software, showing distances calculated by Dice similarity index of SmaI-digested DNA fragments among 34 fusB- carrying S. epidermidis isolates. The degree of similarity is shown in the scale. Different degrees of gray
    Figure Legend Snippet: Dendrogram produced by BioNumerics software, showing distances calculated by Dice similarity index of SmaI-digested DNA fragments among 34 fusB- carrying S. epidermidis isolates. The degree of similarity is shown in the scale. Different degrees of gray

    Techniques Used: Produced, Software

    4) Product Images from "High-throughput methylation profiling by MCA coupled to CpG island microarray"

    Article Title: High-throughput methylation profiling by MCA coupled to CpG island microarray

    Journal:

    doi: 10.1101/gr.6417007

    Schematic diagram of the MCAM method. Enrichment for methylated DNA and reduction of genome complexity is achieved by serial digestion with SmaI (methylation sensitive) and XmaI (methylation insensitive) restriction enzymes, followed by ligation of adaptors
    Figure Legend Snippet: Schematic diagram of the MCAM method. Enrichment for methylated DNA and reduction of genome complexity is achieved by serial digestion with SmaI (methylation sensitive) and XmaI (methylation insensitive) restriction enzymes, followed by ligation of adaptors

    Techniques Used: Methylation, Ligation

    5) Product Images from "Oxacilin-resistant Coagulase-negative staphylococci (CoNS) bacteremia in a general hospital at S?o Paulo city, Brasil"

    Article Title: Oxacilin-resistant Coagulase-negative staphylococci (CoNS) bacteremia in a general hospital at S?o Paulo city, Brasil

    Journal: Brazilian Journal of Microbiology

    doi: 10.1590/S1517-83822008000400006

    PFGE profile of SmaI-digested chromosomal DNA of CoNS isolates, obtained from patients in 9 de Julho Hospital in São Paulo city, Brazil. λ lamba ladder DNA markers; lanes 1–5 S. epidermidis ; lanes 6,7 and 10: S. haemolyticus ; lane 8: S. hominis ; lane 9: S. warneri ; lane 11: S. cohnii spp urealyticus .
    Figure Legend Snippet: PFGE profile of SmaI-digested chromosomal DNA of CoNS isolates, obtained from patients in 9 de Julho Hospital in São Paulo city, Brazil. λ lamba ladder DNA markers; lanes 1–5 S. epidermidis ; lanes 6,7 and 10: S. haemolyticus ; lane 8: S. hominis ; lane 9: S. warneri ; lane 11: S. cohnii spp urealyticus .

    Techniques Used:

    6) Product Images from "A DNA insulator prevents repression of a targeted X-linked transgene but not its random or imprinted X inactivation"

    Article Title: A DNA insulator prevents repression of a targeted X-linked transgene but not its random or imprinted X inactivation

    Journal:

    doi: 10.1073/pnas.0603754103

    DNA methylation status of the GFPn transgenes on the Xa and Xi. ( A ) The structure of the transgenes integrated into the Hprt locus. The location of EcoRI (E) and SmaI (S) restriction sites in the transgenes and the sizes of the fragments detected after
    Figure Legend Snippet: DNA methylation status of the GFPn transgenes on the Xa and Xi. ( A ) The structure of the transgenes integrated into the Hprt locus. The location of EcoRI (E) and SmaI (S) restriction sites in the transgenes and the sizes of the fragments detected after

    Techniques Used: DNA Methylation Assay

    7) Product Images from "Aspartoacylase-LacZ Knockin Mice: An Engineered Model of Canavan Disease"

    Article Title: Aspartoacylase-LacZ Knockin Mice: An Engineered Model of Canavan Disease

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0020336

    Generation of aspa lacZ mice by homologous recombination. A, Genomic structure of the murine aspa locus, which spans 6 exons (black boxes). Homologous recombination of the targeting vector inserts the βgeo cassette (blue box), encoding β-Galactosidase (lacZ), into intron 1 of the intact aspa gene. This cassette is flanked by frt-sites (green circles). Additional loxP sites flank exon 2 (red triangles). The vector includes a DTA cassette for negative selection. In the targeted allele, exon 1 is spliced to the splice acceptor site preceding βgeo, and transcription is terminated at the introduced pA site. The conditional aspa flox allele is produced by breeding aspa lacZ mutants to FLP-deleter mice [40] for recombination of the βgeo cassette in vivo . Note that frt and loxP sites are not drawn to scale. βgeo, β-galactosidase-neomycin resistance cassette; SA, splice acceptor; pA, polyadenylation site, DTA, Diphteria toxin gene. B, For Southern blot analysis genomic DNA was digested with SmaI/ApaI. The neo probe (white box in A) detected the expected 7.5 kb band in heterozygotes and homozygotes. C, Genomic PCR of littermates with primers 1 3 produces the expected amplicons in aspa +/+ (336 bp), aspa lacZ/+ (387 bp and 336 bp) and aspa lacZ/lacZ (387 bp) animals. The upstream loxP site was detected by PCR with primers 1 2 yielding a 307 bp band. D, Q-RT-PCR using a TaqMan probe for quantification of aspa mRNA levels in brains of aspa +/+ , aspa lacZ/+ , and aspa lacZ/lacZ mice (n = 3) confirms the attenuation of transcription downstream of exon 2 in the targeted allele. E, Representative Western blot of whole brain lysates of a spa +/+ , aspa lacZ/+ and aspa lacZ/lacZ mice (aged 4 months, n = 3). The 37 kD protein ASPA was detected in aspa +/+ and aspa lacZ/+ mice but not in aspa lacZ/lacZ mutants. β-Galactosidase is expressed in aspa lacZ/+ and aspa lacZ/lacZ brain but not controls. α-Tubulin was used as loading control. F. Representative picture of a male aspa lacZ/lacZ mutant (asterisk) and an aspa +/+ littermate at P70.
    Figure Legend Snippet: Generation of aspa lacZ mice by homologous recombination. A, Genomic structure of the murine aspa locus, which spans 6 exons (black boxes). Homologous recombination of the targeting vector inserts the βgeo cassette (blue box), encoding β-Galactosidase (lacZ), into intron 1 of the intact aspa gene. This cassette is flanked by frt-sites (green circles). Additional loxP sites flank exon 2 (red triangles). The vector includes a DTA cassette for negative selection. In the targeted allele, exon 1 is spliced to the splice acceptor site preceding βgeo, and transcription is terminated at the introduced pA site. The conditional aspa flox allele is produced by breeding aspa lacZ mutants to FLP-deleter mice [40] for recombination of the βgeo cassette in vivo . Note that frt and loxP sites are not drawn to scale. βgeo, β-galactosidase-neomycin resistance cassette; SA, splice acceptor; pA, polyadenylation site, DTA, Diphteria toxin gene. B, For Southern blot analysis genomic DNA was digested with SmaI/ApaI. The neo probe (white box in A) detected the expected 7.5 kb band in heterozygotes and homozygotes. C, Genomic PCR of littermates with primers 1 3 produces the expected amplicons in aspa +/+ (336 bp), aspa lacZ/+ (387 bp and 336 bp) and aspa lacZ/lacZ (387 bp) animals. The upstream loxP site was detected by PCR with primers 1 2 yielding a 307 bp band. D, Q-RT-PCR using a TaqMan probe for quantification of aspa mRNA levels in brains of aspa +/+ , aspa lacZ/+ , and aspa lacZ/lacZ mice (n = 3) confirms the attenuation of transcription downstream of exon 2 in the targeted allele. E, Representative Western blot of whole brain lysates of a spa +/+ , aspa lacZ/+ and aspa lacZ/lacZ mice (aged 4 months, n = 3). The 37 kD protein ASPA was detected in aspa +/+ and aspa lacZ/+ mice but not in aspa lacZ/lacZ mutants. β-Galactosidase is expressed in aspa lacZ/+ and aspa lacZ/lacZ brain but not controls. α-Tubulin was used as loading control. F. Representative picture of a male aspa lacZ/lacZ mutant (asterisk) and an aspa +/+ littermate at P70.

    Techniques Used: Mouse Assay, Homologous Recombination, Plasmid Preparation, Selection, Produced, In Vivo, Southern Blot, Polymerase Chain Reaction, Reverse Transcription Polymerase Chain Reaction, Western Blot, Mutagenesis

    8) Product Images from "Diversity of Proteolytic Clostridium botulinum Strains, Determined by a Pulsed-Field Gel Electrophoresis Approach"

    Article Title: Diversity of Proteolytic Clostridium botulinum Strains, Determined by a Pulsed-Field Gel Electrophoresis Approach

    Journal:

    doi: 10.1128/AEM.71.3.1311-1317.2005

    Digestion patterns of five proteolytic C. botulinum strains, two of type A (ATCC 3502 and ATCC 19397), two of type B (FT 243 and 4B), and one of type F (ATCC 35415), using the rare-cutting restriction enzymes SacII and SmaI. The pulse time ramp was 1
    Figure Legend Snippet: Digestion patterns of five proteolytic C. botulinum strains, two of type A (ATCC 3502 and ATCC 19397), two of type B (FT 243 and 4B), and one of type F (ATCC 35415), using the rare-cutting restriction enzymes SacII and SmaI. The pulse time ramp was 1

    Techniques Used:

    9) Product Images from "Detection of a New cfr-Like Gene, cfr(B), in Enterococcus faecium Isolates Recovered from Human Specimens in the United States as Part of the SENTRY Antimicrobial Surveillance Program"

    Article Title: Detection of a New cfr-Like Gene, cfr(B), in Enterococcus faecium Isolates Recovered from Human Specimens in the United States as Part of the SENTRY Antimicrobial Surveillance Program

    Journal:

    doi: 10.1128/AAC.01473-15

    PFGE of E. faecium genomic DNA digests and Southern blot hybridization with a cfr -specific digoxigenin-labeled probe. (A) SmaI digests; (B) EcoRI digests; (C) ICeuI digests. Lanes λ, 1, and 2 represent lambda DNA molecular marker, E. faecium 18203R, and E. faecium 18961R, respectively.
    Figure Legend Snippet: PFGE of E. faecium genomic DNA digests and Southern blot hybridization with a cfr -specific digoxigenin-labeled probe. (A) SmaI digests; (B) EcoRI digests; (C) ICeuI digests. Lanes λ, 1, and 2 represent lambda DNA molecular marker, E. faecium 18203R, and E. faecium 18961R, respectively.

    Techniques Used: Southern Blot, Hybridization, Labeling, Lambda DNA Preparation, Marker

    10) Product Images from "Molecular Evolutionary Pathways toward Two Successful Community-Associated but Multidrug-Resistant ST59 Methicillin-Resistant Staphylococcus aureus Lineages in Taiwan: Dynamic Modes of Mobile Genetic Element Salvages"

    Article Title: Molecular Evolutionary Pathways toward Two Successful Community-Associated but Multidrug-Resistant ST59 Methicillin-Resistant Staphylococcus aureus Lineages in Taiwan: Dynamic Modes of Mobile Genetic Element Salvages

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0162526

    Selection for loss of antibiotic resistance in ST59/SCC mec V (5C2 5) MRSA strain PM1. (A) Cartoon representation of MES structures in PM1 and the generated colonies PM1-1 to PM1-5. (B) SmaI-digested PFGE of PM1 and PM1-1 to PM1-5 and the Southern blot. The SmaI-digested DNA separated by PFGE is shown in the left. The DNA was transferred to a nylon membrane and detected by Southern blot hybridization with DIG-labeled cat -, ermB - and IS 1216V -specific probes, which are shown on the right. Abbreviations: E r , erythromycin resistant; K r , kanamycin resistant; S r , streptomycin resistant; C r , chloramphenicol resistant.
    Figure Legend Snippet: Selection for loss of antibiotic resistance in ST59/SCC mec V (5C2 5) MRSA strain PM1. (A) Cartoon representation of MES structures in PM1 and the generated colonies PM1-1 to PM1-5. (B) SmaI-digested PFGE of PM1 and PM1-1 to PM1-5 and the Southern blot. The SmaI-digested DNA separated by PFGE is shown in the left. The DNA was transferred to a nylon membrane and detected by Southern blot hybridization with DIG-labeled cat -, ermB - and IS 1216V -specific probes, which are shown on the right. Abbreviations: E r , erythromycin resistant; K r , kanamycin resistant; S r , streptomycin resistant; C r , chloramphenicol resistant.

    Techniques Used: Selection, Generated, Southern Blot, Hybridization, Labeling

    11) Product Images from "Characteristics of Community-Associated Methicillin-Resistant Staphylococcus aureus (CA-MRSA) Strains Isolated from Skin and Soft-Tissue Infections in Uruguay"

    Article Title: Characteristics of Community-Associated Methicillin-Resistant Staphylococcus aureus (CA-MRSA) Strains Isolated from Skin and Soft-Tissue Infections in Uruguay

    Journal: International Journal of Microbiology

    doi: 10.1155/2009/472126

    (a) Pulsed-field gel electrophoresis (PFGE) patterns of the SmaI-digested genomic DNA obtained from CA-MRSA isolates in Uruguay. Lane 1, pulsotype A (strain IH 23); lane 2, pulsotype B (strain IH 48); lane 3, pulsotype C (strain IH 46); lane 4, pulsotype D (strain IH 7); lane 5, pulsotype E (strain IH 22) and lane 6 pulsotype F (strain IH 69). (b) Pulsed-field gel electrophoresis (PFGE) patterns of the SmaI-digested genomic DNA obtained from CA-MRSA isolates in Uruguay. Lane 1, pulsotype A1 (strain IH 36); lane 2, pulsotype A (strain IH 44); lane 3, pulsotype A2 (strain IH 9).
    Figure Legend Snippet: (a) Pulsed-field gel electrophoresis (PFGE) patterns of the SmaI-digested genomic DNA obtained from CA-MRSA isolates in Uruguay. Lane 1, pulsotype A (strain IH 23); lane 2, pulsotype B (strain IH 48); lane 3, pulsotype C (strain IH 46); lane 4, pulsotype D (strain IH 7); lane 5, pulsotype E (strain IH 22) and lane 6 pulsotype F (strain IH 69). (b) Pulsed-field gel electrophoresis (PFGE) patterns of the SmaI-digested genomic DNA obtained from CA-MRSA isolates in Uruguay. Lane 1, pulsotype A1 (strain IH 36); lane 2, pulsotype A (strain IH 44); lane 3, pulsotype A2 (strain IH 9).

