Research Of Transferred Synthetic Nattokinase Gene In Tobacco

In order to enhance the expression of nattokinase gene, the nattokinase gene sNK, which was artificially synthesized according to plant preferential codons, and the nattokinase gene sNK-E8i, which has been got by inserting the first intron of tomato gene E8into sNK, were transformed into tobacco NC89. We obtained the best conditions for transient expression of nattokinase gene in tobacco NC89by orthogonal experiments. The nattokinase gene were expressed in sNK-E8i plants and sNK plants by RT-qPCR. It proved that our synthetic methods is feasible.We converted sNK gene and sNK-E8i gene into tobacco NC89through the way of Agrobacterium infection.12strains of sNK transgenic tobacco plants and10strains of sNK-E8i transgenic tobacco plants were acquired through PCR test. It preliminarily demonstrated that the target genes were integrated into tobacco genomes. It was found that9strains of sNK transgenic tobacco plants and10strains of sNK-E8i transgenic tobacco plants were positive by using RT-PCR. The result showed that the expression level of sNK-E8i transgenic tobacco plants was higher than the level of sNK transgenic tobacco plants by RT-qPCR. Besides, the enzyme activity was tested through agarose-fibrin plate analysis, and the dissolving circles of5strains of sNK transgenic tobacco plants and3strains of sNK-E8i transgenic tobacco plants were detected. The result showed that target genes could be transcribed and translated normally in a part of transgenic tobacco and presented the activity of thrombolysis. The T1transgenic straints were obtained by generation-adding cultivation.