earticle

영어

Porcine spermatogonial stem cells (SSCs) are an useful model for a successful gene modification of sperm compatible, and various gene delivery systems to transport specific genes into the cytoplasm of porcine SSCs have been introduced for successfully producing transgenic sperms. However, each gene delivery system still suffers from low transfection efficiency and transfected gene expression in porcine SSCs, resulting in making it difficult to produce transgenic offspring. Accordingly, in order to determine type of promoters optimized to porince SSCs, impact of each Cytomegalovirus (CMV), Simian virus 40 (SV40), and chicken b-actin promoter on transfection efficiency of porcine SSCs was investigated using electroporation. For these, the highest transfection efficiency and cell viability were observed in porcine SSCs transfected with 1 μg of the each transgenic vector with a single electric pulse from an electroporator at a voltage of 200 V and a capacitor setting of 500 μF in a 0.4 cm cuvette containing 200μL of standized SSCs medium. Cell viability was measured by trypan blue exclusion assay, and analysis of transfection efficiency and intensity were conducted as counting SSCs expressing enhanced green fluorescent protein (EGFP) and measuring EGFP fluorescent intensity under flow cytometer system. As the results, the highest EGFP expression efficiency and intensity were observed in porcine SSCs introduced with CMV promoter-containing vectors, compared to those with SV40 and chicken b-actin protmoter- containing vectors. However, cell viability showed no significant difference among porcine SSCs experiencing transfection of SV40, CMV and chicken b-actin promotercontaining vector. In conclusion, we found that CMV promoter could effectively stimulate expression of genes transfected to porcine SSCs. This will contribute advances in further research related in SSCs transplantation and production of transgenic animals.