    Techniques Used: Pulsed-Field Gel, Electrophoresis

    12) Product Images from "The nsp3 Macrodomain Promotes Virulence in Mice with Coronavirus-Induced Encephalitis"

    Article Title: The nsp3 Macrodomain Promotes Virulence in Mice with Coronavirus-Induced Encephalitis

    Journal:

    doi: 10.1128/JVI.02596-14

    Engineering a full-length JHMV infectious BAC. (A) Strategy used to construct the JHMV BAC. First, a synthetic DNA was designed containing a CMV promoter, 1,991 bp of 5′ JHM sequence, and 1,197 bp of 3′ JHM sequence separated by a NotI site, and finally a poly(A) tail (pA), ribozyme cleavage site, HDV ribozyme, and BGH termination signal at the end. This DNA was then digested with SmaI and HindIII and ligated into the parental BAC plasmid, and the resulting clone was termed pBAC-JHMV 5′3′. Subsequently, pBAC-JHMV 5′3′ and a full-length JHMV ligated in vitro were digested with PacI and SanDI, ligated, and transformed into competent DH10B E. coli cells. The resulting BAC was termed pBAC-JHMV. (B) Confirmation of BAC clones. Representative clones of pBAC-JHMVIA and pBAC-JHMVSD were digested with EcoRI and EcoRV. The expected banding pattern for EcoRI digestion is 14.5, 11, 7.7, and 6.3 kb, while the expected pattern from EcoRV digestion is 22.3, 12.6, 4.7, and 0.2 kb. The resulting digests matched the expected digestion patterns (0.2-kb fragment is undetectable on the gel).
    Figure Legend Snippet: Engineering a full-length JHMV infectious BAC. (A) Strategy used to construct the JHMV BAC. First, a synthetic DNA was designed containing a CMV promoter, 1,991 bp of 5′ JHM sequence, and 1,197 bp of 3′ JHM sequence separated by a NotI site, and finally a poly(A) tail (pA), ribozyme cleavage site, HDV ribozyme, and BGH termination signal at the end. This DNA was then digested with SmaI and HindIII and ligated into the parental BAC plasmid, and the resulting clone was termed pBAC-JHMV 5′3′. Subsequently, pBAC-JHMV 5′3′ and a full-length JHMV ligated in vitro were digested with PacI and SanDI, ligated, and transformed into competent DH10B E. coli cells. The resulting BAC was termed pBAC-JHMV. (B) Confirmation of BAC clones. Representative clones of pBAC-JHMVIA and pBAC-JHMVSD were digested with EcoRI and EcoRV. The expected banding pattern for EcoRI digestion is 14.5, 11, 7.7, and 6.3 kb, while the expected pattern from EcoRV digestion is 22.3, 12.6, 4.7, and 0.2 kb. The resulting digests matched the expected digestion patterns (0.2-kb fragment is undetectable on the gel).

    Techniques Used: BAC Assay, Construct, Sequencing, Plasmid Preparation, In Vitro, Transformation Assay, Clone Assay

    13) Product Images from "The preparation of HL-60 cells vaccine expressing BCG heat shock protein 70 and detection of its immunogenicity in vitro"

    Article Title: The preparation of HL-60 cells vaccine expressing BCG heat shock protein 70 and detection of its immunogenicity in vitro

    Journal:

    doi: 10.4161/hv.21321

    Figure 2. Identification of recombinant plasmid pDisplay-HSP70. Lane 1: DNA marker (DL2000); Lane 2: PCR product of HSP70; Lane 3: plasmid pDisplay digested with BglII and SmaI; Lane 4: HSP70 digested with BglII and SmaI; Lane 5: recombinant plasmid
    Figure Legend Snippet: Figure 2. Identification of recombinant plasmid pDisplay-HSP70. Lane 1: DNA marker (DL2000); Lane 2: PCR product of HSP70; Lane 3: plasmid pDisplay digested with BglII and SmaI; Lane 4: HSP70 digested with BglII and SmaI; Lane 5: recombinant plasmid

    Techniques Used: Recombinant, Plasmid Preparation, Marker, Polymerase Chain Reaction

    14) Product Images from "RIP140 in thyroid hormone-repression and chromatin remodeling of Crabp1 gene during adipocyte differentiation"

    Article Title: RIP140 in thyroid hormone-repression and chromatin remodeling of Crabp1 gene during adipocyte differentiation

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp780

    Alterations of restriction enzyme accessibility in the juxtaposed region of Crabp1 promoter along the course of adipocyte differentiation. ( A ) Detection of nucleosome array. Chromatin of MNase partially digested nuclei was separated on agarose gels followed by Southern blot analysis (left panel) and EtBr staining (right panel). A probe used covering the TIS (HinfI-PvuII) was used for Southern blot analysis. ( B–F ) Nuclei (differentiation Days 0, 4, 8) were digested with the indicated restriction enzymes and the recovered chromatin DNAs were completely re-digested with ApaI ( B–D ) or PstI ( E and F ) (the relationship of specific restriction site and each individual nucleosome was depicted in panel G), followed by Southern blot analyses using probe 1 ( D ) or probe 2 (B–C and E–F). ( G ) Schematic description of restriction enzyme digestion, nucleosomes, and predicted fragments detected with probes on the Southern blots. Complete digestion with ApaI produced 1.3 kb, and complete digestion with PstI produced 1.65 kb fragments. The generated fragments by digestion with XhoI, PstI, SmaI, SpeI and ApaLI for the corresponding nucleosome are 0.93, 0.67, 0.86, 0.55 and 0.65 kb, respectively. Nucleosomes on this array were named N5 to N-1, from the 5′- to the 3′-ends of this chromatin segment. Experiments were performed for least three times.
    Figure Legend Snippet: Alterations of restriction enzyme accessibility in the juxtaposed region of Crabp1 promoter along the course of adipocyte differentiation. ( A ) Detection of nucleosome array. Chromatin of MNase partially digested nuclei was separated on agarose gels followed by Southern blot analysis (left panel) and EtBr staining (right panel). A probe used covering the TIS (HinfI-PvuII) was used for Southern blot analysis. ( B–F ) Nuclei (differentiation Days 0, 4, 8) were digested with the indicated restriction enzymes and the recovered chromatin DNAs were completely re-digested with ApaI ( B–D ) or PstI ( E and F ) (the relationship of specific restriction site and each individual nucleosome was depicted in panel G), followed by Southern blot analyses using probe 1 ( D ) or probe 2 (B–C and E–F). ( G ) Schematic description of restriction enzyme digestion, nucleosomes, and predicted fragments detected with probes on the Southern blots. Complete digestion with ApaI produced 1.3 kb, and complete digestion with PstI produced 1.65 kb fragments. The generated fragments by digestion with XhoI, PstI, SmaI, SpeI and ApaLI for the corresponding nucleosome are 0.93, 0.67, 0.86, 0.55 and 0.65 kb, respectively. Nucleosomes on this array were named N5 to N-1, from the 5′- to the 3′-ends of this chromatin segment. Experiments were performed for least three times.

    Techniques Used: Southern Blot, Staining, Produced, Generated

    15) Product Images from "Characterization of Vancomycin-Resistant Enterococcus faecium Isolated from Swine in Three Michigan Counties"

    Article Title: Characterization of Vancomycin-Resistant Enterococcus faecium Isolated from Swine in Three Michigan Counties

    Journal:

    doi: 10.1128/JCM.02346-09

    PFGE of SmaI-digested genomic DNA from six vanA -containing Enterococcus faecium isolates from swine in three Michigan counties.
    Figure Legend Snippet: PFGE of SmaI-digested genomic DNA from six vanA -containing Enterococcus faecium isolates from swine in three Michigan counties.

    Techniques Used:

    Related Articles

    Transduction:

    Article Title: Chromogranin B (CHGB): Intra- and extra-cellular mechanisms to regulate catecholamine storage and release, in catecholaminergic cells and organisms
    Article Snippet: Paragraph title: Over-expression of CHGB in PC12 cells by transduction of a CHGB lentivirus ... The 2034 bp PCR product was digested with SmaI and SalI (New England Biolab), purified (Cat#: 28074, Qiagen), and ligated into corresponding site of SIN18-hPGK-tdTomato-T2A vector to generate SIN18-hPGK-tdTomato-T2A-hCHGB lentiviral vector plasmid ( ).

    Clone Assay:

    Article Title: Immune Mediators Regulate CFTR Expression through a Bifunctional Airway-Selective Enhancer
    Article Snippet: The minimal 350-bp DHS-35kb [DHS-35kb(350)] enhancer element (hg 19, chr7:117085371 to 117085720) was amplified with Pfu DNA polymerase (for primers, see Table S1) and cloned into the pSC-B vector using a StrataClone Blunt PCR cloning kit (Agilent Technologies, Santa Clara, CA). .. Cleavage of this construct with XhoI and SmaI (New England BioLabs, Ipswich, MA) enabled Klenow DNA polymerase (New England BioLabs) fill-in with [32 P]dCTP.

    Article Title: Role of the VP16-Binding Domain of vhs in Viral Growth, Host Shutoff Activity, and Pathogenesis
    Article Snippet: PCR amplification conditions were as follows: 5 min, 95°C for 30 s, 95°C for 30 s, 58°C for 30 s, 72°C for 7 min, 72°C for 35 cycles. .. The resulting fragment was treated with T4 polynucleotide kinase (New England Biolabs, Beverly, Mass.) and cloned into SmaI (New England Biolabs, Beverly, Mass.)-digested, calf intestinal phosphatase (New England Biolabs)-treated pMECA , resulting in the construct pMECA-vhs. .. The 1.5-kb EcoRI fragment containing the vhs open reading frame from pMECA-vhs was ligated into EcoRI-cut pCI in both orientations to generate pCI-vhsF and pCI-vhsR.

    Article Title: A Klebsiella pneumoniae antibiotic resistance mechanism that subdues host defences and promotes virulence
    Article Snippet: For complementation of the 52145‐ΔmgrB mutant strain, a PCR fragment (primers: mgrB _UPFWD and mgrB _DWNRVS) comprising the coding and promoter regions of the K. pneumoniae 52145 mgrB gene was amplified using Phusion® High‐Fidelity DNA Polymerase (New England Biolabs). .. The 1,507‐bp amplicon was gel‐purified and then cloned into SmaI‐digested (New England Biolabs), Antarctic Phosphatase (New England Biolabs)‐treated pUC18R6KT‐mini‐Tn7TKm plasmid (Choi et al , ) to obtain pUC18R6KT‐mini‐Tn7TKm_Kp52145mgrB Com. pUC18R6KT‐mini‐Tn7TKm_Kp52145mgrB Com was then transformed into E. coli SY327 and thereafter into E. coli β2163. .. In addition, the transposase‐containing pTSNSK‐Tp plasmid (Crepin et al , ) was introduced to the 52145‐ΔmgrB strain by electroporation to give 52145‐ΔmgrB /pTSNSK‐Tp.

    Article Title: Primed CRISPR adaptation in Escherichia coli cells does not depend on conformational changes in the Cascade effector complex detected in Vitro
    Article Snippet: These data are consistent with the interference-based model of primed adaptation that is independent of effector complex conformations at the priming site. .. Constructs containing the WT g8 protospacer and its variants were cloned into plasmid pUC19 (NEB) at the single SmaI (NEB) site by blunt end ligation. .. The 73 bp insert DNA carried the target sequence (PAM and protospacer variant) in its center (see ).

    Article Title: LSm1-7 complexes bind to specific sites in viral RNA genomes and regulate their translation and replication
    Article Snippet: Forward primers carried a T7 promoter sequence at their 5′ end. .. Amplified fragments were cloned into SmaI digested pUC18 (New England Biolabs) or pGEM-T vectors (Promega). .. Fragments IR A and IR 3ΔA were synthetically produced by Geneart and cloned into plasmid pPCR-Script-ampR (Stratagene) using KpnI and SacI restriction sites.

    Amplification:

    Article Title: A Robust System for Production of Superabundant VP1 Recombinant AAV Vectors
    Article Snippet: The plasmid pRB21 (with P7.5 promoter) was digested with SmaI (NEB) and used as the backbone to construct all plasmids having individual AAV and Ad genes. .. Rep68 , Rep40 , X-gene , and AAP were generated using PCR amplified from pH22.

    Article Title: Molecular quantification and differentiation of Candida species in biological specimens of patients with liver cirrhosis
    Article Snippet: The amplification steps were as follows: initial denaturation at 95°C for 1 min, 40 cycles of 15 sec at 95°C, 15 sec at 55°C and 10 sec at 72°C, followed by a final elongation for 10 min at 72°C. .. After in silico enzyme digestion of all the standard strains using REPK Version 1.3 [ ], a combination of five enzymes led to the best possible differentiation of the above pathogens: Hpy188I, BfaI, HaeIII, BglI and SmaI (New England Biolabs, Frankfurt, Germany).

