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Heterologous Gene Expression in Chlamydomonas reinhardtii Chloroplast by Heterologous Promoters and Terminators, Intercistronic Expression Elements and Minichromosome

文献类型： 外文期刊

作者： Guo, Yunling 1 ; Xiong, Hui 1 ; Fan, Qiuling 2 ; Duanmu, Deqiang 1 ;

作者机构： 1.Huazhong Agr Univ, State Key Lab Agr Microbiol, Hubei Hongshan Lab, Wuhan, Peoples R China

2.Huazhong Agr Univ, Coll Life Sci & Technol, Wuhan, Peoples R China

3.Chinese Acad Agr Sci, Agr Genom Inst Shenzhen, Shenzhen Branch, Guangdong Lab Lingnan Modern Agr,Genome Anal Lab,M, Shenzhen, Peoples R China

关键词： Chlamydomonas reinhardtii; chloroplast biotechnology; intercistronic expression elements; minichromosome; photosynthetic chassis

期刊名称：MICROBIAL BIOTECHNOLOGY （ 影响因子：5.2； 五年影响因子：5.9 ）

ISSN： 1751-7915

年卷期： 2024 年 17 卷 12 期

页码：

收录情况： SCI

摘要： Chlamydomonas reinhardtii, a model green alga for expressing foreign proteins, faces challenges in multigene expression and enhancing protein expression level in the chloroplast. To address these challenges, we compared heterologous promoters, terminators and intercistronic expression elements (IEEs). We transformed Chlamydomonas chloroplast with a biolistic approach to introduce vectors containing the NanoLuc expression unit regulated by Chlamydomonas or tobacco promoters and terminators. We observed that tobacco promoters PrbcL and PpsbA could not effectively regulate protein expression, whereas tobacco terminators TrbcL and Trps16 did not affect the expression of Nluc protein. Further exploration of IEEs specific to Chlamydomonas revealed that Cr-IEE2 had a minor effect on both upstream and downstream protein expression, whereas Cr-IEE5 significantly influenced downstream protein expression. In contrast, tobacco IEE was found to be unsuitable for driving protein expression in Chlamydomonas. Additionally, VOR element and Rep protein derived from beet curly top geminivirus were able to form a minichromosome in Chlamydomonas chloroplast, and this system could enhance protein expression level compared to the traditional method of site-specific integration in the plastome. This study highlights the potential of IEEs and minichromosome in facilitating heterologous protein expression in Chlamydomonas chloroplast.

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