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| 绿色荧光蛋白为传能对的供体光漂白法测定FRET效率的探讨 | |
| 引用本文： | 周晓娟,郭慧芳,刘德龙,孙大业.绿色荧光蛋白为传能对的供体光漂白法测定FRET效率的探讨[J].分析测试学报,2008,27(2):127-130. |
| 作者姓名： | 周晓娟 郭慧芳 刘德龙 孙大业 |
| 作者单位： | 1. 河北师范大学,化学学院,河北,石家庄,050016 2. 河北师范大学,生命科学学院,河北,石家庄,050016 3. 河北师范大学,化学学院,河北,石家庄,050016;河北师范大学,生命科学学院,河北,石家庄,050016 |
| 摘 要： | 探讨以两种新荧光蛋白MiCy,mKo为传能对并应用供体光漂白法测量荧光共振能量转移(FRET)效率.首先通过基因工程方法表达纯化了这两种蛋白,并测量了荧光光谱及光漂白性质,表明MiCy极易光漂白而mKo抗光漂白.进一步以Ni-NTA-agarose为FRET模型,在Confocal上对MiCy进行光漂白时间常数的测量,并计算了FRET效率.结果表明MiCy-mKo传能对适合用供体光漂白法测量FRET效率,此传能对将在蛋白质相互作用研究中有广泛应用. |
| 关 键 词： | 绿色荧光蛋白 荧光共振能量转移 FRET效率 光漂白 绿色 荧光蛋白 传能 光漂白 测定 FRET 效率 Pair Energy Transfer Green Fluorescent Proteins Method Photobleaching Donor Efficiency 研究 相互作用 蛋白质 结果 计算 时间常数 |
| 文章编号： | 1004-4957（2008）02-0127-04 |
| 收稿时间： | 2007-01-18 |
| 修稿时间： | 2007-04-08 |
Determination of FRET Efficiency Using Donor Photobleaching Method with Green Fluorescent Proteins as Energy Transfer Pair |
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| ZHOU Xiao-juan,GUO Hui-fang,LIU De-long,SUN Da-ye.Determination of FRET Efficiency Using Donor Photobleaching Method with Green Fluorescent Proteins as Energy Transfer Pair[J].Journal of Instrumental Analysis,2008,27(2):127-130. | |
| Authors: | ZHOU Xiao-juan GUO Hui-fang LIU De-long SUN Da-ye |
| Abstract: | In this work, two new green fluorescence proteins, named MiCy and mKo, were used as the energy transfer pair and their FRET efficiency was measured by the donor photobleaching method. The excitation and emission spectra and the photobleaching curves of the purified MiCy and mKo proteins were determined. The results suggested that MiCy is the photobleaching donor and that mKo is a photostable acceptor. By using Ni -NTA -agarose as the FRET model, true transfer efficiency between MiCy and mKo was successfully detected on Confocal microscope. In conclusion, the MiCy - mKo is a proper FRET pair, and the donor photobleaching method can provide a very specific measurement of FRET efficiency. |
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