    Article Title: Immune Mediators Regulate CFTR Expression through a Bifunctional Airway-Selective Enhancer
    Article Snippet: The minimal 350-bp DHS-35kb [DHS-35kb(350)] enhancer element (hg 19, chr7:117085371 to 117085720) was amplified with Pfu DNA polymerase (for primers, see Table S1) and cloned into the pSC-B vector using a StrataClone Blunt PCR cloning kit (Agilent Technologies, Santa Clara, CA). .. Cleavage of this construct with XhoI and SmaI (New England BioLabs, Ipswich, MA) enabled Klenow DNA polymerase (New England BioLabs) fill-in with [32 P]dCTP.

    Article Title: Altered Growth, Pigmentation, and Antimicrobial Susceptibility Properties of Staphylococcusaureus Due to Loss of the Major Cold Shock Gene cspB
    Article Snippet: Agarose plugs containing cells were prepared from 175 μl of overnight cultures of each strain grown in brain heart infusion broth (Becton Dickinson Microbiology Systems, Cockeysville, MD) and digested with SmaI (New England Biolabs, Ipswich, MA). .. Plugs were placed into the well of a 1% (wt/vol) SeaKem Gold (Cambrex Biosciences Rockland Inc., Rockland, ME) agarose gel and run with the following parameters: 200 V (6 V/cm), 14°C, 5-s initial switch, 40-s final switch for 21 h. The gel was stained using ethidium bromide, destained, and then visualized using UV light.

    Article Title: Genome-wide alterations in gene methylation by the BRAF V600E mutation in papillary thyroid cancer cells
    Article Snippet: Paragraph title: Methylated CpG island amplification ... Briefly, ~5 µg of each genomic DNA from cells stably transfected with BRAF shRNA (termed ‘test’ hereafter) and vector (termed ‘control’ hereafter) were digested with 100 units of methylation-sensitive restriction endonuclease SmaI (New England Biolabs, Ipswich, MA, USA) for 16 h at 20 °C in a total volume of 100 µl, followed by digestion with 20 units of methylation-insensitive restriction endonuclease XmaI for 6 h at 37 °C, which leaves sticky ends (C/CCGGG).

    Article Title: A Klebsiella pneumoniae antibiotic resistance mechanism that subdues host defences and promotes virulence
    Article Snippet: For complementation of the 52145‐ΔmgrB mutant strain, a PCR fragment (primers: mgrB _UPFWD and mgrB _DWNRVS) comprising the coding and promoter regions of the K. pneumoniae 52145 mgrB gene was amplified using Phusion® High‐Fidelity DNA Polymerase (New England Biolabs). .. The 1,507‐bp amplicon was gel‐purified and then cloned into SmaI‐digested (New England Biolabs), Antarctic Phosphatase (New England Biolabs)‐treated pUC18R6KT‐mini‐Tn7TKm plasmid (Choi et al , ) to obtain pUC18R6KT‐mini‐Tn7TKm_Kp52145mgrB Com. pUC18R6KT‐mini‐Tn7TKm_Kp52145mgrB Com was then transformed into E. coli SY327 and thereafter into E. coli β2163. .. In addition, the transposase‐containing pTSNSK‐Tp plasmid (Crepin et al , ) was introduced to the 52145‐ΔmgrB strain by electroporation to give 52145‐ΔmgrB /pTSNSK‐Tp.

    Article Title: LSm1-7 complexes bind to specific sites in viral RNA genomes and regulate their translation and replication
    Article Snippet: Forward primers carried a T7 promoter sequence at their 5′ end. .. Amplified fragments were cloned into SmaI digested pUC18 (New England Biolabs) or pGEM-T vectors (Promega). .. Fragments IR A and IR 3ΔA were synthetically produced by Geneart and cloned into plasmid pPCR-Script-ampR (Stratagene) using KpnI and SacI restriction sites.

    Stable Transfection:

    Article Title: Genome-wide alterations in gene methylation by the BRAF V600E mutation in papillary thyroid cancer cells
    Article Snippet: The MCA assay was performed as described previously ( ). .. Briefly, ~5 µg of each genomic DNA from cells stably transfected with BRAF shRNA (termed ‘test’ hereafter) and vector (termed ‘control’ hereafter) were digested with 100 units of methylation-sensitive restriction endonuclease SmaI (New England Biolabs, Ipswich, MA, USA) for 16 h at 20 °C in a total volume of 100 µl, followed by digestion with 20 units of methylation-insensitive restriction endonuclease XmaI for 6 h at 37 °C, which leaves sticky ends (C/CCGGG). .. DNA fragments were then precipitated with ethanol and their sticky ends ligated with 0.5 nmol of unphosphorylated linkers RXMA −24/−12, as described ( ).

    Synthesized:

    Article Title: A Robust System for Production of Superabundant VP1 Recombinant AAV Vectors
    Article Snippet: The plasmid pRB21 (with P7.5 promoter) was digested with SmaI (NEB) and used as the backbone to construct all plasmids having individual AAV and Ad genes. .. The PCR products were digested by EcoRV and blunt-end ligated into pRB21. pRB21-E1a-12S plasmid was generated by replacing Bsg I/Bsm I fragment in E1a-13s with the corresponding sequence from the E1a-12s gene.

    Article Title: Role of the VP16-Binding Domain of vhs in Viral Growth, Host Shutoff Activity, and Pathogenesis
    Article Snippet: The resulting fragment was treated with T4 polynucleotide kinase (New England Biolabs, Beverly, Mass.) and cloned into SmaI (New England Biolabs, Beverly, Mass.)-digested, calf intestinal phosphatase (New England Biolabs)-treated pMECA , resulting in the construct pMECA-vhs. .. Generation of the Δ20 construct containing the deletion of the VP16-binding domain of vhs was generated as follows.

    Construct:

    Article Title: A Robust System for Production of Superabundant VP1 Recombinant AAV Vectors
    Article Snippet: Single-cell-derived cell lines were then evaluated by their abilities to produce rAAV vectors by infecting with Ad/AAV-CMV-EGFP and VW22 in 24-well plates. .. The plasmid pRB21 (with P7.5 promoter) was digested with SmaI (NEB) and used as the backbone to construct all plasmids having individual AAV and Ad genes. .. Rep68 , Rep40 , X-gene , and AAP were generated using PCR amplified from pH22.

    Article Title: Immune Mediators Regulate CFTR Expression through a Bifunctional Airway-Selective Enhancer
    Article Snippet: The minimal 350-bp DHS-35kb [DHS-35kb(350)] enhancer element (hg 19, chr7:117085371 to 117085720) was amplified with Pfu DNA polymerase (for primers, see Table S1) and cloned into the pSC-B vector using a StrataClone Blunt PCR cloning kit (Agilent Technologies, Santa Clara, CA). .. Cleavage of this construct with XhoI and SmaI (New England BioLabs, Ipswich, MA) enabled Klenow DNA polymerase (New England BioLabs) fill-in with [32 P]dCTP. .. DNase I footprinting experiments were then performed as described previously ( ).

    Article Title: Mapping Protective Regions on a Three-Dimensional Model of the Moraxella catarrhalis Vaccine Antigen Oligopeptide Permease A
    Article Snippet: SmaI and XhoI sites were added to the ends of the modified T8 construct PCR product using the primers listed in Table S1 in the supplemental material. .. The T8 DNA fragment and pGEX4T-3 plasmid were digested with SmaI and XhoI (New England BioLabs, Ipswich, MA) according to the manufacturer's instructions and ligated together using T4 DNA ligase (Invitrogen).

    Article Title: Primed CRISPR adaptation in Escherichia coli cells does not depend on conformational changes in the Cascade effector complex detected in Vitro
    Article Snippet: These data are consistent with the interference-based model of primed adaptation that is independent of effector complex conformations at the priming site. .. Constructs containing the WT g8 protospacer and its variants were cloned into plasmid pUC19 (NEB) at the single SmaI (NEB) site by blunt end ligation. .. The 73 bp insert DNA carried the target sequence (PAM and protospacer variant) in its center (see ).

    Article Title: LSm1-7 complexes bind to specific sites in viral RNA genomes and regulate their translation and replication
    Article Snippet: Paragraph title: Constructs used in gel-shift assays ... Amplified fragments were cloned into SmaI digested pUC18 (New England Biolabs) or pGEM-T vectors (Promega).

    Real-time Polymerase Chain Reaction:

    Article Title: Molecular quantification and differentiation of Candida species in biological specimens of patients with liver cirrhosis
    Article Snippet: The DNA from the standard strains, duodenal samples of patients with liver cirrhosis, and control patients that were PCR-positive, were amplified with the panfungal 18S rRNA-based oligonucleotide primer pair that was already used in the quantitative PCR, excluding an additional 5’-FAM-(6-carboxyfluorescein)-labeling of the forward primer. .. After in silico enzyme digestion of all the standard strains using REPK Version 1.3 [ ], a combination of five enzymes led to the best possible differentiation of the above pathogens: Hpy188I, BfaI, HaeIII, BglI and SmaI (New England Biolabs, Frankfurt, Germany).

    Incubation:

    Article Title: Two forms of RNA editing are required for tRNA maturation in Physarum mitochondria
    Article Snippet: Primers were phosphorylated by incubation with polynucleotide kinase prior to PCR reactions containing cirRTmet1.3 and cirmet1.4 (5′-TCGGTAGTTCGATTCTAC-3′), cirRTmet2.1 and cirmet2.2 (5′-TGGTTCGATTCCAGGCC-3′), or cirRTpro1 and cirpro2 (5′-GGGACCGAAAGGTTGC-3′). .. The resulting RT-PCR products were gel purified and ligated to pBSM13+ (Stratagene) that had been digested with SmaI (New England Biolabs) and treated with calf intestine alkaline phosphatase (Roche).

    Article Title: A Klebsiella pneumoniae antibiotic resistance mechanism that subdues host defences and promotes virulence
    Article Snippet: The 1,507‐bp amplicon was gel‐purified and then cloned into SmaI‐digested (New England Biolabs), Antarctic Phosphatase (New England Biolabs)‐treated pUC18R6KT‐mini‐Tn7TKm plasmid (Choi et al , ) to obtain pUC18R6KT‐mini‐Tn7TKm_Kp52145mgrB Com. pUC18R6KT‐mini‐Tn7TKm_Kp52145mgrB Com was then transformed into E. coli SY327 and thereafter into E. coli β2163. .. The 1,507‐bp amplicon was gel‐purified and then cloned into SmaI‐digested (New England Biolabs), Antarctic Phosphatase (New England Biolabs)‐treated pUC18R6KT‐mini‐Tn7TKm plasmid (Choi et al , ) to obtain pUC18R6KT‐mini‐Tn7TKm_Kp52145mgrB Com. pUC18R6KT‐mini‐Tn7TKm_Kp52145mgrB Com was then transformed into E. coli SY327 and thereafter into E. coli β2163.

    In Silico:

    Article Title: Molecular quantification and differentiation of Candida species in biological specimens of patients with liver cirrhosis
    Article Snippet: The purification of PCR products was performed, as described above, and the DNA content of each PCR product was quantified using the Nanodrop ND-1000 spectrophotometer (Thermo Fisher Scientific). .. After in silico enzyme digestion of all the standard strains using REPK Version 1.3 [ ], a combination of five enzymes led to the best possible differentiation of the above pathogens: Hpy188I, BfaI, HaeIII, BglI and SmaI (New England Biolabs, Frankfurt, Germany). .. For T-RFLP profiling, the PCR product from the standard strains (10 ng) and from the patient samples (40 ng) were separately digested for 3 h with 2 U of each restriction endonuclease at 37°C (Hpy188I, BfaI, HaeIII, BglI) or 25°C (SmaI).

    Expressing:

    Article Title: Chromogranin B (CHGB): Intra- and extra-cellular mechanisms to regulate catecholamine storage and release, in catecholaminergic cells and organisms
    Article Snippet: The 2034 bp PCR product was digested with SmaI and SalI (New England Biolab), purified (Cat#: 28074, Qiagen), and ligated into corresponding site of SIN18-hPGK-tdTomato-T2A vector to generate SIN18-hPGK-tdTomato-T2A-hCHGB lentiviral vector plasmid ( ). .. The 2034 bp PCR product was digested with SmaI and SalI (New England Biolab), purified (Cat#: 28074, Qiagen), and ligated into corresponding site of SIN18-hPGK-tdTomato-T2A vector to generate SIN18-hPGK-tdTomato-T2A-hCHGB lentiviral vector plasmid ( ).

    Article Title: Role of the VP16-Binding Domain of vhs in Viral Growth, Host Shutoff Activity, and Pathogenesis
    Article Snippet: In order to generate the vhs expression constructs used in the transient-transfection assays, a fragment containing the entire vhs open reading frame was generated by PCR amplification of the pUL41 plasmid that had been generated previously ( ). .. The resulting fragment was treated with T4 polynucleotide kinase (New England Biolabs, Beverly, Mass.) and cloned into SmaI (New England Biolabs, Beverly, Mass.)-digested, calf intestinal phosphatase (New England Biolabs)-treated pMECA , resulting in the construct pMECA-vhs.

    Article Title: Mapping Protective Regions on a Three-Dimensional Model of the Moraxella catarrhalis Vaccine Antigen Oligopeptide Permease A
    Article Snippet: The T8 DNA fragment and pGEX4T-3 plasmid were digested with SmaI and XhoI (New England BioLabs, Ipswich, MA) according to the manufacturer's instructions and ligated together using T4 DNA ligase (Invitrogen). .. The GST-T8 plasmid construct was transformed and expressed in BL21(DE3) Star cells.

    Modification:

    Article Title: Mapping Protective Regions on a Three-Dimensional Model of the Moraxella catarrhalis Vaccine Antigen Oligopeptide Permease A
    Article Snippet: SmaI and XhoI sites were added to the ends of the modified T8 construct PCR product using the primers listed in Table S1 in the supplemental material. .. The T8 DNA fragment and pGEX4T-3 plasmid were digested with SmaI and XhoI (New England BioLabs, Ipswich, MA) according to the manufacturer's instructions and ligated together using T4 DNA ligase (Invitrogen).

