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    • #9724
      lara
      Participant

      Now this has been puzzling me for sometime.The following is from the
      paper titled "Quorum-sensing system influences root colonization and
      biological control
      ability in Pseudomonas fluorescens 2P24"
      (Hai-Lei Wei,and Li-Qun Zhang).can anyone help me out with this? the
      trouble is can’t visualize it and without it can’t proceed…..

      To create a pcoI gene deletion allele, two fragments
      flanking pcoI gene were amplified by PCR. One was created by primers
      I473
      (5′-TCGTTCGCTGCAGGAAACC-3′) which included the Pst1 site and I2056 (5′-
      ATGGATCCCGGAGTTTGTGCATGCCC-3′)which introduced the BamH1 site and the
      other was created by primers I2328 (5′-
      AAGGATCCGGGTTCGGCTTCTCTGACAC-3′) which introduced the BamH1 site and
      I3966 (5′-ATGAATTCAATGGCCCACTCAACGCA-
      3′) which introduced the EcoR1 site.The standard PCR was carried
      out.After being digested with relevant restriction enzymes, the two
      fragments were inserted into pBSNot6.

    • #84510
      Cat
      Participant

      Normal pcoI gene:

      — Flanking region 1 — pcoI gene — Flanking region 2 —

      Deletion of pcoI gene:

      -(Pst1)- Flanking region 1 — (BamH1) — Flanking region 2 -(EcoR1)–

      Homologous recombination substitutes normal allele with the deletion.

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