Application of the Sleeping Beauty system in Saanen goat fibroblast cells for establishing persistent transgene expression

The Sleeping Beauty (SB) transposon system is a promising new method for establishing persistent transgene expression in vivo. We applied the SB system for enhancing transgenesis in Saanen dairy goat fibroblast cells. We constructed a pKT2/CMV-EGFP-IRES-PURO vector and investigated the influence of...

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Bibliographic Details
Published in:Genetics and molecular research 2011-10, Vol.10 (4), p.3347-3355
Main Authors: Jiang, B C, Kaleri, H A, Zhang, H X, Chen, J, Liu, H L
Format: Article
Language:English
Subjects:
Animals
Cell Count
Cytomegalovirus - genetics
DNA - metabolism
DNA Transposable Elements - genetics
Fibroblasts - cytology
Fibroblasts - metabolism
Gene Expression
Gene Transfer Techniques
Genetic Vectors
Goats - genetics
Green Fluorescent Proteins - metabolism
Karyotyping
Molecular Weight
Transfection
Transgenes - genetics
Transposases - metabolism
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Summary:The Sleeping Beauty (SB) transposon system is a promising new method for establishing persistent transgene expression in vivo. We applied the SB system for enhancing transgenesis in Saanen dairy goat fibroblast cells. We constructed a pKT2/CMV-EGFP-IRES-PURO vector and investigated the influence of transposon and transposase vector ratios on transfection efficiency in the Saanen goat fibroblast cells. To enhance the SB system performance, we developed a new transfection technique (double-transfection method) for the SB system. The cultured cells were transfected with transposase and transposon vectors successively, with a 42-h interval. Consequently, the transposase and DNA donor (transposon vector) can interact, both at the highest level. Compared with the traditional transfection method, this new double-transfection method approximately doubled integration efficiency.
ISSN:1676-5680
1676-5680
DOI:10.4238/2011.October.25.4