Expression of immunologically active porcine recombinant TGF-β1 precursor protein in baculovirus system

초록

In order to express recombinant porcine TGF-β1 protein in a baculovirus expression system the entire TGF-β1 gene containing extra amino acids at the N terminus was cloned into pFBa and pFBb of the Bac-To-Bac™ baculovirus expression system. One of the clones contained 106 extra amino acids and was designated pFBa-106TGF-β1, and the other had 28 extra amino acids and was designated pFBb-28TGF-β1. The orientation of the gene was identified with restriction enzyme mapping and PCR with internal TGF-β1 primers. Sf-9 cells were infected at a m.o.i. of 10 by the recombinant viruses generated from the two different plasmids. Both of the recombinant viruses produced precursor forms of TGF-β1 with expected sizes of 55 kD and 46.4 kD. These precursor forms of TGF-β1 reacted with a polyclonal antibody against human TGF-β1. No mature form of TGF-β1 protein was detected on SDS gels and an immunoblot indicated that TGF-β1 precursor is not properly processed in insect cells.

키워드