Swiderek Halina, Claus Heike, Frosch Matthias, Vogel Ulrich
Institute for Hygiene and Microbiology, University of Würzburg, Josef-Schneider-Str. 2, D-97080 Würzburg, Germany.
Int J Med Microbiol. 2005 Apr;295(1):39-45. doi: 10.1016/j.ijmm.2004.12.005.
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.
多位点序列分型（MLST）已成为多种细菌和真菌微生物分型的金标准。其他人最近报道了成功使用平铺DNA阵列技术对金黄色葡萄球菌进行序列分型。我们现在评估了基于多态性导向寡核苷酸设计的微阵列用于脑膜炎奈瑟菌的分型。这种方法背后的基本原理是通过利用脑膜炎奈瑟菌MLST网站中组合的多态性综合知识来最小化微阵列探针的数量。使用长度为28 - 32个碱基对的模型寡核苷酸进行的初步实验表明，所使用的杂交方案具有高度特异性。然而，尽管经过了几个优化步骤，在使用代表三个MLST位点遗传多样性的阵列进行的连续验证实验中，寡核苷酸的错误识别率仍>1.8%。我们认为，脑膜炎奈瑟菌MLST位点的多态性位点高密度和广泛的GC含量变化阻碍了基于多态性导向寡核苷酸设计的MLST微阵列的成功实施。
