Kim Tae-Wan, Keum Jung-Won, Oh In-Seok, Choi Cha-Yong, Park Chang-Gil, Kim Dong-Myung
School of Chemical and Biological Engineering, College of Engineering, Seoul National University, Seoul 151-742, Republic of Korea.
J Biotechnol. 2006 Dec 1;126(4):554-61. doi: 10.1016/j.jbiotec.2006.05.014. Epub 2006 May 27.
In this study, as a part of our efforts to improve the robustness and economical feasibility of cell-free protein synthesis, we developed a simple method of preparing the cell extracts used for catalyzing cell-free protein synthesis reactions. We found that the high-speed centrifugation, pre-incubation, and dialysis steps of the conventional procedures could be omitted without losing the translational activity of the resulting cell extract. Instead, a simple centrifugation step at low speed (12,000 RCF for 10 min) followed by a brief period of incubation was sufficient for the preparation of an active extract to support cell-free protein synthesis with higher productivity and consistency. Compared to the present standard procedures for the preparation of the S30 extract, the overall cost of the reagents and processing time were reduced by 80 and 60%, respectively.
在本研究中，作为我们提高无细胞蛋白质合成的稳健性和经济可行性的一部分工作，我们开发了一种制备用于催化无细胞蛋白质合成反应的细胞提取物的简单方法。我们发现，传统方法中的高速离心、预孵育和透析步骤可以省略，而不会损失所得细胞提取物的翻译活性。相反，简单的低速离心步骤（12,000相对离心力，10分钟），随后进行短暂孵育，就足以制备出活性提取物，以更高的生产率和一致性支持无细胞蛋白质合成。与目前制备S30提取物的标准方法相比，试剂的总成本和处理时间分别降低了80%和60%。
