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標題: Plasmids carrying cloned fragments of RF DNA from the filamentous phage phi Lf can be integrated into the host chromosome via site-specific integration and homologous recombination
作者: Lin, N.T.
Chang, R.Y.
Lee, S.J.
Tseng, Y.H.
關鍵字: Xanthomonas;filamentous phage;Xer-specific integration system;homologous recombination;campestris pv campestris;gram-negative bacteria;escherichia-coli;xanthomonas-campestris;terminus region;nucleotide-sequence;adsorption;protein;binding protein;coat protein;gene
Project: Molecular Genetics and Genomics
期刊/報告no:: Molecular Genetics and Genomics, Volume 266, Issue 3, Page(s) 425-435.
Different regions of RF DNA from the filamentous bacteriophage phi Lf were cloned in Escherichia coli vectors that can not be maintained in Xanthomonas. After introduction into X. campestris pv. campestris 17 (Xc17), most of these constructs were found to integrate into the host chromosome, either by recA-dependent homologous recombination or recA-independent site-specific integration, Mutations in himA, which codes for the alpha -subunit of the Integration Host Factor, does not affect the integration, Integration occurs into a chromosomal region which harbors a copy of a defective phage (4445 bp) that shares a high degree of identity with the phi Lf genome. While various parts of the 4445-bp region are susceptible to homologous recombination, site-specific integration requires the attB sequence on the chromosome and the phage attP. The attB shows a high level of sequence identity (22 out of 28 bp) to the dif site required for E. coli Xer site-specific recombination, including the 6-bp central region, and 8/11 identity in both the left XerC-binding arm and the right XerD-binding arm, with the innermost 5 nt of the arms forming a dyad symmetry that is also present in dif. The attP has the same central region and shows 10/11 identity to the dif site in the left arm, but the sequence of the right arm is less conserved than that of attB. The smallest regions still capable of mediating integration are a cloned 72-bp phi Lf attP-containing sequence and a 51-bp Xc17 attB-containing sequence, which was reinserted into the Xc17 chromosome after the 4445-bp region had been deleted, indicating that accessory sequences are not necessary and that the integrase required for site-specific integration is neither specified by the 4445-bp Xc17 chromosomal region nor encoded by the phi Lf genome.
ISSN: 1617-4615
DOI: 10.1007/s004380100532
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