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Abstract The production of recombinant human pharmaceuticals in the milk of transgenic farm animals has many advantages when compared with microbial bioreactors or animal cell bioreactors. At the same time, somatic cell nuclear transfer (SCNT) has provided an alternative avenue to the production of transgenic animals. The purpose of this study was to establish human lactoferrin cDNA transgenic donor cell lines and to prepare competent donor cells for the production of transgenic animals by SCNT. Human lactoferrin cDNA was obtained by RT-PCR while 6.5 kb caprine β-casein regulatory elements were amplified by Long and Acute PCR. Subsequently the two fragments were inserted into the corresponding site of the plasmid pEGFP-C1, resulting in the mammary specific expression vector p6.5hLF-EGFP. Then the caprine fetal fibroblast cells were transfected with p6.5hLF-EGFP by LipofectamineTM-2000 and selected by G418 for 3 to 4 weeks. The G418 resistant transfectants were identified by PCR and EGFP detection. The results indicated that the transgene was stably integrated into the open region of the chromatin of G418 resistant fibroblast cells. This study may provide competent transgenic donor cells for the production of transgenic animals by SCNT and improve the efficiency of transgenic cloning.
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Received: 03 October 2007
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