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公开(公告)号：US20140329234A1
公开(公告)日：2014-11-06
申请号：US14294256
申请日：2014-06-03
CPC分类号： G01N33/582 , C12P19/34 , C12P21/02 , C12Q1/6806 , G01N33/5308 , G01N33/6803
摘要： Provided are methods for labeling transfer RNA comprising replacing the uracil component of a dihydrouridine of said transfer RNA with a fluorophore. The disclosed methods may comprise fluorescent labeling of natural tRNAs (i.e., tRNAs that have been synthesized in a cell, for example, in a bacterium, a yeast cell, or a vertebrate cell) at dihydrouridine (D) positions, or fluorescent labeling of synthetic tRNAs. In another aspect, the present invention provides methods for assessing protein synthesis in a translation system comprise providing a tRNA having a fluorophore substitution for the uracil component of a dihydrouridine in a D loop of the tRNA; introducing the labeled tRNA into the translation system; irradiating the translation system with electromagnetic radiation, thereby generating a fluorescence signal from the fluorophore; detecting the fluorescence signal; and, correlating the fluorescence signal to one or more characteristics of the protein synthesis in the translation system. The disclosed methods are useful in single molecule as well as in ensemble settings.
摘要翻译： 提供用于标记转移RNA的方法，包括用荧光团替代所述转移RNA的二氢尿苷的尿嘧啶组分。 所公开的方法可以包括在二氢尿苷（D）位置处天然tRNA（即在细胞中例如细菌，酵母细胞或脊椎动物细胞中合成的tRNA）的荧光标记，或合成的荧光标记 tRNA。 另一方面，本发明提供了用于评估翻译系统中的蛋白质合成的方法，包括在tRNA的D环中提供具有二氢尿苷的尿嘧啶组分的荧光团取代的tRNA; 将标记的tRNA引入翻译系统; 用电磁辐射照射翻译系统，从而产生来自荧光团的荧光信号; 检测荧光信号; 并将荧光信号与翻译系统中蛋白质合成的一个或多个特征相关联。 所公开的方法在单分子以及整体设置中是有用的。
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公开(公告)号：US08785119B2
公开(公告)日：2014-07-22
申请号：US12664952
申请日：2008-06-20
CPC分类号： G01N33/582 , C12P19/34 , C12P21/02 , C12Q1/6806 , G01N33/5308 , G01N33/6803
摘要： Provided are methods for labeling transfer RNA comprising replacing the uracil component of a dihydrouridine of said transfer RNA with a fluorophore. The disclosed methods may comprise fluorescent labeling of natural tRNAs (i.e., tRNAs that have been synthesized in a cell, for example, in a bacterium, a yeast cell, or a vertebrate cell) at dihydrouridine (D) positions, or fluorescent labeling of synthetic tRNAs. In another aspect, the present invention provides methods for assessing protein synthesis in a translation system comprise providing a tRNA having a fluorophore substitution for the uracil component of a dihydrouridine in a D loop of the tRNA; introducing the labeled tRNA into the translation system; irradiating the translation system with electromagnetic radiation, thereby generating a fluorescence signal from the fluorophore; detecting the fluorescence signal; and, correlating the fluorescence signal to one or more characteristics of the protein synthesis in the translation system. The disclosed methods are useful in single molecule as well as in ensemble settings.
摘要翻译： 提供用于标记转移RNA的方法，包括用荧光团替代所述转移RNA的二氢尿苷的尿嘧啶组分。 所公开的方法可以包括在二氢尿苷（D）位置处天然tRNA（即在细胞中例如细菌，酵母细胞或脊椎动物细胞中合成的tRNA）的荧光标记，或合成的荧光标记 tRNA。 另一方面，本发明提供了用于评估翻译系统中的蛋白质合成的方法，包括在tRNA的D环中提供具有二氢尿苷的尿嘧啶组分的荧光团取代的tRNA; 将标记的tRNA引入翻译系统; 用电磁辐射照射翻译系统，从而产生来自荧光团的荧光信号; 检测荧光信号; 并将荧光信号与翻译系统中蛋白质合成的一个或多个特征相关联。 所公开的方法在单分子以及整体设置中是有用的。
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公开(公告)号：US20110262899A1
公开(公告)日：2011-10-27
申请号：US12664952
申请日：2008-06-20
CPC分类号： G01N33/582 , C12P19/34 , C12P21/02 , C12Q1/6806 , G01N33/5308 , G01N33/6803
摘要： Provided are methods for labeling transfer RNA comprising replacing the uracil component of a dihydrouridine of said transfer RNA with a fluorophore. The disclosed methods may comprise fluorescent labeling of natural tRNAs (i.e., tRNAs that have been synthesized in a cell, for example, in a bacterium, a yeast cell, or a vertebrate cell) at dihydrouridine (D) positions, or fluorescent labeling of synthetic tRNAs. In another aspect, the present invention provides methods for assessing protein synthesis in a translation system comprise providing a tRNA having a fluorophore substitution for the uracil component of a dihydrouridine in a D loop of the tRNA; introducing the labeled tRNA into the translation system; irradiating the translation system with electromagnetic radiation, thereby generating a fluorescence signal from the fluorophore; detecting the fluorescence signal; and, correlating the fluorescence signal to one or more characteristics of the protein synthesis in the translation system. The disclosed methods are useful in single molecule as well as in ensemble settings.
摘要翻译： 提供用于标记转移RNA的方法，包括用荧光团替代所述转移RNA的二氢尿苷的尿嘧啶组分。 所公开的方法可以包括在二氢尿苷（D）位置处天然tRNA（即在细胞中例如细菌，酵母细胞或脊椎动物细胞中合成的tRNA）的荧光标记，或合成的荧光标记 tRNA。 另一方面，本发明提供了用于评估翻译系统中的蛋白质合成的方法，包括在tRNA的D环中提供具有二氢尿苷的尿嘧啶组分的荧光团取代的tRNA; 将标记的tRNA引入翻译系统; 用电磁辐射照射翻译系统，从而产生来自荧光团的荧光信号; 检测荧光信号; 并将荧光信号与翻译系统中蛋白质合成的一个或多个特征相关联。 所公开的方法在单分子以及整体设置中是有用的。
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