High-Efficient Transfection of Human Embryonic Stem Cells by Single-Cell Plating and Starvation.

Liu H¹, Ren C¹, Zhu B¹, Wang L¹, Liu W¹, Shi J¹, Lin J¹, Xia X², Zeng F³, Chen J¹, Jiang X⁴.

Author information

11 Key Laboratory for Carcinogenesis of Chinese Ministry of Health, Cancer Research Institute, Collaborative Innovation Center for Cancer Medicine, School of Basic Medical Science, Central South University , Changsha, People’s Republic of China .22 Department of Gynecology and Obstetrics, the Second Xiangya Hospital, Central South University , Changsha, People’s Republic of China .33 Department of Gynecology and Obstetrics, the Third Xiangya Hospital, Central South University , Changsha, People’s Republic of China .44 Department of Neurosurgery, Xiangya Hospital, Central South University , Changsha, People’s Republic of China .

Abstract

Nowadays, the low efficiency of small interfering RNA (siRNA) or plasmid DNA (pDNA) transfection is a critical issue in genetic manipulation of human embryonic stem (hES) cells. Development of an efficient transfection method for delivery of siRNAs and plasmids into hES cells becomes more and more imperative. In this study, we tried to modify the traditional transfection protocol by introducing two crucial processes, single-cell plating and starvation, to increase the transfection efficiency in hES cells. Furthermore, we comparatively examined the transfection efficiency of some commercially available siRNA or pDNA transfection reagents in hES cells. Our results showed that the new developed method markedly enhanced the transfection efficiency without influencing the proliferation and pluripotency of hES cells. Lipofectamine RNAiMAX exhibited much higher siRNA transfection efficiency than the other reagents, and FuGENE HD was identified as the best suitable reagent for efficient pDNA transfection of hES cells among the tested reagents.