    Transformation Assay:

    Article Title: Mapping Protective Regions on a Three-Dimensional Model of the Moraxella catarrhalis Vaccine Antigen Oligopeptide Permease A
    Article Snippet: The T8 DNA fragment and pGEX4T-3 plasmid were digested with SmaI and XhoI (New England BioLabs, Ipswich, MA) according to the manufacturer's instructions and ligated together using T4 DNA ligase (Invitrogen). .. The reaction product was transformed into TOP10 cells for plasmid replication and isolated using a QIAprep miniprep kit (Qiagen, Hilden, Germany).

    Article Title: A Klebsiella pneumoniae antibiotic resistance mechanism that subdues host defences and promotes virulence
    Article Snippet: For complementation of the 52145‐ΔmgrB mutant strain, a PCR fragment (primers: mgrB _UPFWD and mgrB _DWNRVS) comprising the coding and promoter regions of the K. pneumoniae 52145 mgrB gene was amplified using Phusion® High‐Fidelity DNA Polymerase (New England Biolabs). .. The 1,507‐bp amplicon was gel‐purified and then cloned into SmaI‐digested (New England Biolabs), Antarctic Phosphatase (New England Biolabs)‐treated pUC18R6KT‐mini‐Tn7TKm plasmid (Choi et al , ) to obtain pUC18R6KT‐mini‐Tn7TKm_Kp52145mgrB Com. pUC18R6KT‐mini‐Tn7TKm_Kp52145mgrB Com was then transformed into E. coli SY327 and thereafter into E. coli β2163. .. In addition, the transposase‐containing pTSNSK‐Tp plasmid (Crepin et al , ) was introduced to the 52145‐ΔmgrB strain by electroporation to give 52145‐ΔmgrB /pTSNSK‐Tp.

    Article Title: Primed CRISPR adaptation in Escherichia coli cells does not depend on conformational changes in the Cascade effector complex detected in Vitro
    Article Snippet: Constructs containing the WT g8 protospacer and its variants were cloned into plasmid pUC19 (NEB) at the single SmaI (NEB) site by blunt end ligation. .. The 73 bp insert DNA carried the target sequence (PAM and protospacer variant) in its center (see ).

    Over Expression:

    Article Title: Chromogranin B (CHGB): Intra- and extra-cellular mechanisms to regulate catecholamine storage and release, in catecholaminergic cells and organisms
    Article Snippet: Paragraph title: Over-expression of CHGB in PC12 cells by transduction of a CHGB lentivirus ... The 2034 bp PCR product was digested with SmaI and SalI (New England Biolab), purified (Cat#: 28074, Qiagen), and ligated into corresponding site of SIN18-hPGK-tdTomato-T2A vector to generate SIN18-hPGK-tdTomato-T2A-hCHGB lentiviral vector plasmid ( ).

    Hybridization:

    Article Title: Altered Growth, Pigmentation, and Antimicrobial Susceptibility Properties of Staphylococcusaureus Due to Loss of the Major Cold Shock Gene cspB
    Article Snippet: Paragraph title: PFGE and Southern hybridization. ... Agarose plugs containing cells were prepared from 175 μl of overnight cultures of each strain grown in brain heart infusion broth (Becton Dickinson Microbiology Systems, Cockeysville, MD) and digested with SmaI (New England Biolabs, Ipswich, MA).

    MCA Assay:

    Article Title: Genome-wide alterations in gene methylation by the BRAF V600E mutation in papillary thyroid cancer cells
    Article Snippet: Briefly, ~5 µg of each genomic DNA from cells stably transfected with BRAF shRNA (termed ‘test’ hereafter) and vector (termed ‘control’ hereafter) were digested with 100 units of methylation-sensitive restriction endonuclease SmaI (New England Biolabs, Ipswich, MA, USA) for 16 h at 20 °C in a total volume of 100 µl, followed by digestion with 20 units of methylation-insensitive restriction endonuclease XmaI for 6 h at 37 °C, which leaves sticky ends (C/CCGGG). .. Briefly, ~5 µg of each genomic DNA from cells stably transfected with BRAF shRNA (termed ‘test’ hereafter) and vector (termed ‘control’ hereafter) were digested with 100 units of methylation-sensitive restriction endonuclease SmaI (New England Biolabs, Ipswich, MA, USA) for 16 h at 20 °C in a total volume of 100 µl, followed by digestion with 20 units of methylation-insensitive restriction endonuclease XmaI for 6 h at 37 °C, which leaves sticky ends (C/CCGGG).

    Transfection:

    Article Title: Chromogranin B (CHGB): Intra- and extra-cellular mechanisms to regulate catecholamine storage and release, in catecholaminergic cells and organisms
    Article Snippet: The 2034 bp PCR product was digested with SmaI and SalI (New England Biolab), purified (Cat#: 28074, Qiagen), and ligated into corresponding site of SIN18-hPGK-tdTomato-T2A vector to generate SIN18-hPGK-tdTomato-T2A-hCHGB lentiviral vector plasmid ( ). .. The 2034 bp PCR product was digested with SmaI and SalI (New England Biolab), purified (Cat#: 28074, Qiagen), and ligated into corresponding site of SIN18-hPGK-tdTomato-T2A vector to generate SIN18-hPGK-tdTomato-T2A-hCHGB lentiviral vector plasmid ( ).

    Article Title: Genome-wide alterations in gene methylation by the BRAF V600E mutation in papillary thyroid cancer cells
    Article Snippet: The MCA assay was performed as described previously ( ). .. Briefly, ~5 µg of each genomic DNA from cells stably transfected with BRAF shRNA (termed ‘test’ hereafter) and vector (termed ‘control’ hereafter) were digested with 100 units of methylation-sensitive restriction endonuclease SmaI (New England Biolabs, Ipswich, MA, USA) for 16 h at 20 °C in a total volume of 100 µl, followed by digestion with 20 units of methylation-insensitive restriction endonuclease XmaI for 6 h at 37 °C, which leaves sticky ends (C/CCGGG). .. DNA fragments were then precipitated with ethanol and their sticky ends ligated with 0.5 nmol of unphosphorylated linkers RXMA −24/−12, as described ( ).

    Southern Blot:

    Article Title: RIP140 in thyroid hormone-repression and chromatin remodeling of Crabp1 gene during adipocyte differentiation
    Article Snippet: The purified DNA was subjected to Southern blot analysis. .. Isolated nuclei from differentiating 3T3-L1 cells were digested with 100 U of PstI, XhoI, SmaI, SpeI and ApaLI (New England Biolabs) for 30 min.

    Article Title: Altered Growth, Pigmentation, and Antimicrobial Susceptibility Properties of Staphylococcusaureus Due to Loss of the Major Cold Shock Gene cspB
    Article Snippet: Agarose plugs containing cells were prepared from 175 μl of overnight cultures of each strain grown in brain heart infusion broth (Becton Dickinson Microbiology Systems, Cockeysville, MD) and digested with SmaI (New England Biolabs, Ipswich, MA). .. Agarose plugs containing cells were prepared from 175 μl of overnight cultures of each strain grown in brain heart infusion broth (Becton Dickinson Microbiology Systems, Cockeysville, MD) and digested with SmaI (New England Biolabs, Ipswich, MA).

    Ligation:

    Article Title: RIP140 in thyroid hormone-repression and chromatin remodeling of Crabp1 gene during adipocyte differentiation
    Article Snippet: Micrococcal nuclease (MNase) digestion and ligation-mediated PCR (LM-PCR) were performed as described ( ). .. Isolated nuclei from differentiating 3T3-L1 cells were digested with 100 U of PstI, XhoI, SmaI, SpeI and ApaLI (New England Biolabs) for 30 min.

    Article Title: High-throughput methylation profiling by MCA coupled to CpG island microarray
    Article Snippet: In brief, 5 μg of DNA was digested with 100 units of SmaI for 16 h (all restriction enzymes were from New England Biolabs), followed by digestion with 20 units of XmaI for 6 h. DNA fragments were then precipitated with ethanol. .. RXMA PCR adaptors were prepared by incubation of the oligonucleotides RXMA24 (5′-AGCACTCTCCAGCCTCTCACCGAC-3′) and RXMA12 (5′-CCGGGTCGGTGA-3′) for 2 min at 65°C, followed by cooling to room temperature for 1 h. DNA (0.5 μg) was ligated to 0.5 nmol of RXMA adaptor using T4 DNA ligase (New England Biolabs).

    Article Title: Primed CRISPR adaptation in Escherichia coli cells does not depend on conformational changes in the Cascade effector complex detected in Vitro
    Article Snippet: These data are consistent with the interference-based model of primed adaptation that is independent of effector complex conformations at the priming site. .. Constructs containing the WT g8 protospacer and its variants were cloned into plasmid pUC19 (NEB) at the single SmaI (NEB) site by blunt end ligation. .. The 73 bp insert DNA carried the target sequence (PAM and protospacer variant) in its center (see ).

    Footprinting:

    Article Title: Immune Mediators Regulate CFTR Expression through a Bifunctional Airway-Selective Enhancer
    Article Snippet: Paragraph title: In vitro DNase I footprinting. ... Cleavage of this construct with XhoI and SmaI (New England BioLabs, Ipswich, MA) enabled Klenow DNA polymerase (New England BioLabs) fill-in with [32 P]dCTP.

    Generated:

    Article Title: A Robust System for Production of Superabundant VP1 Recombinant AAV Vectors
    Article Snippet: The plasmid pRB21 (with P7.5 promoter) was digested with SmaI (NEB) and used as the backbone to construct all plasmids having individual AAV and Ad genes. .. E1a-13S , E1b-55K , E1b-19k , E2a , E4orf6 , and VA I RNA were amplified using the pFΔ6 plasmid as a template.

    Article Title: Role of the VP16-Binding Domain of vhs in Viral Growth, Host Shutoff Activity, and Pathogenesis
    Article Snippet: In order to generate the vhs expression constructs used in the transient-transfection assays, a fragment containing the entire vhs open reading frame was generated by PCR amplification of the pUL41 plasmid that had been generated previously ( ). .. The resulting fragment was treated with T4 polynucleotide kinase (New England Biolabs, Beverly, Mass.) and cloned into SmaI (New England Biolabs, Beverly, Mass.)-digested, calf intestinal phosphatase (New England Biolabs)-treated pMECA , resulting in the construct pMECA-vhs.

    Article Title: Mapping Protective Regions on a Three-Dimensional Model of the Moraxella catarrhalis Vaccine Antigen Oligopeptide Permease A
    Article Snippet: This shortened version of the construct generated a product that was more stable than the previous His-tagged construct. .. The T8 DNA fragment and pGEX4T-3 plasmid were digested with SmaI and XhoI (New England BioLabs, Ipswich, MA) according to the manufacturer's instructions and ligated together using T4 DNA ligase (Invitrogen).

    Article Title: LSm1-7 complexes bind to specific sites in viral RNA genomes and regulate their translation and replication
    Article Snippet: Unless otherwise mentioned, constructs were generated by PCR amplification of the fragments of interest from BMV RNA1, RNA2, and RNA3 genomes cloned, respectively, in the plasmids pB1tp3, pB2tp5, and pB3tp8 ( ; ). .. Amplified fragments were cloned into SmaI digested pUC18 (New England Biolabs) or pGEM-T vectors (Promega).

    Polymerase Chain Reaction:

    Article Title: RIP140 in thyroid hormone-repression and chromatin remodeling of Crabp1 gene during adipocyte differentiation
    Article Snippet: Micrococcal nuclease (MNase) digestion and ligation-mediated PCR (LM-PCR) were performed as described ( ). .. Isolated nuclei from differentiating 3T3-L1 cells were digested with 100 U of PstI, XhoI, SmaI, SpeI and ApaLI (New England Biolabs) for 30 min.

    Article Title: A Robust System for Production of Superabundant VP1 Recombinant AAV Vectors
    Article Snippet: The plasmid pRB21 (with P7.5 promoter) was digested with SmaI (NEB) and used as the backbone to construct all plasmids having individual AAV and Ad genes. .. E1a-13S , E1b-55K , E1b-19k , E2a , E4orf6 , and VA I RNA were amplified using the pFΔ6 plasmid as a template.

    Article Title: Molecular quantification and differentiation of Candida species in biological specimens of patients with liver cirrhosis
    Article Snippet: The purification of PCR products was performed, as described above, and the DNA content of each PCR product was quantified using the Nanodrop ND-1000 spectrophotometer (Thermo Fisher Scientific). .. After in silico enzyme digestion of all the standard strains using REPK Version 1.3 [ ], a combination of five enzymes led to the best possible differentiation of the above pathogens: Hpy188I, BfaI, HaeIII, BglI and SmaI (New England Biolabs, Frankfurt, Germany).

    Article Title: Immune Mediators Regulate CFTR Expression through a Bifunctional Airway-Selective Enhancer
    Article Snippet: The minimal 350-bp DHS-35kb [DHS-35kb(350)] enhancer element (hg 19, chr7:117085371 to 117085720) was amplified with Pfu DNA polymerase (for primers, see Table S1) and cloned into the pSC-B vector using a StrataClone Blunt PCR cloning kit (Agilent Technologies, Santa Clara, CA). .. Cleavage of this construct with XhoI and SmaI (New England BioLabs, Ipswich, MA) enabled Klenow DNA polymerase (New England BioLabs) fill-in with [32 P]dCTP.

    Article Title: Chromogranin B (CHGB): Intra- and extra-cellular mechanisms to regulate catecholamine storage and release, in catecholaminergic cells and organisms
    Article Snippet: The 5′ primer, 5’- agcCCCGGGatgcagccaacgctgcttct -3’, includes a SmaI site (capitalized), while the 3′ primer, 5′- caaGTCGACtcagcccctttggctgaatt -3’, includes a SalI site (capitalized). .. The 2034 bp PCR product was digested with SmaI and SalI (New England Biolab), purified (Cat#: 28074, Qiagen), and ligated into corresponding site of SIN18-hPGK-tdTomato-T2A vector to generate SIN18-hPGK-tdTomato-T2A-hCHGB lentiviral vector plasmid ( ). .. The SIN18-hPGK-tdTomato is used as control lentiviral vector.

