Abstract：
:Cryo-electron tomography (cryoET) allows 3D structural information to be obtained from cells and other biological samples in their close-to-native state. In combination with subtomogram averaging, detailed structures of repeating features can be resolved. CryoET data is collected as a series of images of the sample from different tilt angles; this is performed by physically rotating the sample in the microscope between each image. The angles at which the images are collected, and the order in which they are collected, together are called the tilt-scheme. Here we describe a "dose-symmetric tilt-scheme" that begins at low tilt and then alternates between increasingly positive and negative tilts. This tilt-scheme maximizes the amount of high-resolution information maintained in the tomogram for subsequent subtomogram averaging, and may also be advantageous for other applications. We describe implementation of the tilt-scheme in combination with further data-collection refinements including setting thresholds on acceptable drift and improving focus accuracy. Requirements for microscope set-up are introduced, and a macro is provided which automates the application of the tilt-scheme within SerialEM.
journal_name
J Struct Bioljournal_title
Journal of structural biologyauthors
Hagen WJH,Wan W,Briggs JAGdoi
10.1016/j.jsb.2016.06.007subject
Has Abstractpub_date
2017-02-01 00:00:00pages
191-198issue
2eissn
1047-8477issn
1095-8657pii
S1047-8477(16)30113-7journal_volume
197pub_type
杂志文章abstract：:Titin is a giant protein responsible for passive-tension generation in muscle sarcomeres. Here, we used single-molecule AFM force spectroscopy to investigate the mechanical characteristics of a recombinant construct from the human cardiac-specific N2B-region, which harbors a 572-residue unique sequence flanked by two ...
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