Electronic microarray analysis of 16S rDNA amplicons for bacterial detection.
Electronic microarray technology is a potential alternative in bacterial detection and identification. However, conditions for bacterial detection by electronic microarray need optimization. Using the NanoChip electronic microarray, we investigated eight marine bacterial species. Based on the 16S rDNA sequences of these species, we constructed primers, reporter probes, and species-specific capture probes. We carried out two separate analyses for longer (533 bp) and shorter (350 and 200 bp) amplified products (amplicons). To detect simultaneously the hybridization signals for the 350- and 200-bp amplicons, we designed a common reporter probe from an overlapping sequence within both fragments. We developed methods to optimize detection of hybridization signals for processing the DNA chips. A matrix analysis was performed for different bacterial species and complementary capture probes on electronic microarrays. Results showed that, when using the longer amplicon, not all bacterial targets hybridized with the complementary capture probes, which was characterized by the presence of false-positive signals. However, with the shorter amplicons, all bacterial species were correctly and completely detected using the constructed complementary capture probes.
Barlaan
Edward A
EA
Nagasaki Industrial Promotion Foundation, Ikeda 2-1303-8, Omura City, Nagasaki 856-0026, Japan. ebarlaan@ngsrdc.mhi.co.jp
Sugimori
Miho
M
Furukawa
Seiji
S
Takeuchi
Kazuhisa
K
D001419:Bacteria / Q000235:genetics* / Q000302:isolation & purification*; D015169:Colony Count, Microbial / Q000379:methods*; D015342:DNA Probes / Q000235:genetics; D004582:Electronics, Medical / Q000295:instrumentation / Q000379:methods; D004784:Environmental Monitoring / Q000295:instrumentation / Q000379:methods*; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods; D012336:RNA, Ribosomal, 16S / Q000032:analysis / Q000235:genetics*; D014871:Water Microbiology
D015342:DNA Probes; D012336:RNA, Ribosomal, 16S