The alc-GR system.: A modified alc gene switch designed for use in plant tissue culture
The alc-GR system.: A modified alc gene switch designed for use in plant tissue culture
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DOI：
10.1104/pp.105.059659
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发表时间：
2005-07-01
期刊：
影响因子：
7.4
通讯作者：
Tomsett, AB
中科院分区：
文献类型：
--
作者：
Roberts, GR;Garoosi, GA;Tomsett, AB
ALCR/alcA（alc）双组分乙醇诱导型基因表达系统提供了对转基因植物中转基因表达的严格控制。ALCR是一种乙醇激活的转录因子，可以驱动ALCR应答启动子（alcA）的表达。然而，alc系统已被证明在植物愈伤组织或细胞悬浮培养物中使用时具有组成型表达，这可能是由于响应于降低的氧可用性而产生的内源诱导物。为了拓宽alc系统在植物细胞培养条件下的应用，将大鼠糖皮质激素受体（GR）的受体结构域与ALCR的C末端进行同源融合以产生ALCR-GR，其形成糖皮质激素诱导系统（alc-GR）的基础。在烟草（Nicotiana tabacum）Bright Yellow-2悬浮细胞中使用组成型表达的ALCR-GR测试alc-GR开关系统，所述ALCR-GR具有四种替代性alcA启动子驱动的报告基因：β-葡糖醛酸酶、内质网靶向绿色荧光蛋白、血凝素和绿色荧光蛋白标记的拟南芥（Arabidopsis thaliana）Arath; CDKA; 1细胞周期蛋白依赖性激酶。基因表达被证明是严格依赖于合成的糖皮质激素地塞米松，在细胞悬浮液中，不再需要乙醇诱导。因此，alc-GR系统允许严格控制细胞培养物中的alcA驱动的基因，并补充了整个植物中使用的常规乙醇开关。
The ALCR/alcA (alc) two-component, ethanol-inducible gene expression system provides stringent control of transgene expression in genetically modified plants. ALCR is an ethanol-activated transcription factor that can drive expression from the ALCR-responsive promoter ( alcA). However, the alc system has been shown to have constitutive expression when used in plant callus or cell suspension cultures, possibly resulting from endogenous inducer produced in response to lowered oxygen availability. To widen the use of the alc system in plant cell culture conditions, the receptor domain of the rat glucocorticoid receptor (GR) was translationally fused to the C terminus of ALCR to produce ALCR-GR, which forms the basis of a glucocorticoid-inducible system (alc-GR). The alc-GR switch system was tested in tobacco (Nicotiana tabacum) Bright Yellow-2 suspension cells using a constitutively expressed ALCR-GR with four alternative alcA promoter-driven reporter genes: beta-glucuronidase, endoplasmic reticulum-targeted green fluorescent protein, haemagglutinin, and green fluorescent protein-tagged Arabidopsis (Arabidopsis thaliana) Arath; CDKA; 1 cyclin-dependent kinase. Gene expression was shown to be stringently dependent on the synthetic glucocorticoid dexamethasone and, in cell suspensions, no longer required ethanol for induction. Thus, the alc-GR system allows tight control of alcA-driven genes in cell culture and complements the conventional ethanol switch used in whole plants.