优化用于野生型莱茵衣藻转化的液滴电穿孔法。

Optimization of the droplet electroporation method for wild type Chlamydomonas reinhardtii transformation.

机构信息

Department of Chemical Engineering, Pukyong National University, 365 Sinseon-Ro, Nam-Gu, Busan 48547, South Korea.

出版信息

Bioelectrochemistry. 2019 Apr;126:29-37. doi: 10.1016/j.bioelechem.2018.11.010. Epub 2018 Nov 22.

DOI:10.1016/j.bioelechem.2018.11.010

Abstract

We performed the transformation of a wild type Chlamydomonas reinhardtii by optimizing previously developed droplet EP method. For more effective and faster optimization, we used DNA dying fluorescent molecule (Yo-Pro-1) for finding optimal EP conditions instead of using protein expression based evaluation method. By examining wider range of electrical parameter space together with the analysis of total current flow of EP process, we found optimal EP conditions. The obtained optimal EP conditions were verified by CFP transgene expression experiments. By applying the optimal EP conditions to the transformation of C. reinhardtii, we obtained transformants and analyzed them using PCR. Finally, implications and future work are discussed.

摘要

我们通过优化先前开发的液滴 EP 方法对野生型莱茵衣藻进行了转化。为了更有效和快速地进行优化，我们使用 DNA 死亡荧光分子（Yo-Pro-1）来寻找最佳 EP 条件，而不是使用基于蛋白质表达的评估方法。通过检查更广泛的电参数空间以及分析 EP 过程中的总电流，我们找到了最佳的 EP 条件。通过 CFP 转基因表达实验验证了获得的最佳 EP 条件。通过将最佳 EP 条件应用于莱茵衣藻的转化，我们获得了转化体，并通过 PCR 对其进行了分析。最后，讨论了其意义和未来的工作。