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  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.
College of Animal Sciences and Technology, Nanjing Agricultural University, Nanjing, China.
B C
BC
Jiang
H A
HA
Kaleri
H X
HX
Zhang
J
J
Chen
H L
HL
Liu
D000818:Animals; D002452:Cell Count; D003587:Cytomegalovirus / Q000235:genetics; D004247:DNA / Q000378:metabolism; D004251:DNA Transposable Elements / Q000235:genetics*; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism*; D015870:Gene Expression; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D006041:Goats / Q000235:genetics*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D007621:Karyotyping; D008970:Molecular Weight; D014162:Transfection; D019076:Transgenes / Q000235:genetics*; D019895:Transposases / Q000378:metabolism*
D004251:DNA Transposable Elements; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D004247:DNA; D019895:Transposases