    Article Title: Altered Growth, Pigmentation, and Antimicrobial Susceptibility Properties of Staphylococcusaureus Due to Loss of the Major Cold Shock Gene cspB
    Article Snippet: Agarose plugs containing cells were prepared from 175 μl of overnight cultures of each strain grown in brain heart infusion broth (Becton Dickinson Microbiology Systems, Cockeysville, MD) and digested with SmaI (New England Biolabs, Ipswich, MA). .. Plugs were placed into the well of a 1% (wt/vol) SeaKem Gold (Cambrex Biosciences Rockland Inc., Rockland, ME) agarose gel and run with the following parameters: 200 V (6 V/cm), 14°C, 5-s initial switch, 40-s final switch for 21 h. The gel was stained using ethidium bromide, destained, and then visualized using UV light.

    Article Title: High-throughput methylation profiling by MCA coupled to CpG island microarray
    Article Snippet: In brief, 5 μg of DNA was digested with 100 units of SmaI for 16 h (all restriction enzymes were from New England Biolabs), followed by digestion with 20 units of XmaI for 6 h. DNA fragments were then precipitated with ethanol. .. RXMA PCR adaptors were prepared by incubation of the oligonucleotides RXMA24 (5′-AGCACTCTCCAGCCTCTCACCGAC-3′) and RXMA12 (5′-CCGGGTCGGTGA-3′) for 2 min at 65°C, followed by cooling to room temperature for 1 h. DNA (0.5 μg) was ligated to 0.5 nmol of RXMA adaptor using T4 DNA ligase (New England Biolabs).

    Article Title: Two forms of RNA editing are required for tRNA maturation in Physarum mitochondria
    Article Snippet: Paragraph title: RT-PCR and PCR reactions ... The resulting RT-PCR products were gel purified and ligated to pBSM13+ (Stratagene) that had been digested with SmaI (New England Biolabs) and treated with calf intestine alkaline phosphatase (Roche).

    Article Title: Mapping Protective Regions on a Three-Dimensional Model of the Moraxella catarrhalis Vaccine Antigen Oligopeptide Permease A
    Article Snippet: SmaI and XhoI sites were added to the ends of the modified T8 construct PCR product using the primers listed in Table S1 in the supplemental material. .. The T8 DNA fragment and pGEX4T-3 plasmid were digested with SmaI and XhoI (New England BioLabs, Ipswich, MA) according to the manufacturer's instructions and ligated together using T4 DNA ligase (Invitrogen).

    Article Title: Genome-wide alterations in gene methylation by the BRAF V600E mutation in papillary thyroid cancer cells
    Article Snippet: Briefly, ~5 µg of each genomic DNA from cells stably transfected with BRAF shRNA (termed ‘test’ hereafter) and vector (termed ‘control’ hereafter) were digested with 100 units of methylation-sensitive restriction endonuclease SmaI (New England Biolabs, Ipswich, MA, USA) for 16 h at 20 °C in a total volume of 100 µl, followed by digestion with 20 units of methylation-insensitive restriction endonuclease XmaI for 6 h at 37 °C, which leaves sticky ends (C/CCGGG). .. The oligonucleotide sequences were as follows: (RXMA − 24) 5′-AGC ACT CTC CAG CCT CTC ACC GAC-3′ and (RXMA − 12) 5′-CCG GGT CGG TGA-3′.

    Article Title: A Klebsiella pneumoniae antibiotic resistance mechanism that subdues host defences and promotes virulence
    Article Snippet: For complementation of the 52145‐ΔmgrB mutant strain, a PCR fragment (primers: mgrB _UPFWD and mgrB _DWNRVS) comprising the coding and promoter regions of the K. pneumoniae 52145 mgrB gene was amplified using Phusion® High‐Fidelity DNA Polymerase (New England Biolabs). .. The 1,507‐bp amplicon was gel‐purified and then cloned into SmaI‐digested (New England Biolabs), Antarctic Phosphatase (New England Biolabs)‐treated pUC18R6KT‐mini‐Tn7TKm plasmid (Choi et al , ) to obtain pUC18R6KT‐mini‐Tn7TKm_Kp52145mgrB Com. pUC18R6KT‐mini‐Tn7TKm_Kp52145mgrB Com was then transformed into E. coli SY327 and thereafter into E. coli β2163.

    Article Title: LSm1-7 complexes bind to specific sites in viral RNA genomes and regulate their translation and replication
    Article Snippet: Unless otherwise mentioned, constructs were generated by PCR amplification of the fragments of interest from BMV RNA1, RNA2, and RNA3 genomes cloned, respectively, in the plasmids pB1tp3, pB2tp5, and pB3tp8 ( ; ). .. Amplified fragments were cloned into SmaI digested pUC18 (New England Biolabs) or pGEM-T vectors (Promega).

    Plasmid Preparation:

    Article Title: A Robust System for Production of Superabundant VP1 Recombinant AAV Vectors
    Article Snippet: Single-cell-derived cell lines were then evaluated by their abilities to produce rAAV vectors by infecting with Ad/AAV-CMV-EGFP and VW22 in 24-well plates. .. The plasmid pRB21 (with P7.5 promoter) was digested with SmaI (NEB) and used as the backbone to construct all plasmids having individual AAV and Ad genes. .. Rep68 , Rep40 , X-gene , and AAP were generated using PCR amplified from pH22.

    Article Title: Immune Mediators Regulate CFTR Expression through a Bifunctional Airway-Selective Enhancer
    Article Snippet: The minimal 350-bp DHS-35kb [DHS-35kb(350)] enhancer element (hg 19, chr7:117085371 to 117085720) was amplified with Pfu DNA polymerase (for primers, see Table S1) and cloned into the pSC-B vector using a StrataClone Blunt PCR cloning kit (Agilent Technologies, Santa Clara, CA). .. Cleavage of this construct with XhoI and SmaI (New England BioLabs, Ipswich, MA) enabled Klenow DNA polymerase (New England BioLabs) fill-in with [32 P]dCTP.

    Article Title: Chromogranin B (CHGB): Intra- and extra-cellular mechanisms to regulate catecholamine storage and release, in catecholaminergic cells and organisms
    Article Snippet: The 5′ primer, 5’- agcCCCGGGatgcagccaacgctgcttct -3’, includes a SmaI site (capitalized), while the 3′ primer, 5′- caaGTCGACtcagcccctttggctgaatt -3’, includes a SalI site (capitalized). .. The 2034 bp PCR product was digested with SmaI and SalI (New England Biolab), purified (Cat#: 28074, Qiagen), and ligated into corresponding site of SIN18-hPGK-tdTomato-T2A vector to generate SIN18-hPGK-tdTomato-T2A-hCHGB lentiviral vector plasmid ( ). .. The SIN18-hPGK-tdTomato is used as control lentiviral vector.

    Article Title: Role of the VP16-Binding Domain of vhs in Viral Growth, Host Shutoff Activity, and Pathogenesis
    Article Snippet: In order to generate the vhs expression constructs used in the transient-transfection assays, a fragment containing the entire vhs open reading frame was generated by PCR amplification of the pUL41 plasmid that had been generated previously ( ). .. The resulting fragment was treated with T4 polynucleotide kinase (New England Biolabs, Beverly, Mass.) and cloned into SmaI (New England Biolabs, Beverly, Mass.)-digested, calf intestinal phosphatase (New England Biolabs)-treated pMECA , resulting in the construct pMECA-vhs.

    Article Title: Mapping Protective Regions on a Three-Dimensional Model of the Moraxella catarrhalis Vaccine Antigen Oligopeptide Permease A
    Article Snippet: SmaI and XhoI sites were added to the ends of the modified T8 construct PCR product using the primers listed in Table S1 in the supplemental material. .. The T8 DNA fragment and pGEX4T-3 plasmid were digested with SmaI and XhoI (New England BioLabs, Ipswich, MA) according to the manufacturer's instructions and ligated together using T4 DNA ligase (Invitrogen). .. The reaction product was transformed into TOP10 cells for plasmid replication and isolated using a QIAprep miniprep kit (Qiagen, Hilden, Germany).

    Article Title: Genome-wide alterations in gene methylation by the BRAF V600E mutation in papillary thyroid cancer cells
    Article Snippet: The MCA assay was performed as described previously ( ). .. Briefly, ~5 µg of each genomic DNA from cells stably transfected with BRAF shRNA (termed ‘test’ hereafter) and vector (termed ‘control’ hereafter) were digested with 100 units of methylation-sensitive restriction endonuclease SmaI (New England Biolabs, Ipswich, MA, USA) for 16 h at 20 °C in a total volume of 100 µl, followed by digestion with 20 units of methylation-insensitive restriction endonuclease XmaI for 6 h at 37 °C, which leaves sticky ends (C/CCGGG). .. DNA fragments were then precipitated with ethanol and their sticky ends ligated with 0.5 nmol of unphosphorylated linkers RXMA −24/−12, as described ( ).

    Article Title: A Klebsiella pneumoniae antibiotic resistance mechanism that subdues host defences and promotes virulence
    Article Snippet: For complementation of the 52145‐ΔmgrB mutant strain, a PCR fragment (primers: mgrB _UPFWD and mgrB _DWNRVS) comprising the coding and promoter regions of the K. pneumoniae 52145 mgrB gene was amplified using Phusion® High‐Fidelity DNA Polymerase (New England Biolabs). .. The 1,507‐bp amplicon was gel‐purified and then cloned into SmaI‐digested (New England Biolabs), Antarctic Phosphatase (New England Biolabs)‐treated pUC18R6KT‐mini‐Tn7TKm plasmid (Choi et al , ) to obtain pUC18R6KT‐mini‐Tn7TKm_Kp52145mgrB Com. pUC18R6KT‐mini‐Tn7TKm_Kp52145mgrB Com was then transformed into E. coli SY327 and thereafter into E. coli β2163. .. In addition, the transposase‐containing pTSNSK‐Tp plasmid (Crepin et al , ) was introduced to the 52145‐ΔmgrB strain by electroporation to give 52145‐ΔmgrB /pTSNSK‐Tp.

    Article Title: Primed CRISPR adaptation in Escherichia coli cells does not depend on conformational changes in the Cascade effector complex detected in Vitro
    Article Snippet: These data are consistent with the interference-based model of primed adaptation that is independent of effector complex conformations at the priming site. .. Constructs containing the WT g8 protospacer and its variants were cloned into plasmid pUC19 (NEB) at the single SmaI (NEB) site by blunt end ligation. .. The 73 bp insert DNA carried the target sequence (PAM and protospacer variant) in its center (see ).

    Article Title: LSm1-7 complexes bind to specific sites in viral RNA genomes and regulate their translation and replication
    Article Snippet: Amplified fragments were cloned into SmaI digested pUC18 (New England Biolabs) or pGEM-T vectors (Promega). .. Fragments IR A and IR 3ΔA were synthetically produced by Geneart and cloned into plasmid pPCR-Script-ampR (Stratagene) using KpnI and SacI restriction sites.

    DNA Labeling:

    Article Title: Altered Growth, Pigmentation, and Antimicrobial Susceptibility Properties of Staphylococcusaureus Due to Loss of the Major Cold Shock Gene cspB
    Article Snippet: Agarose plugs containing cells were prepared from 175 μl of overnight cultures of each strain grown in brain heart infusion broth (Becton Dickinson Microbiology Systems, Cockeysville, MD) and digested with SmaI (New England Biolabs, Ipswich, MA). .. For Southern hybridization, a 391-bp fragment of mecA was PCR amplified from S . aureus COL genomic DNA using primers mecA5F and mecA5R and used as a probe in Southern blot analysis.

    Reverse Transcription Polymerase Chain Reaction:

    Article Title: Two forms of RNA editing are required for tRNA maturation in Physarum mitochondria
    Article Snippet: Primers were phosphorylated by incubation with polynucleotide kinase prior to PCR reactions containing cirRTmet1.3 and cirmet1.4 (5′-TCGGTAGTTCGATTCTAC-3′), cirRTmet2.1 and cirmet2.2 (5′-TGGTTCGATTCCAGGCC-3′), or cirRTpro1 and cirpro2 (5′-GGGACCGAAAGGTTGC-3′). .. The resulting RT-PCR products were gel purified and ligated to pBSM13+ (Stratagene) that had been digested with SmaI (New England Biolabs) and treated with calf intestine alkaline phosphatase (Roche). .. For cDNA synthesis from nascent transcripts, 2 μg of total mitochondrial RNA in 10 mM Tris (pH 8.3, at 42°C) and 250 mM KCl were mixed with primer cirRTpro1 , heated for 3 min to 95°C, incubated for 10 min at 65°C, and then held on ice.

    Recombinant:

    Article Title: A Robust System for Production of Superabundant VP1 Recombinant AAV Vectors
    Article Snippet: Paragraph title: Construction and Generation of Recombinant VV Vectors ... The plasmid pRB21 (with P7.5 promoter) was digested with SmaI (NEB) and used as the backbone to construct all plasmids having individual AAV and Ad genes.

    Article Title: Role of the VP16-Binding Domain of vhs in Viral Growth, Host Shutoff Activity, and Pathogenesis
    Article Snippet: Paragraph title: Generation of plasmids and recombinant viruses. ... The resulting fragment was treated with T4 polynucleotide kinase (New England Biolabs, Beverly, Mass.) and cloned into SmaI (New England Biolabs, Beverly, Mass.)-digested, calf intestinal phosphatase (New England Biolabs)-treated pMECA , resulting in the construct pMECA-vhs.

