3. Alternatively, the 3DFSC program can be run through a web
interface, as described in steps 3a–3h. The web address for
running directional resolution estimation online is:
https://3dfsc.salk.edu
(a) Click “Register” on the navigation bar and follow the
instructions to create an account.
(b) Once an account is created, navigate to the processing
form via the “Submit job” link.
(c) The online job submission requires the user to enter a
valid email address and other parameters in the form.
(d) You must upload a unique job name, two half-maps (.mrc
format), a full map (also .mrc format) (see Note 13), and
an appropriate pixel size. Click “Submit job”.
(e) Optional parameters include submission of a mask file (.
mrc format), appropriate cone angle, cutoff for FSC, a
threshold for sphericity and a high-pass filter
(in Ångstroms) (see Note 10).
(f) You should receive an email to confirm your processing
job. If you do not receive an email, please check your spam
folder.
(g) When your job is complete, you will receive another email
with a link to view the results.
(h) A typical output will include a compressed folder that
contains the input files used and a range of outputs.
These can be downloaded from the website. There is a
password on the output page that needs to be used in
order to open and decompress the folder. Once the folder
is decompressed, the outputs will be similar to the outputs
within the command-line version.
4. Inspect the outputs. These include the 3D FSC volume (see
Figs. 3 and 4), the global FSC curve overlaid onto a histogram
plot computed from all individual angular FSC values (individual angular FSC curves corresponding to Æ1 standard deviation of the FSC values within the histogram are also displayed
on the plot, see Fig. 5), and raw data used for computing these
plots are provided as spreadsheets (see Note 14).
5. Open UCSF Chimera and load the “3DFSCPlot_Chimera.
cmd” file from the output directory. This script is designed
for convenient visualization using Chimera, but individual mrc
files can also be opened independently. Once executed, the
Chimera visualization program opens with the full map and
the 3D FSC volume along with an FSC plot that compares
global resolution with the Z-directional resolution relative to
the viewing angle (see Fig. 4).
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interface, as described in steps 3a–3h. The web address for
running directional resolution estimation online is:
https://3dfsc.salk.edu
(a) Click “Register” on the navigation bar and follow the
instructions to create an account.
(b) Once an account is created, navigate to the processing
form via the “Submit job” link.
(c) The online job submission requires the user to enter a
valid email address and other parameters in the form.
(d) You must upload a unique job name, two half-maps (.mrc
format), a full map (also .mrc format) (see Note 13), and
an appropriate pixel size. Click “Submit job”.
(e) Optional parameters include submission of a mask file (.
mrc format), appropriate cone angle, cutoff for FSC, a
threshold for sphericity and a high-pass filter
(in Ångstroms) (see Note 10).
(f) You should receive an email to confirm your processing
job. If you do not receive an email, please check your spam
folder.
(g) When your job is complete, you will receive another email
with a link to view the results.
(h) A typical output will include a compressed folder that
contains the input files used and a range of outputs.
These can be downloaded from the website. There is a
password on the output page that needs to be used in
order to open and decompress the folder. Once the folder
is decompressed, the outputs will be similar to the outputs
within the command-line version.
4. Inspect the outputs. These include the 3D FSC volume (see
Figs. 3 and 4), the global FSC curve overlaid onto a histogram
plot computed from all individual angular FSC values (individual angular FSC curves corresponding to Æ1 standard deviation of the FSC values within the histogram are also displayed
on the plot, see Fig. 5), and raw data used for computing these
plots are provided as spreadsheets (see Note 14).
5. Open UCSF Chimera and load the “3DFSCPlot_Chimera.
cmd” file from the output directory. This script is designed
for convenient visualization using Chimera, but individual mrc
files can also be opened independently. Once executed, the
Chimera visualization program opens with the full map and
the 3D FSC volume along with an FSC plot that compares
global resolution with the Z-directional resolution relative to
the viewing angle (see Fig. 4).
170
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