Development of a real-time PCR method to detect potentially allergenic sesame (Sesamum indicum) in food

Recent papers indicate that the prevalence of allergic reactions to sesame (Sesamum indicum) is increasing in European countries. This paper describes the development of a selective real-time PCR method for the detection of sesame in food. The assay did not show any cross-reactivity with 17 common food ingredients. The real-time PCR method was applied to determine sesame in several crackers, salty snacks, biscuits, tahina sesame paste and sesame oil. With the exception of sesame oil, in all of the samples where sesame was declared, sesame was detected by the real-time PCR assay (Ct value<35). In the samples which might contain sesame or where sesame was not listed, sesame could not be detected (Ct value>35).     

Development of a real-time PCR method for the differential detection and quantification of four solanaceae in GMO analysis: potato (Solanum tuberosum), tomato (Solanum lycopersicum), eggplant (Solanum melongena), and pepper (Capsicum annuum)

The labeling of products containing genetically modified organisms (GMO) is linked to their quantification since a threshold for the presence of fortuitous GMOs in food has been established. This threshold is calculated from a combination of two absolute quantification values: one for the specific GMO target and the second for an endogenous reference gene specific to the taxon. Thus, the development of reliable methods to quantify GMOs using endogenous reference genes in complex matrixes such as food and feed is needed. Plant identification can be difficult in the case of closely related taxa, which moreover are subject to introgression events. Based on the homology of beta-fructosidase sequences obtained from public databases, two couples of consensus primers were designed for the detection, quantification, and differentiation of four Solanaceae: potato (Solanum tuberosum), tomato (Solanum lycopersicum), pepper (Capsicum annuum), and eggplant (Solanum melongena). Sequence variability was studied first using lines and cultivars (intraspecies sequence variability), then using taxa involved in gene introgressions, and finally, using taxonomically close taxa (interspecies sequence variability). This study allowed us to design four highly specific TaqMan-MGB probes. A duplex real time PCR assay was developed for simultaneous quantification of tomato and potato. For eggplant and pepper, only simplex real time PCR tests were developed. The results demonstrated the high specificity and sensitivity of the assays. We therefore conclude that beta-fructosidase can be used as an endogenous reference gene for GMO analysis.     

Use of real-time polymerase chain reaction (PCR) and transformation assay to monitor the persistence and bioavailability of transgenic genes released from genetically modified papaya expressing nptII and PRSV genes in the soil

Soil samples were collected from an isolated field from December 2003 to April 2004 where transgenic papaya were planted, and the persistences of transgenic genes of 796 bp (located between 35S promoter and coat protein, 35S-P/PRSV-CP), 398 bp (located between plasmid pBI121 and NOS terminator, pBI121/NOS-T), and 200 bp (located between NOS promoter and nptII gene, NOS-P/nptII) were studied. At the end of planting, the residues of 398 bp in the soil was 0.06 microg g(-1) of soil, whereas the residues of 769 and 200 bp were less than 30 pg g(-1) of soil (detection limit). Kinetics studies on the persistence of these three fragments in sterile distilled water and nonsterile soil microcosms showed that two mechanisms might be involved: an initial fast exponential degradation pattern in the first week and then followed by a slow-release pattern throughout the experiment. Persistence of transgenic DNA in sterile water was longer than in nonsterile soil microcosms, indicating that enzymatic degradation and soil adsorption played important roles on the persistence of DNA in the environment. The reason for the fragment of 398 bp persisted longer than fragments of 769 and 200 bp is not clear, but the guanine plus cytosin (G plus C) content in the DNA fragment might be involved in the stability of DNA in the environment. Biological availability of soil DNA to bacteria conducted by the transformation assay indicated that gene transformation from soil DNA extracts to two Acinetobacter spp. did not occur.