Max-Planck-Institut für Molekulare Pflanzenphysiologie, Am Mühlenberg 1, D-14476 Potsdam-Golm, Germany.
Plant J. 2012 Oct;72(1):115-28. doi: 10.1111/j.1365-313X.2012.05065.x. Epub 2012 Jul 17.
Although our knowledge about the mechanisms of gene expression in chloroplasts has increased substantially over the past decades, next to nothing is known about the signals and factors that govern expression of the plastid genome in non-green tissues. Here we report the development of a quantitative method suitable for determining the activity of cis-acting elements for gene expression in non-green plastids. The in vivo assay is based on stable transformation of the plastid genome and the discovery that root length upon seedling growth in the presence of the plastid translational inhibitor kanamycin is directly proportional to the expression strength of the resistance gene nptII in transgenic tobacco plastids. By testing various combinations of promoters and translation initiation signals, we have used this experimental system to identify cis-elements that are highly active in non-green plastids. Surprisingly, heterologous expression elements from maize plastids were significantly more efficient in conferring high expression levels in root plastids than homologous expression elements from tobacco. Our work has established a quantitative method for characterization of gene expression in non-green plastid types, and has led to identification of cis-elements for efficient plastid transgene expression in non-green tissues, which are valuable tools for future transplastomic studies in basic and applied research.
尽管在过去几十年中，我们对叶绿体中基因表达机制的了解有了实质性的增加，但对于控制非绿色组织中质体基因组表达的信号和因素，我们几乎一无所知。在这里，我们报告了一种定量方法的发展，该方法适用于确定非绿色质体中基因表达的顺式作用元件的活性。该体内测定基于质体基因组的稳定转化，以及在含有质体翻译抑制剂卡那霉素的幼苗生长过程中发现的根长与转基因烟草质体中抗性基因 nptII 的表达强度直接成正比。通过测试各种启动子和翻译起始信号的组合，我们已经使用该实验系统鉴定了在非绿色质体中高度活跃的顺式元件。令人惊讶的是，来自玉米质体的异源表达元件在赋予根质体中高表达水平方面比来自烟草的同源表达元件更有效。我们的工作建立了一种用于非绿色质体类型基因表达特征描述的定量方法，并确定了非绿色组织中高效质体转基因表达的顺式元件，这是未来基础和应用研究中转质体研究的有价值的工具。
