Synthesis of human erythropoietin in vivo in the oviduct of laying hens by localized in vivo gene transfer using electroporation.
In order to synthesize human erythropoietin protein in the oviduct of laying hens, localized in vivo gene transfer was attempted by using electroporation. In Experiment 1, transcriptional activities were compared by using four viral and cellular promoters, i.e., the 1.35-kbp long ovalbumin promoter, SV40 early promoter, Rous sarcoma virus long terminal repeat (RSV LTR), and the miw promoter, which is a hybrid of RSV LTR and chicken beta-actin promoter. These promoters were fused immediately upstream to the chloramphenicol acetyltransferase reporter gene. The results of chloramphenicol acetyltransferase activity showed that the miw promoter was the strongest, followed by SV40, RSV LTR, and the ovalbumin promoter in decreasing order. The intensity of the miw promoter was 250 times as high as that of the ovalbumin promoter. In Experiment 2, plasmid DNA containing the human erythropoietin gene, driven either by the ovalbumin promoter or the miw promoter, was transfected in vivo, and the production of human erythropoietin protein was detected by ELISA. The results indicated that the synthesis of human erythropoietin protein was attained in the chicken oviduct, and its concentration was higher when driven by the miw promoter than the ovalbumin promoter.
Ochiai
H
H
Department of Biological Resources and Environmental Sciences, School of Agricultural Sciences, Nagoya University, Japan.
Park
H M
HM
Nakamura
A
A
Sasaki
R
R
Okumura
J I
JI
Muramatsu
T
T
D000818:Animals; D002645:Chickens / Q000235:genetics / Q000502:physiology*; D018076:DNA, Complementary / Q000235:genetics; D018274:Electroporation / Q000662:veterinary; D004921:Erythropoietin / Q000096:biosynthesis* / Q000235:genetics; D005260:Female; D018014:Gene Transfer Techniques / Q000662:veterinary*; D006801:Humans; D010057:Oviducts / Q000378:metabolism*; D010957:Plasmids; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics
D018076:DNA, Complementary; D012333:RNA, Messenger; D011994:Recombinant Proteins; D004921:Erythropoietin