    Pulsed-Field Gel:

    Article Title: In Vivo Transfer of the vanA Resistance Gene from an Enterococcus faecium Isolate of Animal Origin to an E. faecium Isolate of Human Origin in the Intestines of Human Volunteers
    Article Snippet: Paragraph title: Pulsed-field gel electrophoresis (PFGE). ... A small slice of the agarose plug was digested with 30 U of the restriction enzyme SmaI (New England BioLabs Inc., Medinova Scientific A/S, Denmark) for a minimum of 4 h at 25°C.

    Article Title: Interference in Pheromone-Responsive Conjugation of a High-Level Bacitracin Resistant Enterococcus faecalis Plasmid of Poultry Origin
    Article Snippet: Paragraph title: 2.9. Pulsed-Field Gel Electrophoresis (PFGE) ... Briefly, a plug slice of 2 to 4 mm wide was suspended in a total volume of 227 µL of the manufacturer’s recommended restriction buffer and 20 U of SmaI (New England Biolabs Inc., Mississauga, ON, Canada).

    Article Title: Novel Structure of Enterococcus faecium-Originated ermB-Positive Tn1546-Like Element in Staphylococcus aureus
    Article Snippet: The genetic relatedness of the 97 ermB -carrying MSSA isolates was determined by pulsed-field gel electrophoresis (PFGE). .. The DNA in gel plugs was digested with SmaI (New England BioLabs, Ipswich, MA) and then separated in a CHEF-DR III apparatus (Bio-Rad Laboratories).

    Article Title: Altered Growth, Pigmentation, and Antimicrobial Susceptibility Properties of Staphylococcusaureus Due to Loss of the Major Cold Shock Gene cspB
    Article Snippet: Pulsed-field gel electrophoresis (PFGE) was performed as outlined by PulseNet, the National Molecular Subtyping Network for Foodborne Disease Surveillance of the CDC (Atlanta, GA) ( ). .. Agarose plugs containing cells were prepared from 175 μl of overnight cultures of each strain grown in brain heart infusion broth (Becton Dickinson Microbiology Systems, Cockeysville, MD) and digested with SmaI (New England Biolabs, Ipswich, MA).

    Methylation:

    Article Title: High-throughput methylation profiling by MCA coupled to CpG island microarray
    Article Snippet: Paragraph title: Methylated CpG island amplication (MCA) ... In brief, 5 μg of DNA was digested with 100 units of SmaI for 16 h (all restriction enzymes were from New England Biolabs), followed by digestion with 20 units of XmaI for 6 h. DNA fragments were then precipitated with ethanol.

    Article Title: Genome-wide alterations in gene methylation by the BRAF V600E mutation in papillary thyroid cancer cells
    Article Snippet: The MCA assay was performed as described previously ( ). .. Briefly, ~5 µg of each genomic DNA from cells stably transfected with BRAF shRNA (termed ‘test’ hereafter) and vector (termed ‘control’ hereafter) were digested with 100 units of methylation-sensitive restriction endonuclease SmaI (New England Biolabs, Ipswich, MA, USA) for 16 h at 20 °C in a total volume of 100 µl, followed by digestion with 20 units of methylation-insensitive restriction endonuclease XmaI for 6 h at 37 °C, which leaves sticky ends (C/CCGGG). .. DNA fragments were then precipitated with ethanol and their sticky ends ligated with 0.5 nmol of unphosphorylated linkers RXMA −24/−12, as described ( ).

    Mutagenesis:

    Article Title: Role of the VP16-Binding Domain of vhs in Viral Growth, Host Shutoff Activity, and Pathogenesis
    Article Snippet: The resulting fragment was treated with T4 polynucleotide kinase (New England Biolabs, Beverly, Mass.) and cloned into SmaI (New England Biolabs, Beverly, Mass.)-digested, calf intestinal phosphatase (New England Biolabs)-treated pMECA , resulting in the construct pMECA-vhs. .. An oligonucleotide complementary to 20 bp on either side of the VP16-binding domain was synthesized.

    Article Title: A Klebsiella pneumoniae antibiotic resistance mechanism that subdues host defences and promotes virulence
    Article Snippet: Paragraph title: Complementation of the 52145‐ΔmgrB mutant ... The 1,507‐bp amplicon was gel‐purified and then cloned into SmaI‐digested (New England Biolabs), Antarctic Phosphatase (New England Biolabs)‐treated pUC18R6KT‐mini‐Tn7TKm plasmid (Choi et al , ) to obtain pUC18R6KT‐mini‐Tn7TKm_Kp52145mgrB Com. pUC18R6KT‐mini‐Tn7TKm_Kp52145mgrB Com was then transformed into E. coli SY327 and thereafter into E. coli β2163.

    Isolation:

    Article Title: RIP140 in thyroid hormone-repression and chromatin remodeling of Crabp1 gene during adipocyte differentiation
    Article Snippet: Restriction enzyme accessibility assay was carried out as described ( ). .. Isolated nuclei from differentiating 3T3-L1 cells were digested with 100 U of PstI, XhoI, SmaI, SpeI and ApaLI (New England Biolabs) for 30 min. .. The purified genomic DNA was re-digested with 100 U of ApaI completely (for first digestion with XhoI, PstI and SmaI) or 100 U of PstI (for first digestion with SpeI and ApaLI).

    Article Title: Mapping Protective Regions on a Three-Dimensional Model of the Moraxella catarrhalis Vaccine Antigen Oligopeptide Permease A
    Article Snippet: The T8 DNA fragment and pGEX4T-3 plasmid were digested with SmaI and XhoI (New England BioLabs, Ipswich, MA) according to the manufacturer's instructions and ligated together using T4 DNA ligase (Invitrogen). .. The GST-T8 plasmid construct was transformed and expressed in BL21(DE3) Star cells.

    Size-exclusion Chromatography:

    Article Title: Molecular quantification and differentiation of Candida species in biological specimens of patients with liver cirrhosis
    Article Snippet: The amplification steps were as follows: initial denaturation at 95°C for 1 min, 40 cycles of 15 sec at 95°C, 15 sec at 55°C and 10 sec at 72°C, followed by a final elongation for 10 min at 72°C. .. After in silico enzyme digestion of all the standard strains using REPK Version 1.3 [ ], a combination of five enzymes led to the best possible differentiation of the above pathogens: Hpy188I, BfaI, HaeIII, BglI and SmaI (New England Biolabs, Frankfurt, Germany).

    Electrophoretic Mobility Shift Assay:

    Article Title: LSm1-7 complexes bind to specific sites in viral RNA genomes and regulate their translation and replication
    Article Snippet: Paragraph title: Constructs used in gel-shift assays ... Amplified fragments were cloned into SmaI digested pUC18 (New England Biolabs) or pGEM-T vectors (Promega).

    Purification:

    Article Title: RIP140 in thyroid hormone-repression and chromatin remodeling of Crabp1 gene during adipocyte differentiation
    Article Snippet: The purified DNA was subjected to Southern blot analysis. .. Isolated nuclei from differentiating 3T3-L1 cells were digested with 100 U of PstI, XhoI, SmaI, SpeI and ApaLI (New England Biolabs) for 30 min.

    Article Title: Molecular quantification and differentiation of Candida species in biological specimens of patients with liver cirrhosis
    Article Snippet: The purification of PCR products was performed, as described above, and the DNA content of each PCR product was quantified using the Nanodrop ND-1000 spectrophotometer (Thermo Fisher Scientific). .. After in silico enzyme digestion of all the standard strains using REPK Version 1.3 [ ], a combination of five enzymes led to the best possible differentiation of the above pathogens: Hpy188I, BfaI, HaeIII, BglI and SmaI (New England Biolabs, Frankfurt, Germany).

    Article Title: Chromogranin B (CHGB): Intra- and extra-cellular mechanisms to regulate catecholamine storage and release, in catecholaminergic cells and organisms
    Article Snippet: The 5′ primer, 5’- agcCCCGGGatgcagccaacgctgcttct -3’, includes a SmaI site (capitalized), while the 3′ primer, 5′- caaGTCGACtcagcccctttggctgaatt -3’, includes a SalI site (capitalized). .. The 2034 bp PCR product was digested with SmaI and SalI (New England Biolab), purified (Cat#: 28074, Qiagen), and ligated into corresponding site of SIN18-hPGK-tdTomato-T2A vector to generate SIN18-hPGK-tdTomato-T2A-hCHGB lentiviral vector plasmid ( ). .. The SIN18-hPGK-tdTomato is used as control lentiviral vector.

    Article Title: Two forms of RNA editing are required for tRNA maturation in Physarum mitochondria
    Article Snippet: Primers were phosphorylated by incubation with polynucleotide kinase prior to PCR reactions containing cirRTmet1.3 and cirmet1.4 (5′-TCGGTAGTTCGATTCTAC-3′), cirRTmet2.1 and cirmet2.2 (5′-TGGTTCGATTCCAGGCC-3′), or cirRTpro1 and cirpro2 (5′-GGGACCGAAAGGTTGC-3′). .. The resulting RT-PCR products were gel purified and ligated to pBSM13+ (Stratagene) that had been digested with SmaI (New England Biolabs) and treated with calf intestine alkaline phosphatase (Roche). .. For cDNA synthesis from nascent transcripts, 2 μg of total mitochondrial RNA in 10 mM Tris (pH 8.3, at 42°C) and 250 mM KCl were mixed with primer cirRTpro1 , heated for 3 min to 95°C, incubated for 10 min at 65°C, and then held on ice.

    Article Title: Primed CRISPR adaptation in Escherichia coli cells does not depend on conformational changes in the Cascade effector complex detected in Vitro
    Article Snippet: Constructs containing the WT g8 protospacer and its variants were cloned into plasmid pUC19 (NEB) at the single SmaI (NEB) site by blunt end ligation. .. Ligation products were transformed into NEB 5α E. coli cells.

    Sequencing:

    Article Title: A Robust System for Production of Superabundant VP1 Recombinant AAV Vectors
    Article Snippet: The plasmid pRB21 (with P7.5 promoter) was digested with SmaI (NEB) and used as the backbone to construct all plasmids having individual AAV and Ad genes. .. E1a-13S , E1b-55K , E1b-19k , E2a , E4orf6 , and VA I RNA were amplified using the pFΔ6 plasmid as a template.

    Article Title: Two forms of RNA editing are required for tRNA maturation in Physarum mitochondria
    Article Snippet: The resulting RT-PCR products were gel purified and ligated to pBSM13+ (Stratagene) that had been digested with SmaI (New England Biolabs) and treated with calf intestine alkaline phosphatase (Roche). .. One-tenth of the cDNA product was used in a PCR reaction using primers 1ssu (5′-TCACGTACAGACCGCCC-3′) and tRNAK3 (5′-TGGTTGGCTCCACAGGACTTGC-3′) and Taq polymerase (New England BioLabs) under recommended conditions.

    Article Title: Primed CRISPR adaptation in Escherichia coli cells does not depend on conformational changes in the Cascade effector complex detected in Vitro
    Article Snippet: Constructs containing the WT g8 protospacer and its variants were cloned into plasmid pUC19 (NEB) at the single SmaI (NEB) site by blunt end ligation. .. Ligation products were transformed into NEB 5α E. coli cells.

    Article Title: LSm1-7 complexes bind to specific sites in viral RNA genomes and regulate their translation and replication
    Article Snippet: Forward primers carried a T7 promoter sequence at their 5′ end. .. Amplified fragments were cloned into SmaI digested pUC18 (New England Biolabs) or pGEM-T vectors (Promega).

    Labeling:

    Article Title: Molecular quantification and differentiation of Candida species in biological specimens of patients with liver cirrhosis
    Article Snippet: After in silico enzyme digestion of all the standard strains using REPK Version 1.3 [ ], a combination of five enzymes led to the best possible differentiation of the above pathogens: Hpy188I, BfaI, HaeIII, BglI and SmaI (New England Biolabs, Frankfurt, Germany). .. After in silico enzyme digestion of all the standard strains using REPK Version 1.3 [ ], a combination of five enzymes led to the best possible differentiation of the above pathogens: Hpy188I, BfaI, HaeIII, BglI and SmaI (New England Biolabs, Frankfurt, Germany).

    Activated Clotting Time Assay:

    Article Title: Genome-wide alterations in gene methylation by the BRAF V600E mutation in papillary thyroid cancer cells
    Article Snippet: Briefly, ~5 µg of each genomic DNA from cells stably transfected with BRAF shRNA (termed ‘test’ hereafter) and vector (termed ‘control’ hereafter) were digested with 100 units of methylation-sensitive restriction endonuclease SmaI (New England Biolabs, Ipswich, MA, USA) for 16 h at 20 °C in a total volume of 100 µl, followed by digestion with 20 units of methylation-insensitive restriction endonuclease XmaI for 6 h at 37 °C, which leaves sticky ends (C/CCGGG). .. Briefly, ~5 µg of each genomic DNA from cells stably transfected with BRAF shRNA (termed ‘test’ hereafter) and vector (termed ‘control’ hereafter) were digested with 100 units of methylation-sensitive restriction endonuclease SmaI (New England Biolabs, Ipswich, MA, USA) for 16 h at 20 °C in a total volume of 100 µl, followed by digestion with 20 units of methylation-insensitive restriction endonuclease XmaI for 6 h at 37 °C, which leaves sticky ends (C/CCGGG).

    Terminal Restriction Fragment Length Polymorphism:

    Article Title: Molecular quantification and differentiation of Candida species in biological specimens of patients with liver cirrhosis
    Article Snippet: Paragraph title: T-RFLP analysis for the differentiation of Candida DNA ... After in silico enzyme digestion of all the standard strains using REPK Version 1.3 [ ], a combination of five enzymes led to the best possible differentiation of the above pathogens: Hpy188I, BfaI, HaeIII, BglI and SmaI (New England Biolabs, Frankfurt, Germany).

