Production of Transgenic Silver Sea Bream (Sparus sarba) by Different Gene Transfer Methods |
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Authors: | Jenn-Kan Lu Bo-Hua Fu Jen-Leh Wu Thomas T Chen |
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Institution: | (1) Department of Aquaculture, National Taiwan Ocean University, Keelung 202, Taiwan, Republic of China, CN;(2) Institute of Zoology, Academia Sinica, Nankang, Taipei 115, Taiwan, Republic of China, CN;(3) Biotechnology Center and Department of Molecular and Cell Biology, University of Connecticut, Storrs, CT 06269, U.S.A., US |
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Abstract: | We have been interested in developing convenient mass gene transfer methods for producing strains of silver sea bream (Sparus
sarba) with superior genetic traits for aquaculture. A transgene construct carrying rainbow trout growth hormone (rtGH) complementary
DNA driven by a common carp b-actin promoter was introduced into silver sea bream by electroporating the sperm with the rtGH
transgene and using the treated sperm to fertilize eggs stripped from mature females. The presence of the GH transgene in
presumptive transgenic individuals was detected by polymerase chain reaction (PCR) analysis. Between 56% and 70% of the animals
carried the GH transgene. We refer to this method as sperm-mediated gene transfer (SMGT). Since the handling stress of stripping
gametes from female sliver sea bream brood fish could cause severe mortality, an alternative gene transfer method would be
highly desirable. We developed a liposome-based method to transfer the GH transgene into the fish. This method, referred as
testis-mediated gene transfer (TMGT), involves injecting the liposome-transgene mixture into the gonads of male sea bream
at least 48 hours before spawning. The males were mated to reproductively active females, and fertilized eggs were collected
for further incubation. Between 59% and 76% of the hatched fry were found by PCR analysis to carry the rtGH transgene. The
efficiency of gene transfer was improved more than 80% by injecting multiple doses of the liposome-transgene mixture into
the gonads of treated males. Results of Southern blot analysis of DNA isolated from PCR-positive animals showed that the transgene
was integrated into the host genome and could be transmitted to its offspring. The rtGH transgene was expressed in many of
the rtGH-transgenic fish. Several P1 GH-transgenic silver sea bream exhibited significant growth enhancement compared with
nontransgenic controls. Our studies showed that faster-growing silver sea bream could be produced by a variety of mass gene
transfer technologies. These gene transfer technologies would be of great value to aquaculture. |
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