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用于高通量功能筛选应用的海肾荧光素酶变体。

Gaussia luciferase variant for high-throughput functional screening applications.

机构信息

Molecular Neurogenetics Unit, Department of Neurology, Harvard Medical School, Boston Massachusetts 02115, USA.

出版信息

Anal Chem. 2009 Aug 15;81(16):7102-6. doi: 10.1021/ac901234r.

DOI:10.1021/ac901234r
PMID:19601604
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC2846205/
Abstract

Gaussia luciferase (Gluc) is a sensitive reporter for studying different biological processes such as gene expression, promoter activity, protein-protein interactions, signal transduction, as well as tumor cell growth and response to therapy. Since Gluc is naturally secreted, the kinetics of these processes can be monitored in real-time by measuring an aliquot of conditioned medium in culture or a few microliters of blood in vivo at different time points. Gluc catalyzes light emission with a short half-life which is unfavorable for certain applications. We isolated a Gluc mutant that catalyzes enhanced light stability in the presence of a detergent, in combination with high sensitivity, making it an attractive luciferase for high-throughput functional screening applications.

摘要

海肾荧光素酶(Gluc)是一种灵敏的报告基因,可用于研究多种生物学过程,如基因表达、启动子活性、蛋白质-蛋白质相互作用、信号转导以及肿瘤细胞的生长和对治疗的反应等。由于 Gluc 是天然分泌的,因此可以通过在不同时间点测量培养物中的条件培养基的等分试样或体内的几微升血液,实时监测这些过程的动力学。Gluc 催化的发光具有半衰期短的缺点,不利于某些应用。我们分离出一种 Gluc 突变体,该突变体在去污剂存在下催化增强的光稳定性,同时保持高灵敏度,使其成为高通量功能筛选应用的有吸引力的荧光素酶。

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本文引用的文献

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Nat Protoc. 2009;4(4):582-91. doi: 10.1038/nprot.2009.28.
2
Sensitive in vivo imaging of T cells using a membrane-bound Gaussia princeps luciferase.使用膜结合型海肾荧光素酶对T细胞进行灵敏的体内成像。
Nat Med. 2009 Mar;15(3):338-44. doi: 10.1038/nm.1930. Epub 2009 Feb 15.
3
A pathway sensor for genome-wide screens of intracellular proteolytic cleavage.一种用于细胞内蛋白水解切割的全基因组筛选的途径传感器。
Genome Biol. 2008 Apr 3;9(4):R64. doi: 10.1186/gb-2008-9-4-r64.
4
A secreted luciferase for ex vivo monitoring of in vivo processes.一种用于体内过程离体监测的分泌型荧光素酶。
Nat Methods. 2008 Feb;5(2):171-3. doi: 10.1038/nmeth.1177. Epub 2008 Jan 20.
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Development of a dual-luciferase reporter system for in vivo visualization of MicroRNA biogenesis and posttranscriptional regulation.用于体内可视化微小RNA生物合成和转录后调控的双荧光素酶报告系统的开发。
J Nucl Med. 2008 Feb;49(2):285-94. doi: 10.2967/jnumed.107.042507. Epub 2008 Jan 16.
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Identification of two catalytic domains in a luciferase secreted by the copepod Gaussia princeps.在桡足类动物高斯伪镖水蚤分泌的荧光素酶中鉴定出两个催化结构域。
Biochem Biophys Res Commun. 2008 Jan 4;365(1):96-101. doi: 10.1016/j.bbrc.2007.10.152. Epub 2007 Nov 5.
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FEBS Lett. 2007 Oct 2;581(24):4551-6. doi: 10.1016/j.febslet.2007.08.036. Epub 2007 Aug 27.
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PLoS One. 2007 Jun 27;2(6):e571. doi: 10.1371/journal.pone.0000571.
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