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DNA 和核小体相衬电镜成像的相衬技术进展。

Advances in phase plate cryo-EM imaging of DNA and nucleosomes.

机构信息

a School of Biological Sciences , Nanyang Technological University , Singapore.

b NTU Institute of Structural Biology , Nanyang Technological University , Singapore.

出版信息

Nucleus. 2017 May 4;8(3):275-278. doi: 10.1080/19491034.2017.1287643. Epub 2017 Feb 7.

DOI:10.1080/19491034.2017.1287643
PMID:28340334
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC5499919/
Abstract

Contrast in electron cryo-microscopy (cryo-EM) is limited by the weak phase and radiation sensitive nature of biologic samples embedded in vitrified ice. We have recently shown that a new contrast enhancement technique utilizing the Volta phase plate can be combined with single particle analysis to determine the structure of a small chromatin complex, the nucleosome core particle, at near-atomic resolution. Here, we discuss advantages and limitations of the technique in terms of data collection, particle detection, and visualization of individual DNA molecules and higher-order chromatin structure.

摘要

电子冷冻透射显微镜(cryo-EM)的对比度受到限制,因为嵌入在玻璃态冰中的生物样本的相位很弱,对辐射也很敏感。我们最近表明,一种利用伏特相衬板的新技术可以与单颗粒分析结合使用,以确定一个小染色质复合物(核小体核心颗粒)的结构,达到近原子分辨率。在这里,我们将讨论该技术在数据收集、颗粒检测以及单个 DNA 分子和高级染色质结构可视化方面的优势和局限性。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/6e3c/5499919/6d1eccdf9446/kncl-08-03-1287643-g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/6e3c/5499919/6d1eccdf9446/kncl-08-03-1287643-g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/6e3c/5499919/6d1eccdf9446/kncl-08-03-1287643-g001.jpg

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本文引用的文献

1
Cryo-EM structure of haemoglobin at 3.2 Å determined with the Volta phase plate.采用 Volta 相板技术获得的 3.2 Å 分辨率血影蛋白冷冻电镜结构。
Nat Commun. 2017 Jun 30;8:16099. doi: 10.1038/ncomms16099.
2
Using the Volta phase plate with defocus for cryo-EM single particle analysis.在冷冻电镜单颗粒分析中使用带散焦的伏打相位板。
Elife. 2017 Jan 21;6:e23006. doi: 10.7554/eLife.23006.
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3.9 Å structure of the nucleosome core particle determined by phase-plate cryo-EM.通过相板冷冻电镜确定的核小体核心颗粒的3.9埃结构。
Sci Rep. 2021 Jan 11;11(1):380. doi: 10.1038/s41598-020-79654-8.
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Spectral comparisons of mammalian cells and intact organelles by solid-state NMR.固态 NMR 技术对哺乳动物细胞和完整细胞器的光谱比较。
J Struct Biol. 2019 Apr 1;206(1):49-54. doi: 10.1016/j.jsb.2018.05.007. Epub 2018 May 30.
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Cell. 2016 Jun 16;165(7):1698-1707. doi: 10.1016/j.cell.2016.05.040. Epub 2016 May 26.
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Cryo-EM single particle analysis with the Volta phase plate.冷冻电镜单颗粒分析与 Volta 相衬板。
Elife. 2016 Mar 7;5:e13046. doi: 10.7554/eLife.13046.
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Visualizing the molecular sociology at the HeLa cell nuclear periphery.可视化 HeLa 细胞核周缘的分子社会学。
Science. 2016 Feb 26;351(6276):969-72. doi: 10.1126/science.aad8857.
7
Volta phase plate cryo-EM of the small protein complex Prx3.小蛋白复合物Prx3的伏打相位板冷冻电镜技术
Nat Commun. 2016 Jan 28;7:10534. doi: 10.1038/ncomms10534.
8
Electron cryotomography of vitrified cells with a Volta phase plate.使用Volta相板对玻璃化细胞进行电子冷冻断层扫描。
J Struct Biol. 2015 May;190(2):143-54. doi: 10.1016/j.jsb.2015.03.004. Epub 2015 Mar 12.
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Proteasomes. A molecular census of 26S proteasomes in intact neurons.蛋白酶体。完整神经元中 26S 蛋白酶体的分子普查。
Science. 2015 Jan 23;347(6220):439-42. doi: 10.1126/science.1261197.
10
Volta potential phase plate for in-focus phase contrast transmission electron microscopy.用于明场相位对比透射电子显微镜的沃尔塔位相板。
Proc Natl Acad Sci U S A. 2014 Nov 4;111(44):15635-40. doi: 10.1073/pnas.1418377111. Epub 2014 Oct 20.