Identification of pathogenic Helicobacter species by chaperonin-60 differentiation on plastic DNA arrays.
A microarray method for bacterial species identification based on cpn60 and 16S rDNA hybridization was developed. Specific cpn60 or 16S rDNA oligonucleotides from various Helicobacter or Campylobacter species were printed and immobilized onto a proprietary plastic solid support. Using universal primers, fragments derived from either cpn60 or 16S rDNA genes from single isolates or from a complex human waste sludge DNA sample spiked with Helicobacter pylori were biotinylated and hybridized to the plastic slide. Subsequent querying with a streptavidin-horseradish peroxidase conjugate followed by color development using tetramethylbenzidine resulted in accurate Helicobacter species identification with no cross-hybridization to either the 16S rDNA or the cpn60 sequence of a closely related strain of Campylobacter jejuni. The combination of a nonfluorescence visual detection system with a polymer-based DNA microarray slide has resulted in a molecular tool that should prove useful in numerous applications requiring rapid, low-cost bacterial species identification.
Masson
Luke
L
Biotechnology Research Institute, National Research Council of Canada, Montréal, QC, Canada. luke.masson@nrc-cnrc.gc.ca
Maynard
Christine
C
Brousseau
Roland
R
Goh
Swee-Han
SH
Hemmingsen
Sean M
SM
Hill
Janet E
JE
Paccagnella
Ana
A
Oda
Ryuichi
R
Kimura
Naoki
N
D016123:Campylobacter jejuni / Q000235:genetics; D018834:Chaperonin 60 / Q000032:analysis / Q000235:genetics*; D004269:DNA, Bacterial / Q000032:analysis / Q000235:genetics*; D004275:DNA, Ribosomal / Q000032:analysis / Q000235:genetics*; D016998:Helicobacter / Q000235:genetics*; D006824:Hybridization, Genetic; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods; D012336:RNA, Ribosomal, 16S / Q000032:analysis / Q000235:genetics*; D012680:Sensitivity and Specificity; D013045:Species Specificity
D018834:Chaperonin 60; D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S