Event-specific qualitative and quantitative detection of transgenic rice Kefeng-6 by characterization of the transgene flanking sequence

摘要

Using TAIL-PCR (thermal asymmetric interlaced PCR), we analyzed the flanking sequences of transfer DNA (T-DNA) in transgenic rice Kefeng-6, which has insect-resistant genes, Cry1Ac and CpTi. Two junction sequences of T-DNA were identified in the Kefeng-6 genome: one integrated in chromosome 6 with an additional 22-bp insertion and another one in chromosome 9 with a 4-bp insertion between rice genomic DNA and T-DNA. Based on these inserts and border sequences, the event-specific qualitative and quantitative PCR system was established for this line. The relative limit of qualitative detection was assessed to be between 0.1 and 0.05%, and the absolute limit of detection in the quantitative PCR was approximately five initial copies. Two mixed rice samples with known Kefeng-6 contents were used to verify the quantitative method, from which the expected results were observed. This study provides a reliable method and information for detection, identification, and quantification of the presence of non-authorized GM rice Kefeng-6.