The detection of nucleotide sequences specific for genetically modified organisms (GMOs) in raw and processed food is based on different technological strategies, such as the extraction of DNA and the amplification by polymerase chain reaction (PCR), which allow to obtain qualitative and quantitative information. We developed a multiplex PCR-based DNA assay for simultaneously detecting multiple target sequences in genetically modified (GM) soybean (Roundup Ready(TM)). Internal control target (lectin gene) was included both to assess the efficiency of all reactions and eliminating any false negatives. The post-PCR analysis was carried out by 2.5% agarose gel electrophoresis followed by ethidium bromide staining and densitometric analysis. The multiplex PCR method, showing high sensitivity and specificity, was tested on DNA extracted from certified reference samples containing GM soybean, and from food samples (feeds, food supplements, etc.). Comparison of this method with a quantitative evaluation, carried out by real-time PCR, suggests a possible utilization of the multiplex approach for semi-quantitative determinations. The method reported in this work can considerably reduce the time and the costs of the GM soybean detection, especially in the screening of a large number of food samples.
A multiplex PCR-based assay for the detection of genetically modified soybean
Maccarrone M;
2004-01-01
Abstract
The detection of nucleotide sequences specific for genetically modified organisms (GMOs) in raw and processed food is based on different technological strategies, such as the extraction of DNA and the amplification by polymerase chain reaction (PCR), which allow to obtain qualitative and quantitative information. We developed a multiplex PCR-based DNA assay for simultaneously detecting multiple target sequences in genetically modified (GM) soybean (Roundup Ready(TM)). Internal control target (lectin gene) was included both to assess the efficiency of all reactions and eliminating any false negatives. The post-PCR analysis was carried out by 2.5% agarose gel electrophoresis followed by ethidium bromide staining and densitometric analysis. The multiplex PCR method, showing high sensitivity and specificity, was tested on DNA extracted from certified reference samples containing GM soybean, and from food samples (feeds, food supplements, etc.). Comparison of this method with a quantitative evaluation, carried out by real-time PCR, suggests a possible utilization of the multiplex approach for semi-quantitative determinations. The method reported in this work can considerably reduce the time and the costs of the GM soybean detection, especially in the screening of a large number of food samples.Pubblicazioni consigliate
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