Integration Sites Analysis Of Exogenous Genes In Transgenic Animals

Transgenic animal has a broad development prospects,its showed in many aspects, such as agriculture, food, medicine, environment. In 1974, Jaenisch and his team injected ape cavitation virus(SV40) DNA into mouse blastocyst, from then until now, a large number of genetically modified(gm) plants and animals have sprung up. When people immersed in huge success brought by transgenic plants and animals, the safety problems appeared which brought by transgenic animals and plants as well. On September 21, 2012, an article named "More than half of mice what eating genetically modified corn two years long had tumors", the report does waked people up. The article reminds us that we should not blindly in production and application of transgenic animals, avoiding damage.safety problems of genetically modified(gm) plants and animals testing must be done. Safety test should be carried out in various aspects, first of all, is the detection of gene, including the detection of integration sites of exogenous gene in the genetically modified(gm) host genome, and the influence of the expression of the host genome; Secondly, the detection of transcription level, we should detect the level of m RNA of exogenous genes and host gene; Third, the testing of protein translation, we should measure the expression of host genes and exogenous genes; Fourth, the detection of actual application. Evaluate weather the transgenic plants and animals are safe. we detected the integration sites and m RNA expression of transgenic mice and transgenic cattle.We detected integration sites of three transgenic mice and a transgenic cattle with TAIL-PCR, hi TAIL-PCR and improved PCR, and we detected expression situation of IMMP2 L gene by fluorescence quantitative PCR. Provide the basis for the safety assessment of genetically modified(gm) animals.1 In transgenic mice A, we amplificated a special band. We found in the segment, there is two segments of plasmid connected to each other after blasting with Known nucleic acid sequence of mouse in Gene Bank.2 We got a specific sequence in transgenic mice B,recombinant plasmid integrated after 43264229 site of chromosome 12 of mouse, and Matching sequence homology rate is 99%. Among the sequence, there is a 6 bp bases, it not only belong to recombinant plasmid and also belong to mouse genome sequences. Which is proved that foreign gene integration into host genome by homologous recombination.3 Recombinant plasmid did not broken in enzyme loci not only in transgenic mice or in transgenic cattle, but broken randomly.4 In transgenic mouse C, we found that the 3 ’end of recombinant plasmid inserted before 171864372 site of mouse chromosome 2, recombinant plasmid is not direct connected with host chromosomes, it is unknown sequence between them; and we also found that recombinant plasmid inserted before 13858398 site of mouse chromosome 14, broken site of recombinant plasmid located the forth basis after enzyme loci and integrated way is randomly; We designed special primers at downstream of recombinant plasmid enzyme loci to amplify integration site of transgenic cattle, and found recombinant plasmid inserted before 13904955 site of cattle chromosome 16. These results proved that the integration of foreign genes is polysites random integration, in the process of integrating producted unknown sequences.5 In the detecting process of integration site, we found that both end of host gene connected with unknown sequences from the sequencing result of transgenic mouse A and transgenic cattle, which proved that exogenous gene integrated into the host chromosome can break host genome.6 We often detected unknown sequences while in the process of detecting integration site. We speculated these unknown sequences may came from pollution or sequence formed from all kinds of little segments of host genome in the process of exogenous gene integrating.7.Studing the m RNA expression quantity of IMMP2 L by fluorescence quantitative PCR, proved that exogenous gene can break the expression of host genome near the integration site.