Abstract:

A multiplex PCR assay coupled with liquid bead array was developed for simultaneous identification of nine
genetically modified rice (Oryzae sativa) lines. Based on the junction sequences between host plant genome DNA and the
exogenous gene of the GM rice events (BT63, KF6, KF8, M12, KMD1, T1C-19, T2A-1, LL601, and LL62), the eventspecific
primers and probes were designed for each event. In addition, in order to reduce the number of PCR reactions, two
multiplex PCR systems were adopted and optimized by adjusting the concentration of each primer. Detection specificity was
provided by capture probes designed for each GMO rice event which were covalently attached to florescent coding beads.
The PCR amplicons were biotin-labeled and directly hybridized with the capture probes on the beads, and then the beads
were detected according to protocol of Bio-Rad. The sensitivity of the assay was evaluated and the results indicated that
the limit of detection was around 0.1% for 9 events. With improved detection efficiency and accuracy, the detection system
complied with the requirements of current regulations in EU, Japan and China for the thresholds for the labeling of GMO,
and could be applied in the enforcement of GM regulation in China.

Key words: multiplex PCR, liquid bead array, transgenic rice, detection