The use of Chlamydomonas for biofuel and biopharmaceutical production has been anticipated. However, the genetic engineering technology for Chlamydomonas is not as advanced as that for other organisms. Here, we established transgenic Chlamydomonas strains capable of high and stable transgene expression. The established cells exhibited stable reporter gene expression at a high level throughout long-term culture (∼60 days), even in the absence of drug pressure. The transgene insertion sites in the cell genome that may be suitable for exogenous gene expression were identified. Because the transgene contains a loxP site, the cells can be used as founders for retargeting other transgenes using the Cre-loxP system to generate transgenic Chlamydomonas producing useful substances. As a model biopharmaceutical gene, an interferon expression cassette was integrated into the genomic locus of the cells using Cre recombinase. The transgenic cells stably produced interferon protein in medium for 12 passages under non-selective conditions. These results indicate that the Chlamydomonas cells established in this study can serve as valuable and powerful tools not only for basic research on microalgae but also for the rapid establishment of cell lines expressing exogenous genes.

预计衣藻将用于生物燃料和生物制药生产。然而，衣藻的基因工程技术并不像其他生物那样先进。在这里，我们建立了能够进行高且稳定的转基因表达的转基因衣藻菌株。即使在没有药物压力的情况下，已建立的细胞在整个长期培养（~60 天）中都表现出稳定的高水平报告基因表达。鉴定了细胞基因组中可能适合外源基因表达的转基因插入位点。因为转基因包含一个loxP位点，所以这些细胞可以用作创建者，以使用 CreloxP 重新定位其他转基因。产生产生有用物质的转基因衣藻的系统。作为模型生物制药基因，干扰素表达盒使用 Cre 重组酶整合到细胞的基因组基因座中。转基因细胞在非选择性条件下在培养基中稳定产生干扰素蛋白12代。这些结果表明，本研究建立的衣藻细胞不仅可以作为微藻基础研究的有价值和强大的工具，而且可以作为快速建立表达外源基因的细胞系。