pcr

[lara]

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.

[Cat]

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.

[Author]

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