    Software:

    Article Title: Molecular quantification and differentiation of Candida species in biological specimens of patients with liver cirrhosis
    Article Snippet: After in silico enzyme digestion of all the standard strains using REPK Version 1.3 [ ], a combination of five enzymes led to the best possible differentiation of the above pathogens: Hpy188I, BfaI, HaeIII, BglI and SmaI (New England Biolabs, Frankfurt, Germany). .. Using the ABI Prism 3100 genetic analyzer (Applied Biosystems), terminally labeled 18S rRNA gene fragments were separated using capillary electrophoresis.

    Article Title: Novel Structure of Enterococcus faecium-Originated ermB-Positive Tn1546-Like Element in Staphylococcus aureus
    Article Snippet: The DNA in gel plugs was digested with SmaI (New England BioLabs, Ipswich, MA) and then separated in a CHEF-DR III apparatus (Bio-Rad Laboratories). .. The DNA in gel plugs was digested with SmaI (New England BioLabs, Ipswich, MA) and then separated in a CHEF-DR III apparatus (Bio-Rad Laboratories).

    Article Title: Comparison of an Automated Repetitive Sequence-Based PCR Microbial Typing System to Pulsed-Field Gel Electrophoresis for Analysis of Outbreaks of Methicillin-Resistant Staphylococcus aureus
    Article Snippet: DNA was extracted by standard PFGE methods ( ) and digested with SmaI (New England Biolabs, Beverly, MA). .. Electrophoresis was performed using a GenePath system (Bio-Rad Laboratories, Hercules, CA) with 1% Bio-Rad PFGE-certified agarose-0.5× Tris-boric acid-EDTA running buffer at 6 V for 23.5 h at 14°C and involved ramping from 5 to 50 s as the initial and final switch times.

    Electrophoresis:

    Article Title: Interference in Pheromone-Responsive Conjugation of a High-Level Bacitracin Resistant Enterococcus faecalis Plasmid of Poultry Origin
    Article Snippet: Paragraph title: 2.9. Pulsed-Field Gel Electrophoresis (PFGE) ... Briefly, a plug slice of 2 to 4 mm wide was suspended in a total volume of 227 µL of the manufacturer’s recommended restriction buffer and 20 U of SmaI (New England Biolabs Inc., Mississauga, ON, Canada).

    Article Title: Molecular quantification and differentiation of Candida species in biological specimens of patients with liver cirrhosis
    Article Snippet: After in silico enzyme digestion of all the standard strains using REPK Version 1.3 [ ], a combination of five enzymes led to the best possible differentiation of the above pathogens: Hpy188I, BfaI, HaeIII, BglI and SmaI (New England Biolabs, Frankfurt, Germany). .. After in silico enzyme digestion of all the standard strains using REPK Version 1.3 [ ], a combination of five enzymes led to the best possible differentiation of the above pathogens: Hpy188I, BfaI, HaeIII, BglI and SmaI (New England Biolabs, Frankfurt, Germany).

    Article Title: Multilocus Sequence Typing, Pulsed-Field Gel Electrophoresis, and fla Short Variable Region Typing of Clonal Complexes of Campylobacter jejuni Strains of Human, Bovine, and Poultry Origins in Luxembourg
    Article Snippet: DNA was digested overnight at 25°C with 25 U of SmaI restriction endonuclease (New England Biolabs, Westburg, The Netherlands) in a final volume of 130 μl. .. DNA was digested overnight at 25°C with 25 U of SmaI restriction endonuclease (New England Biolabs, Westburg, The Netherlands) in a final volume of 130 μl.

    shRNA:

    Article Title: Genome-wide alterations in gene methylation by the BRAF V600E mutation in papillary thyroid cancer cells
    Article Snippet: The MCA assay was performed as described previously ( ). .. Briefly, ~5 µg of each genomic DNA from cells stably transfected with BRAF shRNA (termed ‘test’ hereafter) and vector (termed ‘control’ hereafter) were digested with 100 units of methylation-sensitive restriction endonuclease SmaI (New England Biolabs, Ipswich, MA, USA) for 16 h at 20 °C in a total volume of 100 µl, followed by digestion with 20 units of methylation-insensitive restriction endonuclease XmaI for 6 h at 37 °C, which leaves sticky ends (C/CCGGG). .. DNA fragments were then precipitated with ethanol and their sticky ends ligated with 0.5 nmol of unphosphorylated linkers RXMA −24/−12, as described ( ).

    Agarose Gel Electrophoresis:

    Article Title: Altered Growth, Pigmentation, and Antimicrobial Susceptibility Properties of Staphylococcusaureus Due to Loss of the Major Cold Shock Gene cspB
    Article Snippet: Agarose plugs containing cells were prepared from 175 μl of overnight cultures of each strain grown in brain heart infusion broth (Becton Dickinson Microbiology Systems, Cockeysville, MD) and digested with SmaI (New England Biolabs, Ipswich, MA). .. Agarose plugs containing cells were prepared from 175 μl of overnight cultures of each strain grown in brain heart infusion broth (Becton Dickinson Microbiology Systems, Cockeysville, MD) and digested with SmaI (New England Biolabs, Ipswich, MA).

    Article Title: High-throughput methylation profiling by MCA coupled to CpG island microarray
    Article Snippet: In brief, 5 μg of DNA was digested with 100 units of SmaI for 16 h (all restriction enzymes were from New England Biolabs), followed by digestion with 20 units of XmaI for 6 h. DNA fragments were then precipitated with ethanol. .. In brief, 5 μg of DNA was digested with 100 units of SmaI for 16 h (all restriction enzymes were from New England Biolabs), followed by digestion with 20 units of XmaI for 6 h. DNA fragments were then precipitated with ethanol.

    Article Title: Multilocus Sequence Typing, Pulsed-Field Gel Electrophoresis, and fla Short Variable Region Typing of Clonal Complexes of Campylobacter jejuni Strains of Human, Bovine, and Poultry Origins in Luxembourg
    Article Snippet: DNA was digested overnight at 25°C with 25 U of SmaI restriction endonuclease (New England Biolabs, Westburg, The Netherlands) in a final volume of 130 μl. .. DNA was digested overnight at 25°C with 25 U of SmaI restriction endonuclease (New England Biolabs, Westburg, The Netherlands) in a final volume of 130 μl.

    Article Title: Genome-wide alterations in gene methylation by the BRAF V600E mutation in papillary thyroid cancer cells
    Article Snippet: Briefly, ~5 µg of each genomic DNA from cells stably transfected with BRAF shRNA (termed ‘test’ hereafter) and vector (termed ‘control’ hereafter) were digested with 100 units of methylation-sensitive restriction endonuclease SmaI (New England Biolabs, Ipswich, MA, USA) for 16 h at 20 °C in a total volume of 100 µl, followed by digestion with 20 units of methylation-insensitive restriction endonuclease XmaI for 6 h at 37 °C, which leaves sticky ends (C/CCGGG). .. PCR reactions were performed using the digests as templates and subjected to 20 cycles of amplification as described previously ( ).

    In Vitro:

    Article Title: Immune Mediators Regulate CFTR Expression through a Bifunctional Airway-Selective Enhancer
    Article Snippet: Paragraph title: In vitro DNase I footprinting. ... Cleavage of this construct with XhoI and SmaI (New England BioLabs, Ipswich, MA) enabled Klenow DNA polymerase (New England BioLabs) fill-in with [32 P]dCTP.

    Spectrophotometry:

    Article Title: Molecular quantification and differentiation of Candida species in biological specimens of patients with liver cirrhosis
    Article Snippet: The purification of PCR products was performed, as described above, and the DNA content of each PCR product was quantified using the Nanodrop ND-1000 spectrophotometer (Thermo Fisher Scientific). .. After in silico enzyme digestion of all the standard strains using REPK Version 1.3 [ ], a combination of five enzymes led to the best possible differentiation of the above pathogens: Hpy188I, BfaI, HaeIII, BglI and SmaI (New England Biolabs, Frankfurt, Germany).

    Marker:

    Article Title: In Vivo Transfer of the vanA Resistance Gene from an Enterococcus faecium Isolate of Animal Origin to an E. faecium Isolate of Human Origin in the Intestines of Human Volunteers
    Article Snippet: A small slice of the agarose plug was digested with 30 U of the restriction enzyme SmaI (New England BioLabs Inc., Medinova Scientific A/S, Denmark) for a minimum of 4 h at 25°C. .. A small slice of the agarose plug was digested with 30 U of the restriction enzyme SmaI (New England BioLabs Inc., Medinova Scientific A/S, Denmark) for a minimum of 4 h at 25°C.

    Staining:

    Article Title: Comparison of an Automated Repetitive Sequence-Based PCR Microbial Typing System to Pulsed-Field Gel Electrophoresis for Analysis of Outbreaks of Methicillin-Resistant Staphylococcus aureus
    Article Snippet: DNA was extracted by standard PFGE methods ( ) and digested with SmaI (New England Biolabs, Beverly, MA). .. Electrophoresis was performed using a GenePath system (Bio-Rad Laboratories, Hercules, CA) with 1% Bio-Rad PFGE-certified agarose-0.5× Tris-boric acid-EDTA running buffer at 6 V for 23.5 h at 14°C and involved ramping from 5 to 50 s as the initial and final switch times.

    Article Title: High-throughput methylation profiling by MCA coupled to CpG island microarray
    Article Snippet: In brief, 5 μg of DNA was digested with 100 units of SmaI for 16 h (all restriction enzymes were from New England Biolabs), followed by digestion with 20 units of XmaI for 6 h. DNA fragments were then precipitated with ethanol. .. In brief, 5 μg of DNA was digested with 100 units of SmaI for 16 h (all restriction enzymes were from New England Biolabs), followed by digestion with 20 units of XmaI for 6 h. DNA fragments were then precipitated with ethanol.

    Article Title: Oxacilin-resistant Coagulase-negative staphylococci (CoNS) bacteremia in a general hospital at S?o Paulo city, Brasil
    Article Snippet: Chromosomal DNA CoNS was prepared in agarose blocks and digested with SmaI (New England BioLabs, USA), as described elsewhere ( ). .. The isolates were run on a 1% agarose gel (Invitrogen, USA) in a CHEF DRIII system (Bio-Rad, USA) under the following conditions: run time, 23 h; temperature, 13°C; voltage, 200 V; initial forward time, 5 s; final forward time, 60 s. The molecular weight markers (New England BioLabs, USA) were run in the first and in the last lane.

    Article Title: Genome-wide alterations in gene methylation by the BRAF V600E mutation in papillary thyroid cancer cells
    Article Snippet: Briefly, ~5 µg of each genomic DNA from cells stably transfected with BRAF shRNA (termed ‘test’ hereafter) and vector (termed ‘control’ hereafter) were digested with 100 units of methylation-sensitive restriction endonuclease SmaI (New England Biolabs, Ipswich, MA, USA) for 16 h at 20 °C in a total volume of 100 µl, followed by digestion with 20 units of methylation-insensitive restriction endonuclease XmaI for 6 h at 37 °C, which leaves sticky ends (C/CCGGG). .. PCR reactions were performed using the digests as templates and subjected to 20 cycles of amplification as described previously ( ).

    Variant Assay:

    Article Title: Primed CRISPR adaptation in Escherichia coli cells does not depend on conformational changes in the Cascade effector complex detected in Vitro
    Article Snippet: Constructs containing the WT g8 protospacer and its variants were cloned into plasmid pUC19 (NEB) at the single SmaI (NEB) site by blunt end ligation. .. Ligation products were transformed into NEB 5α E. coli cells.

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    New England Biolabs smai
    <t>PFGE-based</t> clustering of C. coli strains investigated in this study. All 68 strains were analyzed by PFGE with <t>SmaI</t> and KpnI, and cluster analysis of the patterns was performed by BioNumerics as described in Materials and Methods. Two strains (1004 A
    Smai, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 73 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    PFGE-based clustering of C. coli strains investigated in this study. All 68 strains were analyzed by PFGE with SmaI and KpnI, and cluster analysis of the patterns was performed by BioNumerics as described in Materials and Methods. Two strains (1004 A

    Journal:

    Article Title: Clonal Population Structure and Specific Genotypes of Multidrug-Resistant Campylobacter coli from Turkeys

    doi: 10.1128/AEM.02346-06

    Figure Lengend Snippet: PFGE-based clustering of C. coli strains investigated in this study. All 68 strains were analyzed by PFGE with SmaI and KpnI, and cluster analysis of the patterns was performed by BioNumerics as described in Materials and Methods. Two strains (1004 A

    Article Snippet: PFGE was performed using SmaI and KpnI (New England Biolabs) by following the protocol described by Ribot et al. ( ) with a few minor modifications; specifically, the prerestriction step was eliminated and the gels were electrophoresed for 22 h. TIFF images of banding patterns resulting from PFGE and fla typing were analyzed by BioNumerics (version 4.6; Applied Maths).

    Techniques:

    PFGE profiles of group I strains. Nineteen MDR strains were characterized by PFGE using SmaI and KpnI, and cluster analysis of the patterns was performed by BioNumerics as described in Materials and Methods. The line labeled “a” indicates

    Journal:

    Article Title: Clonal Population Structure and Specific Genotypes of Multidrug-Resistant Campylobacter coli from Turkeys

    doi: 10.1128/AEM.02346-06

    Figure Lengend Snippet: PFGE profiles of group I strains. Nineteen MDR strains were characterized by PFGE using SmaI and KpnI, and cluster analysis of the patterns was performed by BioNumerics as described in Materials and Methods. The line labeled “a” indicates

    Article Snippet: PFGE was performed using SmaI and KpnI (New England Biolabs) by following the protocol described by Ribot et al. ( ) with a few minor modifications; specifically, the prerestriction step was eliminated and the gels were electrophoresed for 22 h. TIFF images of banding patterns resulting from PFGE and fla typing were analyzed by BioNumerics (version 4.6; Applied Maths).

