title: Characterization of transgene integration loci in transformed Madin Darby bovine kidney cells.
abstract: Generation of transgenic animals is invaluable for both basic and applied research, as it enables the production of biologically active proteins and immunologically compatible organs for xenotransplantation, improvement of livestock production traits, and establishment of animal models of human disease. However, transgene expression is commonly highly variable, even among cell lines independently transformed with the same construct. Consequently, a great number of transfections and screening is needed to achieve transgenic cell lines showing expected phenotype. In this study, we sequenced transgene-host DNA junctions of transgene integration loci in 26 independently transformed Madin Darby bovine kidney (MDBK) cell lines produced by direct liposome transfection. For 15 rescued clones, sequences were of sufficient length and quality to determine unambiguously the position of integration in the bovine genome. Results revealed that transgenes were integrated in 12 different chromosomes, suggesting that there was no chromosomal preference for insertion of exogenous DNA. Most integration events occurred into transcriptionally active regions. No correlation was found between integration into transcribed sequences and the expression level of the beta-gal transgene.
authors.affiliation: Embrapa Recursos Genéticos e Biotecnologia, Parque Estação Biológica, W5 Norte, 70.770-900, Brasília, DF, Brazil.
authors.forename: Sharon F C
authors.initials: SF
authors.lastname: Lisauskas
authors.forename: Elíbio L
authors.initials: EL
authors.lastname: Rech
authors.forename: Francisco J L
authors.initials: FJ
authors.lastname: Aragão
mesh_terms: D000818:Animals; D001483:Base Sequence; D002417:Cattle; D018929:Cell Culture Techniques / Q000379:methods; D004247:DNA / Q000378:metabolism; D015870:Gene Expression; D005818:Genetic Engineering / Q000379:methods*; D005821:Genetic Techniques; D007668:Kidney / Q000166:cytology*; D008969:Molecular Sequence Data; D014158:Transcription, Genetic; D014162:Transfection; D019076:Transgenes; D001616:beta-Galactosidase / Q000378:metabolism
chemical_list: D004247:DNA; D001616:beta-Galactosidase