Department of Plant Sciences, University of Tennessee, Knoxville, TN, USA.
Department of Grassland Science, Sichuan Agricultural University, Chengdu, Sichuan, China.
Plant Cell Rep. 2018 Apr;37(4):587-597. doi: 10.1007/s00299-018-2253-1. Epub 2018 Jan 17.
A switchgrass vascular tissue-specific promoter (PvPfn2) and its 5'-end serial deletions drive high levels of vascular bundle transgene expression in transgenic rice. Constitutive promoters are widely used for crop genetic engineering, which can result in multiple off-target effects, including suboptimal growth and epigenetic gene silencing. These problems can be potentially avoided using tissue-specific promoters for targeted transgene expression. One particularly urgent need for targeted cell wall modification in bioenergy crops, such as switchgrass (Panicum virgatum L.), is the development of vasculature-active promoters to express cell wall-affective genes only in the specific tissues, i.e., xylem and phloem. From a switchgrass expression atlas we identified promoter sequence upstream of a vasculature-specific switchgrass profilin gene (PvPfn2), especially in roots, nodes and inflorescences. When the putative full-length (1715 bp) and 5'-end serial deletions of the PvPfn2 promoter (shortest was 413 bp) were used to drive the GUS reporter expression in stably transformed rice (Oryza sativa L.), strong vasculature-specificity was observed in various tissues including leaves, leaf sheaths, stems, and flowers. The promoters were active in both phloem and xylem. It is interesting to note that the promoter was active in many more tissues in the heterologous rice system than in switchgrass. Surprisingly, all four 5'-end promoter deletions, including the shortest fragment, had the same expression patterns as the full-length promoter and with no attenuation in GUS expression in rice. These results indicated that the PvPfn2 promoter variants are new tools to direct transgene expression specifically to vascular tissues in monocots. Of special interest is the very compact version of the promoter, which could be of use for vasculature-specific genetic engineering in monocots.
一个柳枝稷维管束组织特异性启动子（PvPfn2）及其 5'端的串联缺失驱动转基因水稻中高水平的维管束转基因表达。组成型启动子广泛用于作物遗传工程，这可能导致多种非靶向效应，包括生长不佳和表观遗传基因沉默。通过组织特异性启动子进行靶向转基因表达，可以潜在地避免这些问题。在能源作物（如柳枝稷）中，靶向细胞壁修饰的一个特别紧迫的需求是开发血管活性启动子，以使细胞壁相关基因仅在特定组织（即木质部和韧皮部）中表达。从柳枝稷表达图谱中，我们鉴定出一个血管特异性柳枝稷丝状蛋白基因（PvPfn2）上游的启动子序列，特别是在根、节和花序中。当使用推测的全长（1715bp）和 5'端串联缺失的 PvPfn2 启动子（最短为 413bp）驱动 GUS 报告基因在稳定转化的水稻（Oryza sativa L.）中表达时，在包括叶片、叶鞘、茎和花在内的各种组织中观察到强烈的血管特异性。启动子在韧皮部和木质部都有活性。有趣的是，与柳枝稷相比，该启动子在异源水稻系统中在更多的组织中具有活性。令人惊讶的是，包括最短片段在内的所有四个 5'端启动子缺失都与全长启动子具有相同的表达模式，并且在水稻中 GUS 表达没有减弱。这些结果表明，PvPfn2 启动子变体是将转基因特异性导向单子叶植物维管束组织的新工具。特别有趣的是，该启动子的非常紧凑版本，可用于单子叶植物的血管特异性遗传工程。
