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  • 99
    New England Biolabs factor xa protease
    Factor Xa Protease, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 624 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    rocky mountain labs pbvs2 himar1 c9
    Plasmid maps of pCR2.1-P1196- <t>Himar1</t> C9 and p1898-Tn. (A) In pCR2.1-P1196- Himar1 C9, the promoter from CBU1169 (P1169) drives expression of the C9 variant of the Himar1 transposase. (B) In p1898-Tn, the Himar1 transposon contains CAT and mCherry genes expressed as a single transcriptional unit from P1169. Also contained within the Himar1 ITRs is a ColE1 origin of replication that allows rescue cloning of the Tn in E. coli . Outside of the transposon is the coding sequence of CBU1898.
    Pbvs2 Himar1 C9, supplied by rocky mountain labs, used in various techniques. Bioz Stars score: 85/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Illumina Inc himar1 mariner transposons
    Plasmid maps of pCR2.1-P1196- <t>Himar1</t> C9 and p1898-Tn. (A) In pCR2.1-P1196- Himar1 C9, the promoter from CBU1169 (P1169) drives expression of the C9 variant of the Himar1 transposase. (B) In p1898-Tn, the Himar1 transposon contains CAT and mCherry genes expressed as a single transcriptional unit from P1169. Also contained within the Himar1 ITRs is a ColE1 origin of replication that allows rescue cloning of the Tn in E. coli . Outside of the transposon is the coding sequence of CBU1898.
    Himar1 Mariner Transposons, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 85/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    Roche himar1 insertion site involved alignment
    Immunoblotting of <t>omp10::himar1</t> mutant and WT A. marginale using the specific monoclonal antibody Omp9. Proteins from equal amounts of host cell-free wild-type (WT) and omp10::himar1 A. marginale were separated by SDS-PAGE gel electrophoresis. Immunoblot PVDF membranes of transferred proteins were reacted with monoclonal antibodies and reactions were visualized by chemiluminescence. A . Monoclonal antibody Omp9 (4 μg/ml) with specificity to Omp9 protein (40 kDa) (black arrow). B . Negative control, monoclonal Tryp1E1 (4 μg/ml) (exhibits specificity for a variable surface glycoprotein of Trypanosoma brucei . C . Monoclonal F16C1 (2 μg/ml), reacts with the Msp5 (19 kDa) (blue arrow) protein of A. marginale , was used as loading control. A. marginale str. Virginia and uninfected ISE6 cells were used as positive and negative controls respectively.
    Himar1 Insertion Site Involved Alignment, supplied by Roche, used in various techniques. Bioz Stars score: 86/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Plasmid maps of pCR2.1-P1196- Himar1 C9 and p1898-Tn. (A) In pCR2.1-P1196- Himar1 C9, the promoter from CBU1169 (P1169) drives expression of the C9 variant of the Himar1 transposase. (B) In p1898-Tn, the Himar1 transposon contains CAT and mCherry genes expressed as a single transcriptional unit from P1169. Also contained within the Himar1 ITRs is a ColE1 origin of replication that allows rescue cloning of the Tn in E. coli . Outside of the transposon is the coding sequence of CBU1898.

    Journal: Journal of Bacteriology

    Article Title: Characterization of a Coxiella burnetii ftsZ Mutant Generated by Himar1 Transposon Mutagenesis ▿ Transposon Mutagenesis ▿ †

    doi: 10.1128/JB.01580-08

    Figure Lengend Snippet: Plasmid maps of pCR2.1-P1196- Himar1 C9 and p1898-Tn. (A) In pCR2.1-P1196- Himar1 C9, the promoter from CBU1169 (P1169) drives expression of the C9 variant of the Himar1 transposase. (B) In p1898-Tn, the Himar1 transposon contains CAT and mCherry genes expressed as a single transcriptional unit from P1169. Also contained within the Himar1 ITRs is a ColE1 origin of replication that allows rescue cloning of the Tn in E. coli . Outside of the transposon is the coding sequence of CBU1898.

    Article Snippet: Himar1 C9 from pBVS2- Himar1 C9 (generously provided by P. Stewart, Rocky Mountain Laboratories) was amplified by using Accuprime Taq DNA polymerase and the primers Himar1 C9-NdeI-F and Himar1 C9-HindIII-R and then cloned into pCR2.1-Topo.

