Targeted Transcriptional Activation Using a CRISPR-Associated Transposon System
Targeted Transcriptional Activation Using a CRISPR-Associated Transposon System
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DOI：
10.1021/acssynbio.3c00563
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发表时间：
2023-12-12
影响因子：
4.7
通讯作者：
Chappell，James
中科院分区：
文献类型：
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作者：
Elizondo，Andrea M. Garza;Chappell，James
基因表达的合成扰动是我们可靠地揭示微生物基因型-表型关系的能力的核心。在这里，我们提出了一种新的转录激活策略，该策略使用霍乱弧菌CRISPR相关转座子（CAST）系统选择性地将启动子元件插入感兴趣的基因上游。通过这一策略，我们显示出强大的激活重组和内源基因在theEscherichia colichromosome。然后，我们展示了通过交换插入的启动子元件来精确调节表达水平。最后，我们证明了CAST激活可以用于在大肠杆菌中合成诱导氨苄青霉素抗性表型。杆菌
Synthetic perturbation of gene expression is central to our ability to reliably uncover genotype–phenotype relationships in microbes. Here, we present a novel transcription activation strategy that uses theVibrio choleraeCRISPR-Associated Transposon (CAST) system to selectively insert promoter elements upstream of genes of interest. Through this strategy, we show robust activation of both recombinant and endogenous genes across theEscherichia colichromosome. We then demonstrate the precise tuning of expression levels by exchanging the promoter elements being inserted. Finally, we demonstrate that CAST activation can be used to synthetically induce ampicillin-resistant phenotypes inE. coli.