Automated tilt series alignment and tomographic reconstruction in IMOD.
Automated tilt series alignment and tomographic reconstruction in IMOD.
复制标题
IMOD中的自动倾斜系列对齐和断层摄影重建。
DOI：
10.1016/j.jsb.2016.07.011
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发表时间：
2017-02
影响因子：
3
通讯作者：
Held SR
中科院分区：
文献类型：
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作者：
Mastronarde DN;Held SR
在IMOD软件包中，现在可以实现自动层析成像重建，包括围绕两个正交轴拍摄的层析成像的合并。一些发展使生产高质量的层析成像成为可能。当使用基准标记进行对准时，将自动选择要跟踪的标记；如果有过多的可用标记，则选择一个分布良好的子集，它最有可能跟踪良好。标记位置通过应用边缘增强索贝尔过滤器进行细化，这导致塑料嵌入样品的校准误差提高20%，冷冻水合样品的校准误差提高10%。采用鲁棒拟合方法，在拟合误差计算中对外围点给予较少权重或不给予权重，从而获得对准解，使自动跟踪产生的异常点对对准的影响很小。在合并两幅双轴层析图时，根据局部斑块之间的相关性来改进它们之间的对齐；开发了一种结构测量方法，使结构不充分的斑块无法给出准确的相关性，现在可以自动排除。我们还开发了一个脚本，用于运行重建过程中的所有步骤，具有灵活的参数设置机制，并且我们在IMOD中的Etomo程序中添加了一个用于批量处理倾斜系列的用户界面。批处理与交互式处理完全兼容，即使在自动化不完全成功的情况下也可以提高效率，因为用户可以将精力集中在需要人工干预的步骤上。
Automated tomographic reconstruction is now possible in the IMOD software package, including the merging of tomograms taken around two orthogonal axes. Several developments enable the production of high-quality tomograms. When using fiducial markers for alignment, the markers to be tracked through the series are chosen automatically; if there is an excess of markers available, a well-distributed subset is selected that is most likely to track well. Marker positions are refined by applying an edge-enhancing Sobel filter, which results in a 20% improvement in alignment error for plastic-embedded samples and 10% for frozen-hydrated samples. Robust fitting, in which outlying points are given less or no weight in computing the fitting error, is used to obtain an alignment solution, so that aberrant points from the automated tracking can have little effect on the alignment. When merging two dual-axis tomograms, the alignment between them is refined from correlations between local patches; a measure of structure was developed so that patches with insufficient structure to give accurate correlations can now be excluded automatically. We have also developed a script for running all steps in the reconstruction process with a flexible mechanism for setting parameters, and we have added a user interface for batch processing of tilt series to the Etomo program in IMOD. Batch processing is fully compatible with interactive processing and can increase efficiency even when the automation is not fully successful, because users can focus their effort on the steps that require manual intervention.