PyTom: Localize Macromolecules interactively

Overview

In many applications you may have no idea of the actual structure of your molecule of interest or you may want to deliberately not use any information that might be available. On these cases you will localize putatively identical particles interactively using an appropriate graphical tool, store the respective coordinates, and reconstruct the respective subvolumes.

Instead of reconstructing subvolumes using the coordinates you might also box them out from the tomogram used for particle picking directly - however, in PyTom we typically work with volumes of reduced resolution (binned) for all overview purposes and avoid reconstructing the un-binned tomogram because that would unnnecessarily block disk space. Moreover, CTF correction requires seperate reconstruction of subtomograms anyway and hence we strongly encourage following this route.

The EMAN2 package provides a rather neat interface for interactive particle picking or 'boxing'. Hence, we support importing coordinates of subtomograms localized in EMAN2 rather than programming our own solution. This coordinate list is then transferred to the native PyTom format for dealing with subtomograms (particle list).

Particle boxing using EMAN2's e2spt_boxer.py

EMAN2 provides support for boxing of subtomograms. For a detailed tutorial on the usage we refer to: EMAN2 single particle tomography tutorial.

e2spt_boxer.py "MyTomogram.em" -­-­inmemory
There is also a argument --yshort, which you must not use (this option applies to somewhat weird coordinate systems like the one from IMOD).

In the e2spt_boxer.py you can conveniently mark the centers of the particles of your interest. Before leaving the GUI you save the coordinates in a file, which is then converted for usage in PyTom, e.g., for reference-free subtomogram alignment and averaging.

Convert particle coordinates to particle list using bin/coords2PL.py

The script bin/coords2PL.py conveniently converts the coordinates stored in the EMAN2 boxer tool into a PyTom particle list (XML file). In this script you can also specify the future names of the to-be-generated subtomograms as well as the respective missing wedges.
pytom "MyPyTomPath"/bin/coords2PL.py -p "particleList" -c "CoordinateFile" -s "MySubtomogramNames" -w "MyWedge"
In detail, the parameters are: