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| Optimization of Electroporation Condition for Buffalo Fetal Fibroblast Cells |
| LUO Chan, GONG Yun, REN Yan-ping, YANG Su-fang, RUAN Qiu-yan, GUAN Xiao-mei, JIANG Jian-rong, SHI De-shun |
| Animal Reproduction Institute, State Key Laboratory for Conservation and Utilizationof Subtropical Agro-bioresources, Guangxi University, Nanning 530005, China |
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Abstract A significant aspect of applying somatic cell nuclear transfer (SCNT) technology to generate transgenic animal is the in vitro method utilized to obtain viable donor nuclei for cloning procedures. Therefore, the present study was undertaken to explore the optimal electroporation parameters for transfection of buffalo fetal fibroblasts (BFFs) and determine the ability of these electroporation parameters to produce stably transfected fibroblast colonies. First, the BFFs was obtained by tissue mass inoculation method and were confirmed of possessed a normal diploid karyotype. And then, the transfection efficiency of different voltage (150V, 200V, 250V, and 300V), different pulse length (5ms, 10ms and 15ms) and different cells passage were compared. Results show that one 10ms pulse of 300V can introduced DNA into the cells more effectively, and the passage of cells also has remarkable influence on transfection efficiency. Finally, the utilization of these conditions can produced numerous transgenic fibroblast colonies following G418 selection. This study provides a suitable electroporation condition for introducing DNA into BFFs and improves the efficiency of transgenic SCNT technology.
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Received: 03 June 2013
Published: 25 September 2013
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