The potential of 3D-FISH and super-resolution structured illumination microscopy for studies of 3D nuclear architecture
The potential of 3D-FISH and super-resolution structured illumination microscopy for studies of 3D nuclear architecture
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DOI：
10.1002/bies.201100176
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发表时间：
2012-05-01
期刊：
影响因子：
4
通讯作者：
Cremer, Marion
中科院分区：
文献类型：
--
作者：
Markaki, Yolanda;Smeets, Daniel;Cremer, Marion
三维结构照明显微镜(3D-SIM)为在超微结构水平上研究近100 nm范围内的核结构开辟了新的可能性。我们介绍了第一个结果，并评估了使用3D-SIM结合3D荧光原位杂交(3D-FISH)对已定义的核目标进行地形分析的潜力。我们的研究还涉及到这样一种担忧，即鱼类产生的人工制品可能会抵消分辨率的提高。我们讨论了DAPI染色的DNA在3D-FISH前后的细胞核、核孔和板层、染色体区域、染色质结构域和单个基因座中的形貌。我们还研究了染色中心的复制模式和Xist-RNA在不活跃的X-区域内的地形关系。这些例子表明，适当适应的3D-FISH/3D-SIM方法保留了核超微结构的关键特征，并且3D-SIM获得的信息产生了对功能核组织的新见解。
Three-dimensional structured illumination microscopy (3D-SIM) has opened up new possibilities to study nuclear architecture at the ultrastructural level down to the similar to 100 nm range. We present first results and assess the potential using 3D-SIM in combination with 3D fluorescence in situ hybridization (3D-FISH) for the topographical analysis of defined nuclear targets. Our study also deals with the concern that artifacts produced by FISH may counteract the gain in resolution. We address the topography of DAPI-stained DNA in nuclei before and after 3D-FISH, nuclear pores and the lamina, chromosome territories, chromatin domains, and individual gene loci. We also look at the replication patterns of chromocenters and the topographical relationship of Xist-RNA within the inactive X-territory. These examples demonstrate that an appropriately adapted 3D-FISH/3D-SIM approach preserves key characteristics of the nuclear ultrastructure and that the gain in information obtained by 3D-SIM yields new insights into the functional nuclear organization.