Innovative Application of Fluorescent Microsphere Based Assay for Multiple GMO Detection

摘要

An innovative multiple screening protocol allowing the detection of specific DNA sequences of p35S and epsps in GM soya flours simultaneously was developed. For the application of the Luminex xMAP technology, two different sets of fluorescent beads were cross-linked to the specific oligonucleotide probes previously amplified and labeled by polymerase chain reaction (PCR) in the presence of a biotinylated nucleotide. The detection of the amplification products bounded by the streptavidin-phycoeritrinconjugate was performed using the Luminex-100 instrument. The system allows identification of extremely low amounts of labeled targets as of 7.8 10~(-4) nmol of p35S antiprobe with an associated value of repeatability relative standard deviation (RSD_r)equal to 2.8. Parameters such as repeatability, reproducibility, limit of detection (LOD), and limit of quantification (LOQ) were considered to evaluate the performances of the assay. The potentiality of the system described in this paper enables us to look forward to a multiple target assay that is able to simultaneously detect and eventually quantify tens of target sequences occurring within the same sample preparation. The present study describes the first application of a multitarget fluorescent microsphere-based assay for DNA genetically modified organism (GMO) detection.