    Techniques: Labeling

    A conserved SmaI PFGE pattern (type a ) was disseminated among MDR strains of different STs.

    Journal:

    Article Title: Clonal Population Structure and Specific Genotypes of Multidrug-Resistant Campylobacter coli from Turkeys

    doi: 10.1128/AEM.02346-06

    Figure Lengend Snippet: A conserved SmaI PFGE pattern (type a ) was disseminated among MDR strains of different STs.

    Article Snippet: PFGE was performed using SmaI and KpnI (New England Biolabs) by following the protocol described by Ribot et al. ( ) with a few minor modifications; specifically, the prerestriction step was eliminated and the gels were electrophoresed for 22 h. TIFF images of banding patterns resulting from PFGE and fla typing were analyzed by BioNumerics (version 4.6; Applied Maths).

    Techniques:

    PFGE profiles of group II strains. Forty-two strains were characterized by PFGE using SmaI and KpnI, and cluster analysis of the patterns was performed by BioNumerics as described in Materials and Methods. Strains which were not resistant to all six antibiotics

    Journal:

    Article Title: Clonal Population Structure and Specific Genotypes of Multidrug-Resistant Campylobacter coli from Turkeys

    doi: 10.1128/AEM.02346-06

    Figure Lengend Snippet: PFGE profiles of group II strains. Forty-two strains were characterized by PFGE using SmaI and KpnI, and cluster analysis of the patterns was performed by BioNumerics as described in Materials and Methods. Strains which were not resistant to all six antibiotics

    Article Snippet: PFGE was performed using SmaI and KpnI (New England Biolabs) by following the protocol described by Ribot et al. ( ) with a few minor modifications; specifically, the prerestriction step was eliminated and the gels were electrophoresed for 22 h. TIFF images of banding patterns resulting from PFGE and fla typing were analyzed by BioNumerics (version 4.6; Applied Maths).

    Techniques:

    PFGE profiles of 13 strains with various STs but a conserved PFGE profile (type a ). Cluster analysis of the SmaI and KpnI patterns was performed by BioNumerics as described in Materials and Methods.

    Journal:

    Article Title: Clonal Population Structure and Specific Genotypes of Multidrug-Resistant Campylobacter coli from Turkeys

    doi: 10.1128/AEM.02346-06

    Figure Lengend Snippet: PFGE profiles of 13 strains with various STs but a conserved PFGE profile (type a ). Cluster analysis of the SmaI and KpnI patterns was performed by BioNumerics as described in Materials and Methods.

    Article Snippet: PFGE was performed using SmaI and KpnI (New England Biolabs) by following the protocol described by Ribot et al. ( ) with a few minor modifications; specifically, the prerestriction step was eliminated and the gels were electrophoresed for 22 h. TIFF images of banding patterns resulting from PFGE and fla typing were analyzed by BioNumerics (version 4.6; Applied Maths).

    Techniques:

    Pulsed-field gel electrophoresis (PFGE)-based dendrogram of 13 SmaI digested S. canis genomic DNA from healthy and diseased mink from 2 unrelated farms in Canada and clinical infections in 4 unrelated dogs. Scale bar represents percent similarity based on Dice coefficients and UPGMA clustering.

    Journal:

    Article Title: Staphylococcus spp., Streptococcus canis, and Arcanobacterium phocae of healthy Canadian farmed mink and mink with pododermatitis

    doi:

    Figure Lengend Snippet: Pulsed-field gel electrophoresis (PFGE)-based dendrogram of 13 SmaI digested S. canis genomic DNA from healthy and diseased mink from 2 unrelated farms in Canada and clinical infections in 4 unrelated dogs. Scale bar represents percent similarity based on Dice coefficients and UPGMA clustering.

    Article Snippet: Genomic DNA from both species was digested with SmaI (New England BioLabs, Ipswich, Massachusetts, USA) before electrophoresis in a 1.0% agarose gel, using a CHEF-III electrophoresis unit (Bio-Rad Laboratories, Hercules, California, USA) in 0.5× Trisborate-ethylenediamine tetra-acetic acid (EDTA) buffer containing 200 μM thiourea (Fisher Scientific, Fair Lawn, New Jersey, USA).

    Techniques: Pulsed-Field Gel, Electrophoresis

    Pulsed-field gel electrophoresis (PFGE)-based dendrogram of 21 SmaI digested S. delphini genomic DNA from healthy and diseased mink from 2 unrelated farms in Canada. Scale bar represents percent similarity based on Dice coefficients and UPGMA clustering. Asterisk (*) indicates 2 isolates taken from the same individual mink.

    Journal:

    Article Title: Staphylococcus spp., Streptococcus canis, and Arcanobacterium phocae of healthy Canadian farmed mink and mink with pododermatitis

    doi:

    Figure Lengend Snippet: Pulsed-field gel electrophoresis (PFGE)-based dendrogram of 21 SmaI digested S. delphini genomic DNA from healthy and diseased mink from 2 unrelated farms in Canada. Scale bar represents percent similarity based on Dice coefficients and UPGMA clustering. Asterisk (*) indicates 2 isolates taken from the same individual mink.

    Article Snippet: Genomic DNA from both species was digested with SmaI (New England BioLabs, Ipswich, Massachusetts, USA) before electrophoresis in a 1.0% agarose gel, using a CHEF-III electrophoresis unit (Bio-Rad Laboratories, Hercules, California, USA) in 0.5× Trisborate-ethylenediamine tetra-acetic acid (EDTA) buffer containing 200 μM thiourea (Fisher Scientific, Fair Lawn, New Jersey, USA).

    Techniques: Pulsed-Field Gel, Electrophoresis

    Generation of aspa lacZ mice by homologous recombination. A, Genomic structure of the murine aspa locus, which spans 6 exons (black boxes). Homologous recombination of the targeting vector inserts the βgeo cassette (blue box), encoding β-Galactosidase (lacZ), into intron 1 of the intact aspa gene. This cassette is flanked by frt-sites (green circles). Additional loxP sites flank exon 2 (red triangles). The vector includes a DTA cassette for negative selection. In the targeted allele, exon 1 is spliced to the splice acceptor site preceding βgeo, and transcription is terminated at the introduced pA site. The conditional aspa flox allele is produced by breeding aspa lacZ mutants to FLP-deleter mice [40] for recombination of the βgeo cassette in vivo . Note that frt and loxP sites are not drawn to scale. βgeo, β-galactosidase-neomycin resistance cassette; SA, splice acceptor; pA, polyadenylation site, DTA, Diphteria toxin gene. B, For Southern blot analysis genomic DNA was digested with SmaI/ApaI. The neo probe (white box in A) detected the expected 7.5 kb band in heterozygotes and homozygotes. C, Genomic PCR of littermates with primers 1 3 produces the expected amplicons in aspa +/+ (336 bp), aspa lacZ/+ (387 bp and 336 bp) and aspa lacZ/lacZ (387 bp) animals. The upstream loxP site was detected by PCR with primers 1 2 yielding a 307 bp band. D, Q-RT-PCR using a TaqMan probe for quantification of aspa mRNA levels in brains of aspa +/+ , aspa lacZ/+ , and aspa lacZ/lacZ mice (n = 3) confirms the attenuation of transcription downstream of exon 2 in the targeted allele. E, Representative Western blot of whole brain lysates of a spa +/+ , aspa lacZ/+ and aspa lacZ/lacZ mice (aged 4 months, n = 3). The 37 kD protein ASPA was detected in aspa +/+ and aspa lacZ/+ mice but not in aspa lacZ/lacZ mutants. β-Galactosidase is expressed in aspa lacZ/+ and aspa lacZ/lacZ brain but not controls. α-Tubulin was used as loading control. F. Representative picture of a male aspa lacZ/lacZ mutant (asterisk) and an aspa +/+ littermate at P70.

    Journal: PLoS ONE

    Article Title: Aspartoacylase-LacZ Knockin Mice: An Engineered Model of Canavan Disease

    doi: 10.1371/journal.pone.0020336

    Figure Lengend Snippet: Generation of aspa lacZ mice by homologous recombination. A, Genomic structure of the murine aspa locus, which spans 6 exons (black boxes). Homologous recombination of the targeting vector inserts the βgeo cassette (blue box), encoding β-Galactosidase (lacZ), into intron 1 of the intact aspa gene. This cassette is flanked by frt-sites (green circles). Additional loxP sites flank exon 2 (red triangles). The vector includes a DTA cassette for negative selection. In the targeted allele, exon 1 is spliced to the splice acceptor site preceding βgeo, and transcription is terminated at the introduced pA site. The conditional aspa flox allele is produced by breeding aspa lacZ mutants to FLP-deleter mice [40] for recombination of the βgeo cassette in vivo . Note that frt and loxP sites are not drawn to scale. βgeo, β-galactosidase-neomycin resistance cassette; SA, splice acceptor; pA, polyadenylation site, DTA, Diphteria toxin gene. B, For Southern blot analysis genomic DNA was digested with SmaI/ApaI. The neo probe (white box in A) detected the expected 7.5 kb band in heterozygotes and homozygotes. C, Genomic PCR of littermates with primers 1 3 produces the expected amplicons in aspa +/+ (336 bp), aspa lacZ/+ (387 bp and 336 bp) and aspa lacZ/lacZ (387 bp) animals. The upstream loxP site was detected by PCR with primers 1 2 yielding a 307 bp band. D, Q-RT-PCR using a TaqMan probe for quantification of aspa mRNA levels in brains of aspa +/+ , aspa lacZ/+ , and aspa lacZ/lacZ mice (n = 3) confirms the attenuation of transcription downstream of exon 2 in the targeted allele. E, Representative Western blot of whole brain lysates of a spa +/+ , aspa lacZ/+ and aspa lacZ/lacZ mice (aged 4 months, n = 3). The 37 kD protein ASPA was detected in aspa +/+ and aspa lacZ/+ mice but not in aspa lacZ/lacZ mutants. β-Galactosidase is expressed in aspa lacZ/+ and aspa lacZ/lacZ brain but not controls. α-Tubulin was used as loading control. F. Representative picture of a male aspa lacZ/lacZ mutant (asterisk) and an aspa +/+ littermate at P70.

    Article Snippet: DNA was digested with ApaI and SmaI (NEB, Ipswich, MA), separated on a 0.7% agarose gel, blotted onto Hybond N+ membrane (GE Healthcare, Dassel, Germany) and probed with a 600 bp 32 P-labelled fragment of the Neo-cassette.

    Techniques: Mouse Assay, Homologous Recombination, Plasmid Preparation, Selection, Produced, In Vivo, Southern Blot, Polymerase Chain Reaction, Reverse Transcription Polymerase Chain Reaction, Western Blot, Mutagenesis

    Engineering a full-length JHMV infectious BAC. (A) Strategy used to construct the JHMV BAC. First, a synthetic DNA was designed containing a CMV promoter, 1,991 bp of 5′ JHM sequence, and 1,197 bp of 3′ JHM sequence separated by a NotI site, and finally a poly(A) tail (pA), ribozyme cleavage site, HDV ribozyme, and BGH termination signal at the end. This DNA was then digested with SmaI and HindIII and ligated into the parental BAC plasmid, and the resulting clone was termed pBAC-JHMV 5′3′. Subsequently, pBAC-JHMV 5′3′ and a full-length JHMV ligated in vitro were digested with PacI and SanDI, ligated, and transformed into competent DH10B E. coli cells. The resulting BAC was termed pBAC-JHMV. (B) Confirmation of BAC clones. Representative clones of pBAC-JHMVIA and pBAC-JHMVSD were digested with EcoRI and EcoRV. The expected banding pattern for EcoRI digestion is 14.5, 11, 7.7, and 6.3 kb, while the expected pattern from EcoRV digestion is 22.3, 12.6, 4.7, and 0.2 kb. The resulting digests matched the expected digestion patterns (0.2-kb fragment is undetectable on the gel).

    Journal:

    Article Title: The nsp3 Macrodomain Promotes Virulence in Mice with Coronavirus-Induced Encephalitis

    doi: 10.1128/JVI.02596-14

    Figure Lengend Snippet: Engineering a full-length JHMV infectious BAC. (A) Strategy used to construct the JHMV BAC. First, a synthetic DNA was designed containing a CMV promoter, 1,991 bp of 5′ JHM sequence, and 1,197 bp of 3′ JHM sequence separated by a NotI site, and finally a poly(A) tail (pA), ribozyme cleavage site, HDV ribozyme, and BGH termination signal at the end. This DNA was then digested with SmaI and HindIII and ligated into the parental BAC plasmid, and the resulting clone was termed pBAC-JHMV 5′3′. Subsequently, pBAC-JHMV 5′3′ and a full-length JHMV ligated in vitro were digested with PacI and SanDI, ligated, and transformed into competent DH10B E. coli cells. The resulting BAC was termed pBAC-JHMV. (B) Confirmation of BAC clones. Representative clones of pBAC-JHMVIA and pBAC-JHMVSD were digested with EcoRI and EcoRV. The expected banding pattern for EcoRI digestion is 14.5, 11, 7.7, and 6.3 kb, while the expected pattern from EcoRV digestion is 22.3, 12.6, 4.7, and 0.2 kb. The resulting digests matched the expected digestion patterns (0.2-kb fragment is undetectable on the gel).

    Article Snippet: The pUC57 vector containing this synthetic DNA was digested with SmaI and HindIII while pBeloBAC11 (New England BioLabs) was digested with SfoI and HindIII.

    Techniques: BAC Assay, Construct, Sequencing, Plasmid Preparation, In Vitro, Transformation Assay, Clone Assay