Construction of DNA Tools for Hyperexpression in Marchantia Chloroplasts.
Construction of DNA Tools for Hyperexpression in Marchantia Chloroplasts.
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DOI：
10.1021/acssynbio.0c00637
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发表时间：
2021-07-16
影响因子：
4.7
通讯作者：
Haseloff J
中科院分区：
文献类型：
--
作者：
Frangedakis E;Guzman-Chavez F;Rebmann M;Markel K;Yu Y;Perraki A;Tse SW;Liu Y;Rever J;Sauret-Gueto S;Goffinet B;Schneider H;Haseloff J
叶绿体是合成生物学应用的有吸引力的平台，因为如果mRNA的产生和稳定性得到适当调节，它们能够驱动非常高水平的转基因表达。然而，质体转化是一个缓慢的过程，目前仅限于少数植物物种。地钱是一种简单的模式植物，允许快速转化研究，然而，其蛋白质过表达的潜力尚未得到充分利用。这部分是由于叶绿体转录后调控的特点是在这种植物的事实。我们已经绘制了地钱叶绿体中的转录模式。此外，我们已经获得并比较了51个物种的序列，并确定了pentatricopeptide重复蛋白结合被认为在mRNA稳定中发挥重要作用的推定位点。候选人的结合位点进行了测试，他们的能力，赋予高水平的报告基因表达地钱叶绿体，和蛋白质的生产水平和对生长的影响进行了测量，在同源转化植物。我们已经产生了新的DNA工具，在这个简单的植物系统，是叶绿体工程的试验台蛋白质过表达。
Chloroplasts are attractive platforms for synthetic biology applications since they are capable of driving very high levels of transgene expression, if mRNA production and stability are properly regulated. However, plastid transformation is a slow process and currently limited to a few plant species. The liverwort Marchantia polymorpha is a simple model plant that allows rapid transformation studies; however, its potential for protein hyperexpression has not been fully exploited. This is partially due to the fact that chloroplast post-transcriptional regulation is poorly characterized in this plant. We have mapped patterns of transcription in Marchantia chloroplasts. Furthermore, we have obtained and compared sequences from 51 bryophyte species and identified putative sites for pentatricopeptide repeat protein binding that are thought to play important roles in mRNA stabilization. Candidate binding sites were tested for their ability to confer high levels of reporter gene expression in Marchantia chloroplasts, and levels of protein production and effects on growth were measured in homoplastic transformed plants. We have produced novel DNA tools for protein hyperexpression in this facile plant system that is a test-bed for chloroplast engineering.
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