    Techniques: Plasmid Preparation, Expressing, Variant Assay, Clone Assay, Sequencing

    Immunoblotting of omp10::himar1 mutant and WT A. marginale using the specific monoclonal antibody Omp9. Proteins from equal amounts of host cell-free wild-type (WT) and omp10::himar1 A. marginale were separated by SDS-PAGE gel electrophoresis. Immunoblot PVDF membranes of transferred proteins were reacted with monoclonal antibodies and reactions were visualized by chemiluminescence. A . Monoclonal antibody Omp9 (4 μg/ml) with specificity to Omp9 protein (40 kDa) (black arrow). B . Negative control, monoclonal Tryp1E1 (4 μg/ml) (exhibits specificity for a variable surface glycoprotein of Trypanosoma brucei . C . Monoclonal F16C1 (2 μg/ml), reacts with the Msp5 (19 kDa) (blue arrow) protein of A. marginale , was used as loading control. A. marginale str. Virginia and uninfected ISE6 cells were used as positive and negative controls respectively.

    Journal: BMC Genomics

    Article Title: Knockout of an outer membrane protein operon of Anaplasma marginale by transposon mutagenesis

    doi: 10.1186/1471-2164-15-278

    Figure Lengend Snippet: Immunoblotting of omp10::himar1 mutant and WT A. marginale using the specific monoclonal antibody Omp9. Proteins from equal amounts of host cell-free wild-type (WT) and omp10::himar1 A. marginale were separated by SDS-PAGE gel electrophoresis. Immunoblot PVDF membranes of transferred proteins were reacted with monoclonal antibodies and reactions were visualized by chemiluminescence. A . Monoclonal antibody Omp9 (4 μg/ml) with specificity to Omp9 protein (40 kDa) (black arrow). B . Negative control, monoclonal Tryp1E1 (4 μg/ml) (exhibits specificity for a variable surface glycoprotein of Trypanosoma brucei . C . Monoclonal F16C1 (2 μg/ml), reacts with the Msp5 (19 kDa) (blue arrow) protein of A. marginale , was used as loading control. A. marginale str. Virginia and uninfected ISE6 cells were used as positive and negative controls respectively.

    Article Snippet: Based on this, the strategy that we used to map the Himar1 insertion site involved alignment of the sequencing reads obtained by Roche/454 and Illumina methods to two reference sequences, the A. marginale str.

    Techniques: Mutagenesis, SDS Page, Nucleic Acid Electrophoresis, Negative Control

    Relative gene expression by RT-qPCR. A . location of binding sites for primers and probes designed to target omp8 ( 1 . AB1591, 2 . AB1592, 3 . AB1593), omp9 ( 4 . AB1581, 5 . AB1582, 6 . AB1583), 3’ end of omp10 ( 7 . AB1569, 8 . AB1570, 9 . AB1571), and the 5’ end of omp10 ( 10 . AB1594, 11 . AB1595, 12 . AB1596). B . Bar lengths represent the percentage of expression of omp8 , omp9 , 3’ end of omp10 and 5’ end of omp10 in A. marginale wild-type (red bars) and omp10::himar1 mutant (blue bars). msp5 , rpoH and 16S rRNA were used as reference genes for data normalization. Changes in expression of these genes were calculated using the 2 -ΔΔCt method.* Significant differences (P

    Journal: BMC Genomics

    Article Title: Knockout of an outer membrane protein operon of Anaplasma marginale by transposon mutagenesis

    doi: 10.1186/1471-2164-15-278

    Figure Lengend Snippet: Relative gene expression by RT-qPCR. A . location of binding sites for primers and probes designed to target omp8 ( 1 . AB1591, 2 . AB1592, 3 . AB1593), omp9 ( 4 . AB1581, 5 . AB1582, 6 . AB1583), 3’ end of omp10 ( 7 . AB1569, 8 . AB1570, 9 . AB1571), and the 5’ end of omp10 ( 10 . AB1594, 11 . AB1595, 12 . AB1596). B . Bar lengths represent the percentage of expression of omp8 , omp9 , 3’ end of omp10 and 5’ end of omp10 in A. marginale wild-type (red bars) and omp10::himar1 mutant (blue bars). msp5 , rpoH and 16S rRNA were used as reference genes for data normalization. Changes in expression of these genes were calculated using the 2 -ΔΔCt method.* Significant differences (P

    Article Snippet: Based on this, the strategy that we used to map the Himar1 insertion site involved alignment of the sequencing reads obtained by Roche/454 and Illumina methods to two reference sequences, the A. marginale str.

