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首页> 外文期刊>BMC Biotechnology >Development of an optimized tetracycline-inducible expression system to increase the accumulation of interleukin-10 in tobacco BY-2 suspension cells
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Development of an optimized tetracycline-inducible expression system to increase the accumulation of interleukin-10 in tobacco BY-2 suspension cells

机译：优化的四环素诱导表达系统的开发，以增加白细胞介素10在烟草BY-2悬浮细胞中的积累

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Background Plant cell suspension cultures can be used for the production of valuable pharmaceutical and industrial proteins. When the recombinant protein is secreted into the culture medium, restricting expression to a defined growth phase can improve both the quality and quantity of the recovered product by minimizing proteolytic activity. Temporal restriction is also useful for recombinant proteins whose constitutive expression affects cell growth and viability, such as viral interleukin-10 (vIL-10). Results We have developed a novel, tetracycline-inducible system suitable for tobacco BY-2 suspension cells which increases the yields of vIL-10. The new system is based on a binary vector that is easier to handle than conventional vectors, contains an enhanced inducible promoter and 5′-UTR to improve yields, and incorporates a constitutively-expressed visible marker gene to allow the rapid and straightforward selection of the most promising transformed clones. Stable transformation of BY-2 cells with this vector, without extensive optimization of the induction conditions, led to a 3.5 fold increase in vIL-10 levels compared to constitutive expression in the same host. Conclusions We have developed an effective and straightforward molecular farming platform technology that improves both the quality and the quantity of recombinant proteins produced in plant cells, particularly those whose constitutive expression has a negative impact on plant growth and development. Although we tested the platform using vIL-10 produced in BY-2 cells, it can be applied to other host/product combinations and is also useful for basic research requiring strictly controlled transgene expression.
机译：背景技术植物细胞悬浮培养物可用于生产有价值的药物和工业蛋白质。当重组蛋白分泌到培养基中时，将表达限制在确定的生长期可以通过使蛋白水解活性最小化来提高回收产物的质量和数量。时间限制也可用于重组蛋白，其组成性表达会影响细胞的生长和生存能力，例如病毒白介素10（vIL-10）。结果我们开发了一种适用于烟草BY-2悬浮细胞的新型四环素诱导系统，该系统可提高vIL-10的产量。新系统基于一种二元载体，该二元载体比常规载体更易于操作，包含增强的诱导型启动子和5'-UTR以提高产量，并整合了组成型表达的可见标记基因，可以快速，直接地选择最有希望的转化克隆。与该宿主中的组成型表达相比，在没有广泛优化诱导条件的情况下，用该载体稳定转化BY-2细胞可导致vIL-10水平提高3.5倍。结论我们已经开发了一种有效而直接的分子耕种平台技术，可以提高植物细胞中重组蛋白的质量和数量，特别是那些组成型表达对植物生长和发育产生负面影响的重组蛋白。尽管我们使用BY-2细胞中产生的vIL-10测试了该平台，但它可以应用于其他宿主/产品组合，也可用于需要严格控制转基因表达的基础研究。

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