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A new single-step quantitative pathogen detection system: Template-tagging followed by multiplex asymmetric PCR using common primers and CE-SSCP

Title
A new single-step quantitative pathogen detection system: Template-tagging followed by multiplex asymmetric PCR using common primers and CE-SSCP
Authors
Gi W.S.Yang S.C.Hee S.H.Oh M.-H.Hong G.N.Jin H.P.Gyoo Y.J.
Ewha Authors
오미화
Issue Date
2009
Journal Title
Electrophoresis
ISSN
0173-0835JCR Link
Citation
vol. 30, no. 15, pp. 2728 - 2736
Indexed
SCI; SCIE; SCOPUS WOS scopus
Abstract
Rapid diagnosis of bacterial infection is important for patient management and appropriate therapy during the early phase of bacteria-induced disease. Among the existing techniques for identifying microbial, CE-SSCP combined with 16S ribosomal RNA gene-specific PCR has the benefits of excellent sensitivity, resolution, and reproducibility. However, even though CE-SSCP can separate PCR products with high-resolution, multiplex detection and quantification are complicated by primer-dimer formation and non-specific amplification. Here, we describe a novel technique for multiplex detection and quantification of pathogens by template-tagging followed by multiplex asymmetric PCR and subsequent CE-SSCP. More specifically, we reverse transcribed 16S ribosomal RNAs from seven septicemia-inducing pathogens, tagged the templates with common end sequences, and amplified them using common primers. The resulting amplicons could be successfully separated by CE-SSCP and quantified by comparison to an internal standard. This method yielded results that illustrate the potential of this system for diagnosing infectious disease. © 2009 Wiley-VCH Verlag GmbH & Co. KGaA.
DOI
10.1002/elps.200900074
Appears in Collections:
엘텍공과대학 > 식품공학전공 > Journal papers
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