    Techniques: Expressing, Quantitative RT-PCR, Binding Assay, Mutagenesis

    Mapping of Himar1 transposon insertion site. A . Location of primer pairs (AB1553-AB1554) and (AB1561-AB1562) designed to target the omp6 and omp10 genes respectively, in wild-type (WT) and transformed A. marginale . Based on sequencing results the Himar1 sequences are integrated in the chromosome of transformed A. marginale at nucleotide 245 after the first base of the omp10 start codon (arrows) and mCherry and aadA (Str/Spc resistant) genes are in the opposite orientation to omp10 . B . Agarose Gel electrophoresis. gDNA isolated from ISE6 tick cells infected with wild-type (WT) and transformed A. marginale , was used as template for PCR amplification with primers shown in A. (Lane 1) 100 bp/1Kb DNA ladder, omp6 amplicons in transformed (lane 2) and WT (lane3) A. marginale were of the same size 492 bp. The omp10 amplicon in transformed A. marginale (lane 4) was 2805 bp, while in wild type was 969 bp (lane 5).

    Journal: BMC Genomics

    Article Title: Knockout of an outer membrane protein operon of Anaplasma marginale by transposon mutagenesis

    doi: 10.1186/1471-2164-15-278

    Figure Lengend Snippet: Mapping of Himar1 transposon insertion site. A . Location of primer pairs (AB1553-AB1554) and (AB1561-AB1562) designed to target the omp6 and omp10 genes respectively, in wild-type (WT) and transformed A. marginale . Based on sequencing results the Himar1 sequences are integrated in the chromosome of transformed A. marginale at nucleotide 245 after the first base of the omp10 start codon (arrows) and mCherry and aadA (Str/Spc resistant) genes are in the opposite orientation to omp10 . B . Agarose Gel electrophoresis. gDNA isolated from ISE6 tick cells infected with wild-type (WT) and transformed A. marginale , was used as template for PCR amplification with primers shown in A. (Lane 1) 100 bp/1Kb DNA ladder, omp6 amplicons in transformed (lane 2) and WT (lane3) A. marginale were of the same size 492 bp. The omp10 amplicon in transformed A. marginale (lane 4) was 2805 bp, while in wild type was 969 bp (lane 5).

    Article Snippet: Based on this, the strategy that we used to map the Himar1 insertion site involved alignment of the sequencing reads obtained by Roche/454 and Illumina methods to two reference sequences, the A. marginale str.

    Techniques: Transformation Assay, Sequencing, Agarose Gel Electrophoresis, Isolation, Infection, Polymerase Chain Reaction, Amplification

    Himar1 transposon insertion site in the A. marginale str. Virginia genome. ACT (Artemis Comparison Tool) window showing alignment between the A. marginale genome (CP000030) used as reference, the A. marginale str. Virginia omp10::himar1 contig formed by Roche/454 and Illumina sequencing reads and the Himar1 transposon sequences. Alignment between the A. marginale str St Maries and the omp10::himar1 mutant shows that sequences flanking the transposon insertion site are highly similar sharing an identity of 99% (matching red band). This demonstrates that the transposon insertion site (yellow boxes) occurred at nucleotide 245 after the first base of the omp10 start codon (black dotted line) in the reference strain. Alignment with the Himar1 transposon sequences clearly show the insertion of these sequences in the omp10::himar1 mutant which are not present in the A. marginale str . St Maries (absence of matching band).

    Journal: BMC Genomics

    Article Title: Knockout of an outer membrane protein operon of Anaplasma marginale by transposon mutagenesis

    doi: 10.1186/1471-2164-15-278

    Figure Lengend Snippet: Himar1 transposon insertion site in the A. marginale str. Virginia genome. ACT (Artemis Comparison Tool) window showing alignment between the A. marginale genome (CP000030) used as reference, the A. marginale str. Virginia omp10::himar1 contig formed by Roche/454 and Illumina sequencing reads and the Himar1 transposon sequences. Alignment between the A. marginale str St Maries and the omp10::himar1 mutant shows that sequences flanking the transposon insertion site are highly similar sharing an identity of 99% (matching red band). This demonstrates that the transposon insertion site (yellow boxes) occurred at nucleotide 245 after the first base of the omp10 start codon (black dotted line) in the reference strain. Alignment with the Himar1 transposon sequences clearly show the insertion of these sequences in the omp10::himar1 mutant which are not present in the A. marginale str . St Maries (absence of matching band).

    Article Snippet: Based on this, the strategy that we used to map the Himar1 insertion site involved alignment of the sequencing reads obtained by Roche/454 and Illumina methods to two reference sequences, the A. marginale str.

    Techniques: Activated Clotting Time Assay, Sequencing, Mutagenesis

    Characterization of Himar1 transposon insertion site. A . Artemis (genome browser and annotation tool) window showing the A. marginale genome (CP000030) used as a reference for the location of the omp10 gene ( AM1223 , 1092273–1093555), and the TA dinucleotide (1093290–1093291) at the Himar1 tn insertion site (arrow) determined by high throughput genome sequencing analysis. B . Himar1 tn insertion into the omp10 gene was mediated by the A7 transposase in a cut and paste mechanism leading to the duplication of TA dinucleotide sequences. A. marginale genome (underlined uppercase text, TA dinucleotide duplications (enhanced uppercase text) flanking the tn elements (bold lowercase).

