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首页> 外文期刊>Journal of Experimental Botany >Generating and characterizing single- and multigene mutants of the Rubisco small subunit family in Arabidopsis
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Generating and characterizing single- and multigene mutants of the Rubisco small subunit family in Arabidopsis

机译：在拟南芥中产生和表征Rubisco小亚基家族的单颗粒和多烯叶突变体

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The primary CO2-fixing enzyme Rubisco limits the productivity of plants. The small subunit of Rubisco (SSU) can influence overall Rubisco levels and catalytic efficiency, and is now receiving increasing attention as a potential engineering target to improve the performance of Rubisco. However, SSUs are encoded by a family of nuclear rbcS genes in plants, which makes them challenging to engineer and study. Here we have used CRISPR/Cas9 [clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR-associated protein 9] and T-DNA insertion lines to generate a suite of single and multiple gene knockout mutants for the four members of the rbcS family in Arabidopsis, including two novel mutants 2b3b and 1a2b3b. 1a2b3b contained very low levels of Rubisco (similar to 3% relative to the wild-type) and is the first example of a mutant with a homogenous Rubisco pool consisting of a single SSU isoform (1B). Growth under near-outdoor levels of light demonstrated Rubisco-limited growth phenotypes for several SSU mutants and the importance of the 1A and 3B isoforms. We also identified 1a1b as a likely lethal mutation, suggesting a key contributory role for the least expressed 1B isoform during early development. The successful use of CRISPR/Cas here suggests that this is a viable approach for exploring the functional roles of SSU isoforms in plants.
机译：初级CO2固定酶Rubisco限制了植物的生产率。 Rubisco（SSU）的小亚基可以影响整体鲁斯科水平和催化效率，现在正在接受提高潜在工程目标，以提高Rubisco的性能。然而，SSUS由植物中的一系列核RBCS基因编码，这使得它们对工程师和学习挑战。在这里，我们使用CRISPR / CAS9 [聚类定期间隙的回文重复（CRISPR）/ CRISPR相关蛋白质9]和T-DNA插入线，以产生拟南芥RBCS家族四个成员的单一和多基因敲除突变体的套件，包括两种新突变体2b3b和1a2b3b。图1A2B3B包含了非常低的Rubisco（相对于野生型类似的3％），并且是由单个SSU同种型（1B）组成的均质Rubisco池的突变体的第一实例。近室内光线下的增长显示了几种SSU突变体的鲁代菌类有限的生长表型以及1A和3B同种型的重要性。我们还将1A1B鉴定为可能的致命突变，这表明早期发育期间最不表达1B同种型的关键贡献作用。这里的CRISPR / CA的成功使用表明这是探索SSU同种型在植物中的功能作用的可行方法。

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