Study On Expression Of Human Lactoferrin CDNA In The Transgenic Mammalian Gland Under Control Of Casein Gene Of Bovine Or Goat

This paper was studied on the expression of Human Lactoferrin cDNA in the mammary gland of transgenic goats by using fusion gene constructed with regulative elements of casein gene and LTF cDNA. Then the transgenic goats and mice were produced by microinjecting the fusion gene into pronuclear of fertilized eggs. The biological reactor was made from the transgenic mammary specific- expression goats.The human lactoferrin cDNA (hLTF) was amplified from the templet of human mammary cDNA by PCR with high fidelity Taq enzyme. After cloned and sequenced, the hLTF cDNA sequence was proved correct, hLTF was expressed in the microorganism using PGEX-4T-3 vector. .Subsequently, we amplified 5'-regulative sequence(2.3Kb) and 3'- regulative sequence (1.5Kb) of bovine as 1-casein gene from bovine genome DNA with PCR, connected the regulative sequence with PGEM vector .then cloned the regulative sequence A; 4Kb bovine genome DNA of 5'-upstream regulative sequence was spliced to the sequence A, regulative sequence B was formed; the regulative sequence C of 3 -casein gene had been constructed. Six fusion gene constructs (PCN1, PCN2,261, AF203.P1238, PF84) were obtained by constructing three regulative sequences (A3,C) and two Human Lactoferrin gene cDNA segments(2.18Kb,2.32Kb). These gene constructs were injected into mammary gland tissue of ewe respectively about 10 days before secreting milk and were expressed transiently. Results showed that there were different express levels, especially, expression of PCN2, 261, AF203 showed high expression, it was 88ng/ml, 144ng/ml, 63ng/ml respectively. Then transgenic mammalian was produced with high expressing construct "261 "by microinjecting the fusion gene into pronuclear of fertilized eggs. 148 fertilized eggs was microinjected, 16 lambs were borned, finally, we obtained 4 transgenic goats.