Evaluation of custom-made DNA microarrays for multilocus sequence typing of Neisseria meningitidis.
Multilocus sequence typing (MLST) has become the gold standard for typing of a variety of bacterial and fungal micro-organisms. Others recently reported the successful use of the tiling DNA array technology to sequence-type Staphylococcus aureus. We now evaluated microarrays based on polymorphism-directed oligonucleotide design for typing of Neisseria meningitidis. The rationale behind this approach was to minimize the number of microarray probes by exploiting the comprehensive knowledge of polymorphisms combined in the Neisseria MLST website. Initial experiments using model oligonucleotides of 28-32 base-pairs in length revealed that the hybridization protocols used were highly specific. However, despite of several optimization steps, the rate of misidentification of oligonucleotides remained > 1.8% in consecutive validation experiments using arrays representing the genetic diversity at three MLST loci. We assume that the high density of polymorphic sites and the extensive GC-content variations at N. meningitidis MLST loci hinder the successful implementation of MLST microarrays based on polymorphism-directed oligonucleotide design.
Swiderek
Halina
H
Institute for Hygiene and Microbiology, University of Würzburg, Josef-Schneider-Str. 2, D-97080 Würzburg, Germany.
Claus
Heike
H
Frosch
Matthias
M
Vogel
Ulrich
U
D000483:Alleles; D015373:Bacterial Typing Techniques / Q000379:methods*; D001482:Base Composition; D004269:DNA, Bacterial / Q000032:analysis / Q000302:isolation & purification; D009345:Neisseria meningitidis / Q000145:classification* / Q000302:isolation & purification; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D011110:Polymorphism, Genetic; D012680:Sensitivity and Specificity
D004269:DNA, Bacterial