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To obtain a 3D model, 15′352 particles were selected from the 279 micrographs recorded and subjected to reference free alignment and classification as detailed in the Materials and Methods.
I offer a limited number of free alignment coaching calls per week.
Finally, averages were generated for selected classes using reference free alignment procedures as implemented in the XMIPP software package [22].
Genomic comparisons between the available vtx-phages sequences have been performed by using the BLAST algorithm available at NCBI [ 28] and the Mauve free alignment software [ 24].
Our base shifting algorithm is a greedy search for free alignment positions to the left and right of the insertion which we believe to be equivalent to NAST.
Alignments of predicted peptide sequences for each gene were performed in CLC Genomics Workbench using the Create Alignment tool (Gap open cost = 10, Gap extension cost = 1, End gap cost = Free, Alignment mode = Very accurate).
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Next, we use cross-correlation and reference-free alignment, to align each NPC image with sub-pixel resolution.
The two tilt series were first independently aligned using marker-free alignment [93] and then merged by patch correlation and volume warp using IMOD [94] to produce the raw, dual axis tomogram.
Prior to beginning data collection, coma-free alignment was carried out to align the beam to the column optical axis with the assistance of a Zemlin tableau (Glaeser et al., 2011).
In all, 162 particles on plasmid and 2600 on short DNA were windowed, centred and aligned using the reference-free alignment procedure.
Single particles were aligned and classified using reference-free alignment and k-means classification procedures implemented in SPARX (Hohn et al., 2007).
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