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KMID : 0545120090190080781
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Journal of Microbiology and Biotechnology
2009 Volume.19 No. 8 p.781 ~ p.786
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Enhanced Delivery of siRNA Complexes by Sonoporation in Transgenic Rice Cell Suspension Cultures
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Cheon Su-Hwan
Lee Kyoung-Hoon
Kwon Jun-Young
Choi Sung-Hun
Song Mi-Na
Kim Dong-Il
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Abstract
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Small interfering synthetic double-stranded RNA (siRNA) was applied to suppress the expression of the human cytotoxic- T-lymphocyte antigen 4-immunoglobulin (hCTLA4Ig) gene transformed in transgenic rice cell cultures. The sequence of the 21-nucleotide siRNA was deliberately designed and synthesized with overhangs to inactivate the expression of hCTLA4Ig. The chemically synthesized siRNA duplex was combined with polyethyleneimine (PEI) at a mass ratio of 1:10 (0.33 ￥ig siRNA:3.3 ￥ig PEI) to produce complexes. The siRNA complexes (siRNA+PEI) were labeled with Cy3 in order to subsequently confirm the delivery by fluorescent microscopy. In addition, the cells were treated with sonoporation at 40 kHz and 419 W for 90 s to improve the delivery. The siRNA complexes alone inhibited the expression of hCTLA4Ig to 45% compared with control. The siRNA complexes delivered with sonoporation downregulated the production of hCTLA4Ig to 73%. Therefore, we concluded that the delivery of siRNA complexes into plant cells could be enhanced successfully by sonoporation.
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KEYWORD
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Polyethyleneimine, transgenic plant cell cultures, siRNA, sonoporation, hCTLA4Ig
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