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Abstract
Sperm-mediated gene transfer was used to produce transgenic rabbits that expressed the porcine growth hormone gene under the control of a metallothionein promoter. The gene that encodes the selectable marker green fluorescent protein (GFP) was inserted downstream of the transgene. After lipofectin-mediated gene transfer into sperm cells and after subsequent in vitro fertilization using the transfected sperm cells, 32% of the cultured blastocysts exhibited bright green fluorescence when stimulated with blue light. Of the 74 adult rabbits and five fetal rabbits (age, gestational day 15), 2 fetuses and 29 rabbits were GFP-positive as indicated by PCR analysis. Southern blot analysis of their genomic DNA showed that 13 of 21 GFP-positive rabbits were transgenic. GFP expression was observed in different tissues of transgenic rabbits and the growth rate of four GFP-positive rabbits was greater than that of controls. PCR analysis showed that one of six F1 offspring was transgenic. These results suggest that lipofectin-mediated gene transfer into sperm cells can be used to efficiently produce transgenic rabbits.
ACKNOWLEDGMENTS
We would like to thank Ms. L.M. Ren, Dr. N. Li, Prof. S.E. Zhu, Dr. S.M. Zeng, Ms. H.Y. Hu, Dr. X.J. Liu, Ms. X. Ding, Dr. X. J. Chen and Dr. J.X. Liu for their kind gifts of materials and their skillful technical assistance. Special thanks to Mr. Z. Yang for taking cares of the rabbits and for providing technical assistance. Also, we thank to Drs. Robert. J. Wall and Dr. Angela McArthur for their kindly advice and assistance in the revision of this manuscript.