scholarly journals Simultaneous Detection of 10 Foodborne Pathogens using Capillary Electrophoresis-Based Single Strand Conformation Polymorphism

2012 ◽  
Vol 32 (2) ◽  
pp. 241-246 ◽  
Author(s):  
Mi-Hwa Oh ◽  
Hee-Sung Hwang ◽  
Bo-Ram Chung ◽  
Hyun-Dong Paik ◽  
Sang-Ha Han ◽  
...  
Keyword(s):  
Capillary Electrophoresis ◽  
Foodborne Pathogens ◽  
Simultaneous Detection ◽  
Single Strand Conformation Polymorphism ◽  
Single Strand ◽  
Conformation Polymorphism

Simultaneous Identification of Seven Foodborne Pathogens and Escherichia coli (Pathogenic and Nonpathogenic) Using Capillary Electrophoresis–Based Single-Strand Conformation Polymorphism Coupled with Multiplex PCR

2009 ◽  
Vol 72 (6) ◽  
pp. 1262-1266 ◽  
Author(s):  
MI-HWA OH ◽  
SE-HEE PAEK ◽  
GI WON SHIN ◽  
HAE-YEONG KIM ◽  
GYOO YEOL JUNG ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Capillary Electrophoresis ◽  
Multiplex Pcr ◽  
Simultaneous Detection ◽  
Pcr Amplification ◽  
Single Strand Conformation Polymorphism ◽  
Single Strand ◽  
Rrna Gene ◽  
Optimal Separation ◽  
Conformation Polymorphism

The objective of this study was to develop a novel technique for parallel analysis of eight important foodborne microbes using capillary electrophoresis–based single-strand conformation polymorphism (CE-SSCP) coupled with multiplex PCR. Specific primers for multiplex PCR amplification of the 16S rRNA gene were designed, corresponding to eight species of bacteria, including Escherichia coli, Clostridium perfringens, Campylobacter jejuni, Salmonella enterica, Listeria monocytogenes, Vibrio parahaemolyticus, Staphylococcus aureus, and Bacillus cereus, for the species-specific identification and optimal separation of their PCR products in subsequent analysis by CE-SSCP. Multiplex PCR conditions including annealing temperature, extension time, the number of PCR cycles, and primer concentrations were then optimized for simultaneous detection of all target foodborne bacteria. The diagnostic system using CE-SSCP combined with multiplex PCR developed here can be used for rapid investigation of causative agents of foodborne illness. The simplicity and high sensitivity of the method may lead to improved management of safety and illness related to food.

Multiplex Single Strand Conformation Polymorphism Analysis by Capillary Electrophoresis with On-the-Fly Fluorescence Lifetime Detection

2005 ◽  
Vol 59 (3) ◽  
pp. 335-339 ◽  
Author(s):  
Tara Morcone Snyder ◽  
Linda B. McGown
Keyword(s):  
Capillary Electrophoresis ◽  
Fluorescence Lifetime ◽  
Single Strand Conformation Polymorphism ◽  
Single Strand ◽  
Polymorphism Analysis ◽  
Wild Type ◽  
G20210a Mutation ◽  
Prothrombin Gene ◽  
Fluorescence Lifetime Detection ◽  
Conformation Polymorphism

This paper describes the use of on-the-fly fluorescence lifetime detection (OFLD) for multiplex single strand conformation polymorphism (SSCP) analysis by capillary electrophoresis (CE). The dye labels studied for multiplex SSCP-OFLD-CE analyses included RG, NBD, and BODIPY-FL. The dyes were first investigated for a model system of “Wild Type” and “Mutant” 43-base fragments designed to vary by a single A/T substitution. Two dye pairs, BODIPY-FL/RG and BODIPY-FL/NBD, were then used to detect the G20210A mutation in the human prothrombin gene. Mobility correction was required for the BODIPY-FL/RG system. Three “blind” analyses were performed of three mixtures that combined a control fragment (wild type-BODIPY-FL) with two “unknown” fragments selected among four possibilities (wild type or mutant labeled with NBD or RG). In each multiplex analysis, the “origin” of the unknown fragments was correctly identified on the basis of fluorescence lifetime of the dye label and the presence or absence of the mutation was correctly determined on the basis of conformation-induced differences in migration time.

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