Detection And Identification Of Transgenic Corps Products By Real-time Fluorescent PCR

According to ISAAA count, the cultivated area of transgenic crops in the whole world is 52,600,000 hectares in 2001, More than ten countries have already planted the transgenic crops, 99% of the transgenic crops among them are planted in U.S.A., Argentina, Canada and China. Commercialized transgenic crops are soybeanses, Maize, rapes, cottons, tomatoes, potatoes, sweet pepper, French gourd, Chinese flowering quinces, beets, fragrant China pink, morning glory, flax, tobacco, kinds of food that is processed with the transgenic crops in the whole world.People and animal security from the transgenic products, especially transgenic food healthy and the impact on ecological environment, Since the gene technology appeared, has been the focus question of international organizations, including many countries and UN, etc. and make out a series of relevant policies and laws successively. A lot of countries among them limited the minimum content of the transgenic production, The threshold value is varying from 1% to 5%. It need to detect transgenic products precisely, definitely. In order to prevent some transgenic products with risk from spreading, our country announced and implemented" agricultural transgenic biology safe management rule* on May 9, 2001 too, And announced agricultural transgenic biology safe appraisal, sign and import the management method of three complete networks of safety management on January 5, 2002. Various of transgenic products relevant laws and regulations issue present in 36 countries and regions in the whole world and some regional international organizations are convening to set up the unified detection technique and method standard of the world in the organization ( FAO, WHO, etc.) under the UN, But inconsistent to the security,risk control measures and other interests of the transgenic products, etc., has been reached unanimous suggestion at present.For measuring transgenic products effectively, we collect gene structure of transgenic crops that commercialized already both at home and abroad, Choose 7 genes fragment (35S, nos, nptII,pat, bar, FMV, cry3A) that cover all already commercialized transgenic crops as goal of screening, and design fluorescence prime and probe especially, utilize RT-PCR method to measure transgenic target peculiarly, and sensitivity relativly is 1%. We have set up transgenic RT-PCR measure system determine transgenic Bt - 176 maize and the Roundup ReadyTM soybean and adopt UNG enzyme to reduce pollution possibility at the same time. In order to confirm the genes further, we clone some PCR products to T - carrier, transforme E. coli DH5 a , screen positive clone, and sequencing. The sequence homology between target product and GeneBank reach more than 99. 9%. We cloned two length genes fragment for positive control at the same time.This paper set up transgenic RT-PCR detection method for the first time at home and cooperate with company to put out the related Kit. This will exert a far-reaching influence on measuring import and export the transgenic products and have important realistic meanings in the future.