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Abstract：
Synthetic promoters are considered ideal candidates in driving robust gene expression. Most of the available synthetic promoters are minimal promoters, for which the upstream sequence of the 5\' end of the core region is usually excluded. Although the upstream sequence has been shown to mediate transcription of natural promoters, its impact on synthetic promoters has not been widely studied. Here, a library of chromosomal DNA fragments is randomly fused with the 5\' end of the J23119 synthetic promoter, and the transcriptional performance of the promoter is evaluated through β-galactosidase assay, fluorescence intensity and chemical biosynthesis. Results show that changes in the upstream sequence can induce significant variation in the promoter strength of up to 5.8-fold. The effect is independent of the length of the insertions and the number of potential transcription factor binding sites. Several DNA fragments that are able to enhance the transcription of both the natural and the synthetic promoters are identified. This study indicates that the synthetic minimal promoters are susceptible to the surrounding sequence context. Therefore, the upstream sequence should be treated as an indispensable component in the design and application of synthetic promoters, or as an independent genetic part for the fine-tuning of gene expression.
摘要：
合成启动子被认为是驱动稳健基因表达的理想候选者。大多数可用的合成启动子是最小启动子，通常排除核心区域5'端的上游序列。尽管上游序列已被证明可介导天然启动子的转录，其对合成启动子的影响尚未被广泛研究。这里,染色体DNA片段的文库与J23119合成启动子的5'端随机融合，并通过β-半乳糖苷酶测定评估启动子的转录性能，荧光强度和化学生物合成。结果表明，上游序列的变化可以诱导启动子强度的显着变化高达5.8倍。该效应与插入的长度和潜在转录因子结合位点的数量无关。鉴定了能够增强天然和合成启动子两者的转录的若干DNA片段。该研究表明合成的最小启动子对周围的序列环境敏感。因此,上游序列应被视为合成启动子设计和应用中不可或缺的组成部分，或作为基因表达微调的独立遗传部分。
