Work with other systems utilizing pluripotent embryonal stem (ES) cell cultures for the formation of chimeric animals has succeeded in creating organisms in which foreign genes are expressed, as well as organisms in which endogenous genes have been altered. Dr. Barnes is interested in developing Zebra fish ES cell cultures for derivation of chimeric transgenic animals. He developed serum-free culture techniques allowing derivation of karyotypically normal late-stage mouse embryo cells that could not be derived in conventional serum-containing medium. Applying this approach to fish embryo cells, he found a mitogenic activity in trout embryo extract that was essential for the cultures and adapted transfection techniques to achieve expression of exogenous genes in fish cell cultures. He proposes to: (1) use the mitogenic activity in trout embryos to derive ES cell cultures and characterize the cells through transfection of exogenous genes, injection into developing embryos, and evaluation of contribution to transgenic chimeras; (2) apply this approach to haploid embryos to derive haploid ES cell cultures allowing alteration of endogenous genes.

Agency
National Institute of Health (NIH)
Institute
National Institute of Environmental Health Sciences (NIEHS)
Type
Research Project (R01)
Project #
5R01ES006011-03
Application #
2154837
Study Section
Cellular Biology and Physiology Subcommittee 1 (CBY)
Project Start
1992-09-30
Project End
1995-09-29
Budget Start
1994-09-30
Budget End
1995-09-29
Support Year
3
Fiscal Year
1994
Total Cost
Indirect Cost
Name
Oregon State University
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
053599908
City
Corvallis
State
OR
Country
United States
Zip Code
97339
Helmrich, A; Barnes, D (1999) Zebrafish embryonal cell culture. Methods Cell Biol 59:29-37
Helmrich, A; Barnes, D (1998) Animal cell culture equipment and techniques. Methods Cell Biol 57:3-17
Cheng, R; Bradford, S; Barnes, D et al. (1997) Cloning, sequencing, and embryonic expression of an N-ras proto-oncogene isolated from an enriched zebrafish (Danio rerio) cDNA library. Mol Mar Biol Biotechnol 6:40-7
Cheng, R; Ford, B L; O'Neal, P E et al. (1997) Zebrafish (Danio rerio) p53 tumor suppressor gene: cDNA sequence and expression during embryogenesis. Mol Mar Biol Biotechnol 6:88-97
Bradford, C S; Miller, A E; Toumadje, A et al. (1997) Characterization of cell cultures derived from Fugu, the Japanese pufferfish. Mol Mar Biol Biotechnol 6:279-88
Sun, L; Bradford, C S; Barnes, D W (1995) Feeder cell cultures for zebrafish embryonal cells in vitro. Mol Mar Biol Biotechnol 4:43-50
Sun, L; Bradford, C S; Ghosh, C et al. (1995) ES-like cell cultures derived from early zebrafish embryos. Mol Mar Biol Biotechnol 4:193-9
Bradford, C S; Sun, L; Barnes, D W (1994) Basic fibroblast growth factor stimulates proliferation and suppresses melanogenesis in cell cultures derived from early zebrafish embryos. Mol Mar Biol Biotechnol 3:78-86