Inserted Structures Of Transgenic Rice Bt Shanyou 63 Detected And Event-specific Quantitative Systems Established

Transgenic rice of China is in the world's advanced level, several disease-resistant and insect-resistant transgenic rice lines have been into the stage of approval for safety certificates to commercial applications. But mainly due to safety considerations of them, it has not approved commercial planting. Molecular characteristic analysis is one of the most important aspects to evaluate the safety of transgenic plants. GMO safety management laws and regulations all have this information request both in China and many other countries. We did researches on the genetically modified crops"Bt shanyou 63"which hasn't been approved commercial planting, used PCR,LD-PCR,RT-PCR and qPCR. The main contents and results were as follows:1. Structure of foreign DNA insertion of transgenic cry1Ab/c fusing gene rice line Bt shanyou 63 transformed by particle bombardment method and event-specific qualitative system were validated. Detected the structure of foreign DNA insertion of offspring which were Bt shanyou 63 selfed, the marker gene was founded. The segregation ratio of foreign DNA insertion as single site of offspring was true of Mendelian genetic law2. we designed event-specific primers based on flanking sequences, selected species-specific genes of the genetically modified crops as endogenous reference during PCR amplifications, we established event-specific quantitative systems. The results showed that the quantitative systems had good repeatability after many experiments, the limits of quantification of transgenic rice Bt shanyou 63 event-specific quantitative PCR systems using reference molecules were both 100 copies.3. Detected marker gene and target gene expressed on RNA level of all growth stage in transgenic rice Bt shanyou 63 by RT-PCR, marker gene hasn't expressed of all growth stage and target gene has. On this basis we established quantitative systems of target gene expression. The results showed that the quantitative systems had good repeatability, Efficient and stable expression of target gene in other stage except seedling, The expression of target gene in seedling was mild.4. we established standard curves by the reference molecules, constructed reference molecules by recombinant PCR, detected the mixed samples with known GM content, and then calculated the GM amounts; To constitute the distinction between transgenic rice Bt shanyou 63 and huahui 1 hao; Calculated the amounts of target gene expression. The results showedthe event-specific quantitative systems had good repeatability and reproducibility, recombinant PCR used to construct reference molecules was rapid and feasible.The above researches and conclusions validated structure of foreign DNA insertion of transgenic cry1Ab/c fusing gene rice line Bt shanyou 63 transformed by particle bombardment method and event-specific qualitative system, established event-specific quantitative systems and quantitative systems of target gene expression. Those will provide technical support for detection and safety management of the transgenic rice Bt shanyou 63.