کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
2091192 1081537 2006 15 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Detection of bacterial pathogens in municipal wastewater using an oligonucleotide microarray and real-time quantitative PCR
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی بیوتکنولوژی یا زیست‌فناوری
پیش نمایش صفحه اول مقاله
Detection of bacterial pathogens in municipal wastewater using an oligonucleotide microarray and real-time quantitative PCR
چکیده انگلیسی

As a first step toward building a comprehensive microarray, two low density DNA microarrays were constructed and evaluated for the accurate detection of wastewater pathogens. The first one involved the direct hybridization of wastewater microbial genomic DNA to the functional gene probes while the second involved PCR amplification of 23S ribosomal DNA. The genomic DNA microarray employed 10 functional genes as detection targets. Sensitivity of the microarray was determined to be approximately 1.0 μg of Esherichia coli genomic DNA, or 2 × 108 copies of the target gene, and only E. coli DNA was detected with the microarray assay using municipal raw sewage. Sensitivity of the microarray was enhanced approximately by 6 orders of magnitude when the target 23S rRNA gene sequences were PCR amplified with a novel universal primer set and allowed hybridization to 24 species-specific oligonucleotide probes. The minimum detection limit was estimated to be about 100 fg of E. coli genomic DNA or 1.4 × 102 copies of the 23S rRNA gene. The PCR amplified DNA microarray successfully detected multiple bacterial pathogens in wastewater. As a parallel study to verify efficiency of the DNA microarray, a real-time quantitative PCR assay was also developed based on the fluorescent TaqMan® probes (Applied Biosystems).

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Journal of Microbiological Methods - Volume 65, Issue 3, June 2006, Pages 453–467
نویسندگان
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