[Studies of the clone and cell expression of human lactoferritin gene].

ABSTRACT

In this paper,we directly cloned 2.366 Kb cDNA sequence of human lactoferrin gene and 800 bp 5'flank regulatory sequence of beta/lactoglobulin gene from goat by PCR,then connected them with the expression vector pLNCX. The recombinant plasmid pLNCXHLF containing human lactoferrin gene cDNA was transfected into mice mammary tumor cell line MA/782 after liposome transinfection. Positive single clone cells were selected with G418 and by PCR. After proliferating,the transfected cells immobilized and cultured in sodium alginate were induced by hormone. The result of Western blotting analysis on cultured cell supernatant shows that transfected cells can express the exogenic gene and secrete hLF protein, whose MW is 34 KD. The highest amount detected by ELISA reached 65 mg/l medium/10(5) cells. The result of antibacterial experiment indicates that the recombinant hLF protein has the effect of inhibiting E.coli proliferation;moreover,its activity is superior to the commercial available hLF's.