RESUMO
A microfluidic platform is designed and fabricated to investigate the role of uncharacterized YOR060C (Sld7) protein in aging in yeast cells for the first time. Saccharomyces cerevisiae yeast cells are trapped in the series of C-shaped regions (0.5 nL) of COP (cyclo olefin polymer), PMMA (poly methylmethacrylate), or PS (polystyrene) microbioreactors. The devices are fabricated using hot embossing and thermo-compression bonding methods. Photolithography and electrochemical etching are used to form the steel mold needed for hot embossing. The cell cycle processes are investigated by monitoring green fluorescent protein (GFP) tagged Sld7 expressions under normal as well as calorie restricted conditions. The cells are loaded at 1 µL/min flowrate and trapped successfully within each chamber. The medium is continuously fed at 0.1 µL/min throughout the experiments. Fluorescent signals of the low abundant Sld7 proteins could be distinguished only on COP devices. The background fluorescence of COP is found 1.22 and 7.24 times lower than that of PMMA, and PS, respectively. Hence, experiments are continued with COP, and lasted for more than 40 h without any contamination. The doubling time of the yeast cells are found as 72 min and 150 min, and the growth rates as 9.63 × 10-3 min-1 and 4.62 × 10-3 min-1, in 2% glucose containing YPD and YNB medium, respectively. The product concentration (Sld7p:GFP) increased in accordance with cell growth. The dual role of Sld7 protein in both cell cycle and chronological aging needs to be further investigated following the preliminary experimental results.
