Identification and nucleotide sequence of attachment site of the Cflt filamentous phage from Xanthomonas campestris pv. citri.
It has been reported that the attachment site on the phage attP is located from 69.2 to 73.8 min on Cflt RF DNA. KpnI and PstI were used, which cut respectively at 67.2 and 72.6 min of Cflt RF DNA. A 0.54 kb fragment containing attP was obtained. For isolation of the right (attR) and left (attL) junctions of prophage and host chromosomal DNA, lysogen DNA was digested with HindIII and used to prepare a recombinant plasmid library. With Cflt RF DNA as a probe, three types of recombinant plasmids respectively containing inserts of 1.7, 4.3 and 2.8 kb DNA fragments were obtained. Since 1.7 kb represents the internal HindIII fragment of Cflt, 2.8 kb were from the junctional regions of prophage and host chromosomal DNA. Further analysis of restriction fragment patterns suggested that the 2.8 kb fragment contains attR, while the 4.3 kb fragment contains attL. To isolate the attachment site on the host bacteria (attB), recombinant plasmids constructed from HindIII DNA fragments of uninfected cells are screened by using molecular probes prepared from the host DNA sequences immediately adjacent to the prophage in the lysogenic host chromosomal DNA. A 1.45 kb fragment was obtained. Further analysis with HindIII site on this fragment established that it represents attB. The DNA nucleotide sequences of attP, attB attR and attL were determined. All of them contain a common 15 nucleotide core with the sequence of 5'-TATACATTATGCGAA-3'. The common core region of the recombination site shows two inverted-repeat DNA sequence.
Fann
J H
JH
Department of Botany National Taiwan University, Taipei, Republic of China.
Lee
M H
MH
Kuo
T T
TT
D001287:Attachment Sites, Microbiological; D001435:Bacteriophages / Q000235:genetics*; D001483:Base Sequence; D004279:DNA, Viral / Q000032:analysis*; D008242:Lysogeny; D008969:Molecular Sequence Data; D014974:Xanthomonas / Q000235:genetics*
D004279:DNA, Viral