Optimization of Polyethylenimine to Improve the Transfection Efficiency of Sheep Spermatogonial Stem Cell

Background: Spermatogonial stem cells (SSCs) have the capability to self-renew and to contribute genes to the next generation. Genetic modification of these cells would provide an opportunity to study the gene function, biochemical mapping, mutational analysis, production of recombinant proteins and generation of transgenic animals. The aim of this study was to verify and optimize the transfection efficiency of sheep testicular cells including (SSCs) via polyethylenimine (PEI, branched with Mw 25 kD), one of potent non-viral gene delivery carrier that show promise in stem cell genetic modification.Methods: Following isolation and propagation of one-month lamb testicular cells, the effect of different polymer/plasmid DNA weight ratios (C/P; 2.5 and 5) in various incubation times (4, 24, 48, and 72h) were evaluated on transfection efficiency, viability rate and mean fluorescent intensity (MFI) of sheep testicular cells.Result: The highest transfection efficiency with appropriate cell viability was obtained 48h after culture initiation in C/P 2.5. The most MFI was demonstrated in SSCs among different testicular cell types.Conclusion: PEI as a cationic polymer is an efficient method for transduction of plasmid vector into sheep SSCs. This improvement could greatly boost the application potential of its in clinical cell-based gene therapies.

کلیدواژه ها:

Transfection ، Polyethylenimine ، Spermatogonial Stem Cell