Direct detection of 16S rRNA using oligonucleotide microarrays assisted by base stacking hybridization and tyramide signal amplification.
A simple method has been developed and validated for direct, sensitive detection and specific identification of 16S rRNA. We first report our direct investigation of discrimination efficiency for sequence variations in RNA using oligonucleotide microarrays assisted by base stacking hybridization, and demonstrate that the sequence variations of double base substitution, single base substitution and single base deletion in RNA could be directly identified. With the help of tyramide signal amplification (TSA), the detection sensitivity of this method for four clinically important bacterial species was below 0.5, 5, 1 and 1 ng of total RNA, which are 100-1000 fold more sensitive than the published methods.
Wang
Dong
D
Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing 100084, PR China.
Zhu
Lingxiang
L
Jiang
Di
D
Ma
Xuemei
X
Zhou
Yuxiang
Y
Cheng
Jing
J
D015342:DNA Probes / Q000235:genetics; D009693:Nucleic Acid Hybridization; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D012336:RNA, Ribosomal, 16S / Q000032:analysis* / Q000737:chemistry / Q000235:genetics; D012680:Sensitivity and Specificity; D014439:Tyramine / Q000031:analogs & derivatives* / Q000737:chemistry*
D015342:DNA Probes; D012336:RNA, Ribosomal, 16S; D014439:Tyramine