title: Effect of exogenous DNA microinjection on early development response of the seabream (Sparus aurata).
abstract: DNA transfer techniques allow genetic manipulation of commercial fish. However, marine species have received little attention because of their difficult zootechnical requirements. The seabream (Sparus aurata) has become one of the most important species in the aquaculture of Mediterranean countries, and the development of suitable DNA transfer procedures represents a main step in its genetic improvement. To assess the response of the seabream to exogenous DNA, naturally fertilized eggs were injected with the plasmids pCMV-CAT, pCMVTklacZ, and pEGFP-N1, in supercoiled and linearized forms. Embryo and larval survival, DNA fate, and reporter gene expression were analyzed during early development. The survival results indicate that microinjection is an effective transfer method in spite of the unfavorable conditions. Linearized plasmids were more efficiently polymerized than supercoiled ones; however, no significant differences were detected either in their persistence or in their expression levels. Reporter gene expression was initiated after mid-blastula transition. The duration of transient expression varied between the promoter-gene combinations, and no integration of transgenes into fish chromosomes was detected. Results suggest that the main factor affecting the persistence and expression of DNA seems to be related to developmental processes. Among the markers used, CAT proved to be the most sensitive, but GFP had obvious methodologic advantages over the spatial marker lacZ. The usefulness of GFP for diagnosis of transgenesis is enhanced by the transparency of embryos and larvae in S. aurata.
authors.affiliation: Departamento de Genética, Facultad de Ciencias, Universidad de Málaga, 29071 Málaga, Spain.
authors.forename: S
authors.initials: S
authors.lastname: García-Pozo
authors.forename: J
authors.initials: J
authors.lastname: Béjar
authors.forename: M
authors.initials: M
authors.lastname: Shaw
authors.forename: M C
authors.initials: MC
authors.lastname: Alvarez
mesh_terms: D000818:Animals; D015500:Chloramphenicol O-Acetyltransferase / Q000096:biosynthesis / Q000235:genetics; D004247:DNA / Q000008:administration & dosage; D004625:Embryo, Nonmammalian / Q000502:physiology; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D007814:Larva; D008845:Microinjections; D010473:Perciformes / Q000254:growth & development*; D011994:Recombinant Proteins / Q000096:biosynthesis; D001616:beta-Galactosidase / Q000096:biosynthesis
chemical_list: D011994:Recombinant Proteins; D004247:DNA; D015500:Chloramphenicol O-Acetyltransferase; D001616:beta-Galactosidase