Identification of a novel repetitive DNA element and its use as a molecular marker for strain typing and discrimination of ara- from ara+ Burkholderia pseudomallei isolates.
A novel 10-bp repeat (5'-CGACGCAGGC-3')34 was identified in a strain of Burkholderia pseudomallei, the first repetitive element found in this species. A pair of primers, based on the flanking sequences of the repetitive region, was used in PCR and DNA sequence analysis to determine the genomic structure and distribution of the repetitive element in 76 arabinose- (ara-) and 7 ara+ B. pseudomallei isolates. DNA fragments of 400-700 bp were amplified in all ara- isolates. Ara+ isolates were characterised by a uniform fragment of 402 bp. Nucleotide sequence analysis of these fragments revealed broad heterogeneity of the variable-number tandem repeats with 26 distinct alleles ranging between (5'-CGACGCAGGC-3')13 and (5'-CGACGCAGGC-3')45 identified in the ara- isolates. In contrast, a novel non-repetitive sequence was identified in each of the ara+ isolates. This was confirmed by Southern blot analysis. Such biotype-specific variable-number tandem repeats may be useful as genetic markers for rapid strain differentiation of ara- isolates.
Liu
Yichun
Y
Wang
Dongling
D
Yap
Eu Hian
EH
Biomedical Sciences Laboratory, Defence Medical Research Institute and *Department of Microbiology, National University of Singapore, Singapore.
Yap
Eric P H
EPH
Lee
May-Ann
MA
D000483:Alleles; D000595:Amino Acid Sequence; D000818:Animals; D001089:Arabinose / Q000378:metabolism*; D015373:Bacterial Typing Techniques; D001483:Base Sequence; D015139:Blotting, Southern; D016957:Burkholderia pseudomallei / Q000145:classification* / Q000235:genetics* / Q000378:metabolism / Q000472:pathogenicity; D004269:DNA, Bacterial / Q000032:analysis* / Q000737:chemistry; D004587:Electrophoresis, Agar Gel; D005819:Genetic Markers; D006801:Humans; D008969:Molecular Sequence Data; D008970:Molecular Weight; D021141:Nucleic Acid Amplification Techniques; D010802:Phylogeny; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length; D017422:Sequence Analysis, DNA; D020080:Tandem Repeat Sequences; D014774:Virulence
D004269:DNA, Bacterial; D005819:Genetic Markers; D001089:Arabinose