Herein, a Broccoli/mCherry and an EGFP/mCherry dual-color fluorescent reporting systems have been established to quantify the promoter activity at transcription and translation level in eukaryotic cells. Based on those systems, four commonly used promoters (CMV and SV40 of Pol II and U6, H1 of Pol III) were accurately evaluated at both the transcriptional and translational levels by combining accurate protein and RNA quantification. Furthermore, we verified that Pol III promoters can induce proteins expression, and Pol II promoter can be applied to express RNA molecules with defined length by combining a self-cleaving ribozyme and an artificial poly(A) tail. The dual-color fluorescence reporting systems described here could play a significant role in evaluating other gene expression regulators for gene therapy.

中文翻译：

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使用双色荧光报告系统结合蛋白质和 RNA 定量评估启动子活性。

在此，建立了一个 Broccoli/mCherry 和一个 EGFP/mCherry 双色荧光报告系统，以量化真核细胞中转录和翻译水平的启动子活性。基于这些系统，通过结合准确的蛋白质和 RNA 定量，在转录和翻译水平上准确评估了四种常用启动子（Pol II 和 U6 的 CMV 和 SV40，Pol III 的 H1）。此外，我们验证了 Pol III 启动子可以诱导蛋白质表达，并且通过结合自切割核酶和人工 poly(A) 尾巴，可以将 Pol II 启动子用于表达具有确定长度的 RNA 分子。此处描述的双色荧光报告系统可以在评估其他基因表达调节剂用于基因治疗方面发挥重要作用。