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用于……的嵌合开放阅读框融合表型报告基因

A Chimeric ORF Fusion Phenotypic Reporter for .

作者信息

Phillips-Rose Louis S, Yu Chendi K, West Nicholas P, Fraser James A

机构信息

Australian Infectious Diseases Research Centre, School of Chemistry & Molecular Biosciences, The University of Queensland, Brisbane, QLD 4072, Australia.

出版信息

J Fungi (Basel). 2024 Aug 12;10(8):567. doi: 10.3390/jof10080567.

DOI:10.3390/jof10080567
PMID:39194893
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC11355724/
Abstract

The plethora of genome sequences produced in the postgenomic age has not resolved many of our most pressing biological questions. Correlating gene expression with an interrogatable and easily observable characteristic such as the surrogate phenotype conferred by a reporter gene is a valuable approach to gaining insight into gene function. Many reporters including , and the fluorescent proteins mRuby3 and mNeonGreen have been used across all manners of organisms. Described here is an investigation into the creation of a robust, synthetic, fusion reporter system for that combines some of the most useful fluorophores available in this system with the versatility of the counter-selectable nature of . The reporters generated include multiple composition and orientation variants, all of which were investigated for differences in expression. Evaluation of known promoters from the and genes was undertaken, elucidating novel expression tendencies of these biologically relevant regulators of transcription. Smaller than but providing multiple useful surrogate phenotypes for interrogation, the fusion ORF serves as a superior whole-cell assay compared to traditional systems. Ultimately, the work described here bolsters the array of relevant genetic tools that may be employed in furthering manipulation and understanding of the WHO fungal priority group pathogen .

摘要

后基因组时代产生的大量基因组序列并未解决我们许多最紧迫的生物学问题。将基因表达与可询问且易于观察的特征(如报告基因赋予的替代表型)相关联,是深入了解基因功能的一种有价值的方法。包括……以及荧光蛋白mRuby3和mNeonGreen在内的许多报告基因已在各种生物体中使用。本文描述了一项针对……创建强大的合成融合报告系统的研究,该系统将该系统中一些最有用的荧光团与……的反选择性质的多功能性结合起来。产生的报告基因包括多种组成和方向变体,所有这些都对表达差异进行了研究。对来自……和……基因的已知启动子进行了评估,阐明了这些生物学相关的转录调节因子的新表达趋势。融合开放阅读框比……小,但提供了多种有用的替代表型以供询问,与传统系统相比,它是一种更优越的全细胞分析方法。最终,本文所述的工作加强了一系列相关的遗传工具,这些工具可用于进一步操纵和了解世界卫生组织真菌重点病原体……

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e696/11355724/168254c54bb9/jof-10-00567-g006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e696/11355724/c96f1ec5a33a/jof-10-00567-g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e696/11355724/ead780998c93/jof-10-00567-g002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e696/11355724/895806d976de/jof-10-00567-g003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e696/11355724/e473da86a05e/jof-10-00567-g004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e696/11355724/50ecd3b314d8/jof-10-00567-g005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e696/11355724/168254c54bb9/jof-10-00567-g006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e696/11355724/c96f1ec5a33a/jof-10-00567-g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e696/11355724/ead780998c93/jof-10-00567-g002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e696/11355724/895806d976de/jof-10-00567-g003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e696/11355724/e473da86a05e/jof-10-00567-g004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e696/11355724/50ecd3b314d8/jof-10-00567-g005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e696/11355724/168254c54bb9/jof-10-00567-g006.jpg

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本文引用的文献

1
Cryptococcosis in Africa: What the data tell us.非洲的隐球菌病:数据告诉我们什么。
Med Mycol. 2023 Jun 5;61(6). doi: 10.1093/mmy/myad049.
2
UniProt: the Universal Protein Knowledgebase in 2023.UniProt:2023 年的通用蛋白质知识库。
Nucleic Acids Res. 2023 Jan 6;51(D1):D523-D531. doi: 10.1093/nar/gkac1052.
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Characterization of Bicistronic Transcription in Budding Yeast.芽殖酵母中双顺反子转录的表征
mSystems. 2021 Feb 23;6(1):e01002-20. doi: 10.1128/mSystems.01002-20.
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Broadening the spectrum of fluorescent protein tools for use in the encapsulated human fungal pathogen Cryptococcus neoformans.拓宽荧光蛋白工具在包被人真菌病原体新型隐球菌中的应用范围。
Fungal Genet Biol. 2020 May;138:103365. doi: 10.1016/j.fgb.2020.103365. Epub 2020 Mar 4.
5
Comparing the performance of mScarlet-I, mRuby3, and mCherry as FRET acceptors for mNeonGreen.比较 mScarlet-I、mRuby3 和 mCherry 作为 mNeonGreen 的 FRET 受体的性能。
PLoS One. 2020 Feb 5;15(2):e0219886. doi: 10.1371/journal.pone.0219886. eCollection 2020.
6
Quantitative global studies reveal differential translational control by start codon context across the fungal kingdom.定量全局研究揭示了真菌界中起始密码子上下文对翻译控制的差异。
Nucleic Acids Res. 2020 Mar 18;48(5):2312-2331. doi: 10.1093/nar/gkaa060.
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Construction of a RFP-lacZα bicistronic reporter system and its application in lead biosensing.构建 RFP-lacZα 双顺反子报告系统及其在铅生物传感中的应用。
PLoS One. 2020 Jan 30;15(1):e0228456. doi: 10.1371/journal.pone.0228456. eCollection 2020.
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amdS as a dominant recyclable marker in Cryptococcus neoformans.amdS 作为新型隐球菌中的优势可回收标记。
Fungal Genet Biol. 2019 Oct;131:103241. doi: 10.1016/j.fgb.2019.103241. Epub 2019 Jun 17.
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FPbase: a community-editable fluorescent protein database.FPbase:一个可由社区编辑的荧光蛋白数据库。
Nat Methods. 2019 Apr;16(4):277-278. doi: 10.1038/s41592-019-0352-8.
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Introns in Cryptococcus.隐球菌中的内含子
Mem Inst Oswaldo Cruz. 2018;113(7):e170519. doi: 10.1590/0074-02760170519. Epub 2018 Feb 19.