Construction of transposon-mediated baculovirus vector and expression of green fluorescent protein in insect cells and larvae

A transposon-shuttle vector Hanpvid was constructed by using wild-type geno mic DNA from Heliothis armigera nuclear polyhedrosis virus (HaNPV). It coul d replicate in E. coli cells as a large plasmid and remain infectious when being induced into insect cells. Hanpvid comprises HaNPV DNA and a transpos on cassette which includes a miniF replicon, a kanamycin resistance gene (k an), lacZ alpha and an attachment site for Tn7 (attTn7). Recombinant virus rHa-FaGP was obtained after transposition of a donor plasmid carrying green fluorescent protein gene (gfp) and polyhedrin gene (ocu) into attTn7. SDS- PAGE analysis shows that both gfp and ocu genes were highly expressed in He liothis armigera cells. Green Hemolymphocytes can be seen under a fluoresce nt microscope 4 d after recombinant virus rHa-FaGP infected the third-insta r larvae. The infected larvae show strong green fluorescence 6 d post infec tion.