Establishment Of Transgenic Mouse Model Expressing Human Lactoperoxidase

The mammary gland bioreactor is the technique that a kind of using animal transgenic technology to express proteins, in mammalian cells, such as pharmaceutical polypeptide, industry enzyme , vaccine, antibody and etc. This potential technique is characterized by producing high output in low cost, its efficiency is as 100 times as that of cultivating echerichia coli and animal cells. In 1987, the Simons first successfully expressed sheep milk globulin gene in mammary glands of transgenic mice. What' s more, the concentation of this recombinant protein is up to 23 g/1, which is about 400 times higher than that expressed by animal cells. The technique advanced with remarkable speed since its appear. Many famous scientists all consider it is an unprecedented revolution in animal husbandry which may bring the society enormous economic results. Especially, with the development of clone, the transgenic technology also has got well developed. This will make the large-scale production of the mammary glands bioreactor more realistic, therefore an international competition around this technique will be inevitable.This research is to establish a model of transgenic mice which can express human lactoperoxidase (LPO)efficiently in mammary gland cells mainly. Lactoperoxidase belongs to mammalian heme peroxidase superfamily , it can widely be found in milk and other secretory fluid . It can oxidate thiocyanate and othe substrates under the circumstances of H202, by this reaction, human lactoperoxidase can kill or inhibit the germs , and eliminate H-A at the same time. In consequence.it is also a natural health care goods which can delay slow death in addition to preventing disease by repressing and killing bacterium and is an essential component of the defence system in mammalian.Its value is more and more valueed by people , Nowadays, it has been widely used in keeping in store of dairy products fodder increasing , and on the clinical medicine actoperoxidase whose activity is correspond to 1/20 of cows represents about 0. 004% of the total protein in crude human milk. With increase in milk consumptions, the quality of milk is more concerned. It is through mammary gland bioreactor that milk of bovine and sheep may become similar to human milk, which has nutritional and medical function. The reconstructive milk is a promising product, which promotes children growth and development of nerve system and brain and immunity ability. Our research is to establish transgenic mouse model which highly expresses recombinant human lactoperoxidase in mammary gland.In order to establish the mouse model, pBCl-hLPO was constructed.Transgenic mice were generated by fertilized oocyte microinjection with the constructs.pBCl-hLPO expression vector is composed of Chicken beta-globin insulator, Goat beta-casein promoter, beta-casein exon K 2, beta-casein intron 1, the human lactoperoxidase encoding region and 3' termination region. As for pBOhLPO expression vector, 28 mice were born, and Spositive transgenic mice were identified by PCR and confirmed by Southern Blotting analysis of tail DNA. The integration rate is 17.86%. In order to examinate repression of human lactoperoxidase in the milk of transgenic mice ,based on the third class predict construction of human lactoperoxidase, Polypeptide CSAID KLDLSPWASV KN is synthetised in the design .united with KLH and then immunited white rabbits , examinated antibody got from the rabbits in the way of ELISA' s law , The effect price is up to 5 million times , that is to say , it may be used in Western blotting examinition the milk of transgenic mice completely...