Abstract
BackgroundTransposon-mediated, non-viral gene delivery is a powerful tool for generating stable cell lines and transgenic animals. However, as multi-copy insertion is the preferred integration pattern, there is the potential for uncontrolled changes in endogenous gene expression and detrimental effects in cells or animals. Our group has previously reported on the generation of several transgenic cattle by using microinjection of the Sleeping Beauty (SB) and PiggyBac (PB) transposons and seeks to explore the long-term effects of this technology on cattle.ResultsTransgenic cattle, one female (SNU-SB-1) and one male (SNU-PB-1), reached over 36 months of age with no significant health issues and normal blood parameters. The detection of transgene integration and fluorescent signal in oocytes and sperm suggested the capacity for germline transmission in both of the founder animals. After natural breeding, the founder transgenic cow delivered a male calf and secreted milk containing fluorescent transgenic proteins. The calf expressed green fluorescent protein in primary cells from ear skin, with no significant change in overall genomic stability and blood parameters. Three sites of transgene integration were identified by next-generation sequencing of the calf’s genome.ConclusionsOverall, these data demonstrate that transposon-mediated transgenesis can be applied to cattle without being detrimental to their long-term genomic stability or general health. We further suggest that this technology may be usefully applied in other fields, such as the generation of transgenic animal models.
Highlights
Transposon-mediated, non-viral gene delivery is a powerful tool for generating stable cell lines and transgenic animals
There was no significant difference in various serum chemical parameters, including alanine transaminase (ALT), aspartate transaminase (AST), and blood urea nitrogen (BUN), between the transgenic animals and the reference (Additional file 1)
Germline transmission of the transgene To assess the germline transmission in sperm of transgenes introduced by transposons, sperm was harvested from SNU-PB-1 after puberty using manual ejaculation and cryopreserved to create frozen semen stocks
Summary
Transposon-mediated, non-viral gene delivery is a powerful tool for generating stable cell lines and transgenic animals. Of the several DNA transposons available, Sleeping Beauty (SB) and PiggyBac (PB) transposons have been the most widely used to deliver exogenous genes into cell lines and to generate transgenic animals. In. The transposon system is valuable as it mediates both the efficient genomic integration and stable expression of transgenes in target cells or animals. Yum et al BMC Genomics (2018) 19:387 backbone DNA [12]. This system can be used in conjunction with various non-viral delivery systems, such as chemical-based transfection, electroporation or microinjection. In comparison to viral delivery and transgene integration systems, transposon systems are more secure and safe to use [13, 14] and this has led to NIH-OBA and FDA approval for testing of the SB transposon in humans [15,16,17]
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