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    Thermo Fisher ecor v enzyme
    Construction strategy of the low-copy pMKP ccd B reverse genetic vector to clone unstable HA segments of IAV. The resulting vector is a hybrid of pSMART-LC-Kan and the Pol-I/ ccd B/Pol-II cassette of pMP ccd B with either AarI or <t>Bsa</t> I or BsmB I sites for restriction/cloning of the desired insert. Briefly, the sequence representing the Pol-I/ ccd B/Pol-II cassette excised from pMP ccd B by digestion with BsrB I were blunt end phosphorylated, and ligated into the <t>EcoR</t> V linearized pSMART-LC-Kan plasmid to generate pMKP ccd B. The pMKP ccd B was suitable to clone and maintain the genetically unstable RT-PCR product of H9N2/SA- and H5N1/VSVRI-HA. Transcriptional terminator (T); Repressor of Primer (ROP); low copy origin of replication (Ori); kanamycin resistance gene (Kanr), ampicillin resistance gene (Ampr).
    Ecor V Enzyme, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 89/100, based on 90 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/ecor v enzyme/product/Thermo Fisher
    Average 89 stars, based on 90 article reviews
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    ecor v enzyme - by Bioz Stars, 2020-08
    89/100 stars
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    Construction strategy of the low-copy pMKP ccd B reverse genetic vector to clone unstable HA segments of IAV. The resulting vector is a hybrid of pSMART-LC-Kan and the Pol-I/ ccd B/Pol-II cassette of pMP ccd B with either AarI or Bsa I or BsmB I sites for restriction/cloning of the desired insert. Briefly, the sequence representing the Pol-I/ ccd B/Pol-II cassette excised from pMP ccd B by digestion with BsrB I were blunt end phosphorylated, and ligated into the EcoR V linearized pSMART-LC-Kan plasmid to generate pMKP ccd B. The pMKP ccd B was suitable to clone and maintain the genetically unstable RT-PCR product of H9N2/SA- and H5N1/VSVRI-HA. Transcriptional terminator (T); Repressor of Primer (ROP); low copy origin of replication (Ori); kanamycin resistance gene (Kanr), ampicillin resistance gene (Ampr).

    Journal: PLoS ONE

    Article Title: Efficient Generation of Recombinant Influenza A Viruses Employing a New Approach to Overcome the Genetic Instability of HA Segments

    doi: 10.1371/journal.pone.0116917

    Figure Lengend Snippet: Construction strategy of the low-copy pMKP ccd B reverse genetic vector to clone unstable HA segments of IAV. The resulting vector is a hybrid of pSMART-LC-Kan and the Pol-I/ ccd B/Pol-II cassette of pMP ccd B with either AarI or Bsa I or BsmB I sites for restriction/cloning of the desired insert. Briefly, the sequence representing the Pol-I/ ccd B/Pol-II cassette excised from pMP ccd B by digestion with BsrB I were blunt end phosphorylated, and ligated into the EcoR V linearized pSMART-LC-Kan plasmid to generate pMKP ccd B. The pMKP ccd B was suitable to clone and maintain the genetically unstable RT-PCR product of H9N2/SA- and H5N1/VSVRI-HA. Transcriptional terminator (T); Repressor of Primer (ROP); low copy origin of replication (Ori); kanamycin resistance gene (Kanr), ampicillin resistance gene (Ampr).

    Article Snippet: Since the cloned HA-segments of H9N2/SA showed instability in the genetic backbone of pCR2.1 sequencing vector, the full-length RT-PCR products of HA segments amplified with primers containing EcoRV and Bsa I sites were digested with EcoR V enzyme (Thermo Scientific, USA) for 1 h at 37°C to produce blunt-ends to allow the ligation into the EcoRV -linearized pSMART-LC-Kan (Lucigen, USA).

    Techniques: Plasmid Preparation, Clone Assay, Sequencing, Reverse Transcription Polymerase Chain Reaction