Production of recombinant goldfish growth hormone I in a baculovirus expression system.
The cDNA sequence encoding the growth hormone (GH) I gene of goldfish (gf GH-I) was cloned into the pSXIVVI+ X3 baculovirus transfer vector. This transfer vector, without the initiation codon (ATG) and using a SynXIV promoter to activate transcription, was constructed for a baculovirus expression system. The recombinant transfer vector with the gf GH-I insert was used to produce the recombinant baculovirus TnNPV-SX+ gf GH-I 21a. Both in vivo and in vitro approaches were used to test the expression of growth hormone I gene using TnNPV-SX+ gf GH-I 21a. For in vivo studies, larvae of Trichoplusia ni were infected with the recombinant baculovirus. Four days later, growth hormone-like immunoreactivity was detected in the hemolymph of these infected larvae. For in vitro studies, insect Spodoptera frugiperda 9(Sf9) cells were infected with TnNPV-SX+ gf GH-I 21a. After incubation for at least 24 hours, growth hormone-like immunoreactivity was detected in Sf9 insect cells as well as in the culture medium. Western blotting analysis of larval hemolymph and Sf9 cell contents after viral infection revealed a protein band of 22.5 kDa immunoreactive to goldfish growth hormone antiserum. This is consistent with the predicted molecular weight deduced from the cDNA of goldfish growth hormone I gene. These results, taken together, suggest that the baculovirus expression system can be used to produce the recombinant growth hormone of a fish species.
Lin
G
G
State Key Laboratory for Biological Control, Zhongshan University, Guangzhou, China.
Wang
X
X
Long
Q
Q
Pang
Y
Y
Wong
A O
AO
Yu
K
K
D000818:Animals; D002460:Cell Line; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D015870:Gene Expression; D005822:Genetic Vectors; D006054:Goldfish; D013006:Growth Hormone / Q000096:biosynthesis* / Q000235:genetics; D007814:Larva; D009036:Moths; D017924:Nucleopolyhedroviruses; D018411:Spodoptera / Q000166:cytology
D013006:Growth Hormone; C024408:endodeoxyribonuclease SacI; D015252:Deoxyribonucleases, Type II Site-Specific