Overview of a single-particle reconstruction algorithm1 for helical filaments as applied to TMV.
Twenty-nine filaments were extracted from a raw TMV dataset and used to create a SPIDER stack of 3924 highly-overlapping square boxes. Multi-Reference Alignment (MRA) was performed on these particles using 180 2D projections of an initial starting volume (shown in gold; produced with Phoelix) as references. Euler angles were then assigned and used to backproject the images, creating an asymmetric 3D volume (green). The axial rise and twist were calculated by least squares and used to impose helical symmetry on the asymmetric volume, producing a helically symmetric one (coral). Projections of this volume were then used as references for the next round of MRA. Pictured in the lower-left corner are the first, seventh, fourteenth, and twentieth volumes from a preliminary experiment of 20 iterations. 1. E.H. Egelman. Ultramicroscopy 85 (2000) 225-234 Kathryn Thompson, NRAMM