PT  - JOURNAL ARTICLE
AU  - Tanmay A.M. Bharat
AU  - Sjors H.W. Scheres
TI  - Sub-tomogram averaging in RELION
AID  - 10.1101/030544
DP  - 2015 Jan 01
TA  - bioRxiv
PG  - 030544
4099  - http://biorxiv.org/content/early/2015/11/03/030544.short
4100  - http://biorxiv.org/content/early/2015/11/03/030544.full
AB  - Electron cryo-tomography (cryo-ET) and sub-tomogram averaging allow structure determination of macromolecules in situ, and are gaining in popularity for initial model generation for single-particle analysis. We describe herein, a protocol for sub-tomogram averaging from cryo-ET data using the RELION software. We describe how to calculate newly developed three-dimensional models for the contrast transfer function and the missing wedge of each sub-tomogram, and how to use these models for regularized-likelihood refinement. This approach has been implemented in the existing workflow for single-particle analysis, so that users may conveniently tap into existing capabilities of the RELION software. As example applications, we present analyses of purified hepatitis B capsid particles and S. cerevisiae 80S ribosomes. In both cases, we show that following initial classification, sub-tomogram averaging in RELION allows de novo generation of initial models, and provides high-resolution maps where secondary structure elements are resolved.