TY  - JOUR
T1  - Expression of Human Granulocyte-Macrophage Colony-Stimulating Factor in Stably-Transformed BmN and Sf-9 Cells and Silkworms by a Non-Transposon Vector
AU - Gong, Cheng Liang AU - Zhang, Hao Kun AU - Cao, Guang Li AU - Li, Yan Mei AU - Xue, Ren Yu 
JO  - Journal of Animal and Veterinary Advances
VL  - 11
IS  - 16
SP  - 2890
EP  - 2897
PY  - 2012
DA  - 2001/08/19
SN  - 1680-5593
DO  - javaa.2012.2890.2897
UR  - https://makhillpublications.co/view-article.php?doi=javaa.2012.2890.2897
KW  - Non-transposon vector
KW  -BmN cells
KW  -Sf-9 cells
KW  -Bombyx mori
KW  -hGM-CSF
KW  -transgene
AB  - This study aimed to explore the possibility of non-transposon vector mediated foreign gene expression in cultured insect cells and transgenic silk worms. To this end, the human Granulocyte-Macrophage Colony-Stimulating Factor (<I>hGM-CSF</I>) gene was inserted into the insect cell expression vector pIZT-V5-His to generate the recombinant vector pIZT-hGM-CSF. After transfection of BmN and Sf-9 cells with the pIZT-hGM-CSF vector, stably-transformed cells expressing the <I>hGM-CSF</I> gene were selected using the antibiotic zeocin at a final concentration of 300-400 &#956;g mL<SUP>-1</SUP>. Expression of a 22 kDa protein band representing hGM-CSF was detected in the transformed cells by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting. The expression levels of hGM-CSF in BmN and Sf-9 cells were determined by Enzyme-Linked Immunosorbent Assay (ELISA) to be about 0.7 and 0.3 ng/10<SUP>6</SUP> cells, respectively. The transgenic vector pIZT-hGM-CSF was transferred into silkworm eggs using sperm-mediated gene transfer. Transgenic silkworms were obtained after screening for the <I>gfp </I>gene and were verified by polymerase chain reaction, dot hybridization and Western blotting. The expression level of hGM-CSF determined by ELISA was about 4.7 ng g<SUP>-1</SUP> of freeze-dried silk glands in the G5 generation. These results suggest that heterologous genes can be integrated into cultured BmN and Sf-9 cells and into the silkworm genome using a non-transposon vector and can be expressed successfully.
ER  -