Transcriptomic analysis of silver-resistant Saccharomyces cerevisiae mutant obtained by evolutionary engineering

Silver-resistant Saccharomyces cerevisiae mutant was obtained by evolutionary engineering method. Briefly, genetic diversity in reference strain, CEN.PK.113-7D, was increased by ethyl methane sulfonate (EMS)-mutagenesis. The mutant population was passaged several times in gradually increasing silver stress. Several mutant individuals were selected from the final population. Among selected mutant individuals, one of them was much more resistant to silver stress than the reference strain, called as 2E. Whole-genome transcriptomic analysis was performed to identify the silver resistance mechanisms in the silver-resistant mutant strain. Silver-resistant Saccharomyces cerevisiae (2E) mutant strain and reference strain were grown in yeast minimal medium. When cultures’ Optical Density (OD600) reached to about 1, which correspond to the logarithmic growth phase of the yeast cells, total RNA isolation was carried out by using RNeasy Mini Kit (Qiagen). The experiment was performed triplicates. Whole-genome microarray analysis was carried out by using Agilent DNA Microarray Technology.

本研究通过进化工程方法获得抗银酿酒酵母（Saccharomyces cerevisiae）突变株。具体流程如下：以参考菌株CEN.PK.113-7D为出发菌株，通过甲基磺酸乙酯（ethyl methane sulfonate，EMS）诱变提升其遗传多样性；将诱变获得的突变体群体在逐步升高的银胁迫条件下连续传代培养多代；从最终培养群体中筛选得到多株突变体个体，其中一株对银胁迫的抗性显著高于参考菌株，将其命名为2E。
为解析该抗银突变株的银抗性分子机制，本研究开展全基因组转录组分析：将抗银酿酒酵母（2E）突变株与参考菌株接种于酵母基本培养基中培养，待培养液光密度（Optical Density，OD600）值达到约1.0（对应酵母细胞的对数生长期）时，使用RNeasy Mini Kit（Qiagen）提取总RNA。实验设置三次生物学重复，采用Agilent DNA微阵列技术完成全基因组微阵列分析。