The Expression Of Silkworm Transgenic Vector PBacA3EG,pBacA4R In Insect Cell

In recent years, the transgenic research on animals has been becoming one of the best hot-points in Genetics and Biology. Silkworm (Bombyx mori) , is not only an important economic insect, but also an important organisms for research classical genetics. Studies on transgenic silkworm possess theoretical significance and practical value. PiggyBac is enable foreign gene to transposion in chromosome genome, and gene heredity gives to the future generation. The studies of silkworm transgenic are set up on the foundation of classical genetics, molecular genetics, structural genetics and DNA recombinant technique. The whole gene order-checking of silkworm finishing recently (Xia et al.,2004), it will promote the basic research of the silkworm, and make the silkworm play a positive role in the development of insect's discipline as the mode insect. After the draft of the silkworm genome, the present groundwork is to carry on research to a large number of genes obtained of genome plan of silkworm, in order to explore its concrete biological function. Because transgenic technology can realize the disappearance of excessive expression or genetic function of silkworm's function gene, the study of silkworm transgenic will become one of silkworm's genetic function important methods. At the same time.it is the key to utilizing the function gene to develop new assortment of silkworm, even still realize utilizing silkworms to formate the useful protein, technological key of developing silkworm's bioreactor.This thesis mainly includes the constructing transgenic vectors, expressing, cells being trained and screened. The main research work includes the following several aspects:1. Designing the primers according to the promoter sequences of the reported A3 gene of silkworm and cloning A3 promoter. Throught the comparative analysis with the actin 4 gene of silkworm, 3 groups primers, A4-1, A4-2, and A4-3, were designed. The produce of A4-2 primer was ligated to the transgenic vector finally and succeeded to promote the expression of the reporter gene.2. By using of three basic elements, actin3 promoter of Bombyx mori, enhancing green fluorescent protein gene (EGFP) and the identified sequence of polyadenylation acid in SV40, the piggyBac transposon vector pBacA3EG was constructed. This vector was further confirmed by PCR, enzyme digestion and sequencing analysis, and the results show that all elements have been inserted into the piggyBac vector correctly. We transfected the piggyBac expression vector into silkworm cell line BME and observed the red fluorogenic cells 48 hours later. Through testing at the molecular...