    Journal: BMC Genomics

    Article Title: Knockout of an outer membrane protein operon of Anaplasma marginale by transposon mutagenesis

    doi: 10.1186/1471-2164-15-278

    Figure Lengend Snippet: Characterization of Himar1 transposon insertion site. A . Artemis (genome browser and annotation tool) window showing the A. marginale genome (CP000030) used as a reference for the location of the omp10 gene ( AM1223 , 1092273–1093555), and the TA dinucleotide (1093290–1093291) at the Himar1 tn insertion site (arrow) determined by high throughput genome sequencing analysis. B . Himar1 tn insertion into the omp10 gene was mediated by the A7 transposase in a cut and paste mechanism leading to the duplication of TA dinucleotide sequences. A. marginale genome (underlined uppercase text, TA dinucleotide duplications (enhanced uppercase text) flanking the tn elements (bold lowercase).

    Article Snippet: Based on this, the strategy that we used to map the Himar1 insertion site involved alignment of the sequencing reads obtained by Roche/454 and Illumina methods to two reference sequences, the A. marginale str.

    Techniques: High Throughput Screening Assay, Sequencing

    Transcriptional analysis of the effect of the insertion of the Himar1 transposon within the omp10 gene by RT-PCR. A . Binding sites of primers (AB1553-AB1554), (AB1555-AB1556), (AB1591-AB1592), (AB1559-AB1560), and (AB1561-AB1562), designed to amplify transcripts on omp6, 7, 8, 9 and 10 , respectively, in wild-type (WT) and o mp10::himar1 mutant. Complementary DNA from WT and o mp10::himar1 mutant grown in ISE6 tick cells was used for PCR amplification for o mp6 through 10 with specific primers to evaluate gene expression. B . Agarose gel analysis of PCR products for omp6 through 10 in omp10::himar1 mutant (lanes 2, 8, 14, 20, and 26). PCR products for o mp6 through 10 in WT (lanes 5, 11, 17, 23, and 29). Genomic DNA was used as positive control (lanes 4, 7, 10, 13, 16, 19, 22, 25, 28, and 31). Complementary DNA from reactions without reverse transcriptase were used as negative controls (lanes, 3, 6, 9, 12, 15, 18, 21, 24, 27, and 30). 100 bp/1Kb DNA ladder lane 1). 16S rRNA (AB1572-AB1573) was used as an internal control to ensure integrity of cDNA (lanes 32–37).

    Journal: BMC Genomics

    Article Title: Knockout of an outer membrane protein operon of Anaplasma marginale by transposon mutagenesis

    doi: 10.1186/1471-2164-15-278

    Figure Lengend Snippet: Transcriptional analysis of the effect of the insertion of the Himar1 transposon within the omp10 gene by RT-PCR. A . Binding sites of primers (AB1553-AB1554), (AB1555-AB1556), (AB1591-AB1592), (AB1559-AB1560), and (AB1561-AB1562), designed to amplify transcripts on omp6, 7, 8, 9 and 10 , respectively, in wild-type (WT) and o mp10::himar1 mutant. Complementary DNA from WT and o mp10::himar1 mutant grown in ISE6 tick cells was used for PCR amplification for o mp6 through 10 with specific primers to evaluate gene expression. B . Agarose gel analysis of PCR products for omp6 through 10 in omp10::himar1 mutant (lanes 2, 8, 14, 20, and 26). PCR products for o mp6 through 10 in WT (lanes 5, 11, 17, 23, and 29). Genomic DNA was used as positive control (lanes 4, 7, 10, 13, 16, 19, 22, 25, 28, and 31). Complementary DNA from reactions without reverse transcriptase were used as negative controls (lanes, 3, 6, 9, 12, 15, 18, 21, 24, 27, and 30). 100 bp/1Kb DNA ladder lane 1). 16S rRNA (AB1572-AB1573) was used as an internal control to ensure integrity of cDNA (lanes 32–37).

    Article Snippet: Based on this, the strategy that we used to map the Himar1 insertion site involved alignment of the sequencing reads obtained by Roche/454 and Illumina methods to two reference sequences, the A. marginale str.

    Techniques: Reverse Transcription Polymerase Chain Reaction, Binding Assay, Mutagenesis, Polymerase Chain Reaction, Amplification, Expressing, Agarose Gel Electrophoresis, Positive Control