A microfluidic device for electrofusion of two plant cells

In this paper, we have electrofused two cells which are P. japonicum and G. littoralis at the ‘Lab on a Chip’ level. The microchannel platform was fabricated using poly(dimethylsiloxane)(PDMS) via the conventional softlithographic procedure, and the microelectrode patterns (we ebeam evaporated the titanium (as seed layer) and the gold with the thickness of 200 Å and 3000 Å and patterned by the chemical etching) were created on the surface of glass. Both the PDMS platform and electrode glass were aligned and combined via the oxygen plasma-treatment. The channel has two inlets and two types of cells were slowly introduced by using the syringe pumps and we stopped when the cells were positioned around the electrodes. We have applied 1 - 2 MHz AC field (Amplitude: 8 - 10 p-p) with rectangular wave shape to the microelectrodes to form the pearl-chain between the electrodes. Then, the cell-membranes contact closely each other. For the cell fusion, we again applied short DC pulse (amplitude: 250V/mm, duration: 10 – 100 ms) to the microelectrodes. A little pore was generated and finally two cells were fused to single cell. The fused cells were cultured in Nitsch medium containing growth regulators and some cells were cultured. We will investigate their viability with FDA (fluorescein diacetate). This results may encourage the maximum yield of 1: 1 heterologous fusion under the micro environment.