Department of Applied Research, National Marine Biodiversity Institute of Korea, Jangsan-ro 101-75, Seocheon-gun, Chungcheongnam-do, 33662, South Korea.
Mol Biotechnol. 2020 May;62(5):297-305. doi: 10.1007/s12033-020-00249-9.
Microalgal chloroplasts have a substantial potential as a sustainable alternative to conventional hosts for recombinant protein production, due to their photosynthetic ability. However, realization of microalgal chloroplast as a platform for the production of recombinant proteins has suffered from difficulties in genetic manipulation and development of molecular tools, including reporter proteins. Here, we investigated the suitability of a fluorescent protein, mCherry, as a reporter for quantitative in vivo monitoring of gene expression in the chloroplast of Chlamydomonas reinhardtii. By analyzing cell growth, the fluorescence intensity of a mCherry-expressing strain, as well as auto-fluorescence, under different photoautotrophic culture conditions, we demonstrated a strong correlation between the fluorescence intensity of mCherry expressed in the chloroplast and its protein expression level. In addition, we found that the supply of CO and light energy can be an important factor for the synthesis of recombinant proteins in the microalgal chloroplast. Our results identified mCherry as a reliable and quantitative reporter for the study of gene expression in chloroplasts, which is essential for the biotechnological application of microalgal chloroplasts and for improved production of valuable recombinant proteins.
微藻叶绿体由于其光合作用能力，具有作为传统宿主的替代物来生产重组蛋白的巨大潜力。然而，由于遗传操作和分子工具（包括报告蛋白）的开发困难，微藻叶绿体作为生产重组蛋白的平台的实现受到了阻碍。在这里，我们研究了荧光蛋白 mCherry 作为 Chlamydomonas reinhardtii 叶绿体中基因表达定量体内监测的报告蛋白的适用性。通过分析细胞生长、mCherry 表达菌株的荧光强度以及在不同自养培养条件下的自发荧光，我们证明了在叶绿体中表达的 mCherry 的荧光强度与其蛋白表达水平之间存在很强的相关性。此外，我们发现 CO 和光能的供应可能是微藻叶绿体中重组蛋白合成的一个重要因素。我们的结果确定 mCherry 是研究叶绿体中基因表达的可靠和定量报告蛋白，这对于微藻叶绿体的生物技术应用以及提高有价值的重组蛋白的生产至关重要。
