Classification example

Download data and reference

Download the test data set comprising 10,000 ribosome images as deposited by Joachim Frank at the EBI-EMDB from here. The corresponding metadata is stored in this PDF file. Save both files in your working directory.

We will use EMDB entry 1056 as a reference, save this file in the same working directory. Note this reference has the same pixel size (2.8A) and the same box size (130x130) as the data set above. Therefore, no re-scaling or windowing operations are necessary.

Unpack the data as follows:

tar -xf J-Frank_70s_real_data.tar
gunzip emd_1056.map.gz
mv emd_1056.map emd_1056.mrc

10,000 70S ribosomes wi/wo EFG

The test data set that will be used below was provided by Joachim Frank and is stored at the EBI-EMDB.

Prepare the input STAR file

From the PDF provided by Joachim Frank select the following (14) lines and save them in a text file called defocus.dat

1 3 1347.0 1347.0 21580.
2 3 505.00 1852.0 24833.
3 3 989.00 2841.0 26450.
4 3 857.00 3698.0 28320.
5 3 475.00 4173.0 30993.
6 3 349.00 4522.0 33150.
7 3 478.00 5000.0 34588.
8 3 1242.0 6242.0 21580.
9 3 713.00 6955.0 24833.
10 3 1255.0 8210.0 26450.
11 3 1022.0 9232.0 28320.
12 3 304.00 9536.0 30993.
13 3 232.00 9768.0 33150.
14 3 232.00 10000. 34588.

Then save the following lines as a file called make_star.csh

#!/usr/bin/env csh
ls -l win/*dat | awk '{print $NF}' >imagelist
#
relion_star_loopheader rlnImageName rlnMicrographName rlnDefocusU rlnVoltage rlnSphericalAberration rlnAmplitudeContrast > all_images.star
#
set ngr = 14
set gr = 0
while ($gr < $ngr)
 @ gr++
 set nn=`head -n $gr defocus.dat  | tail -1 | awk '{print int($3)}'`
 set tot=`head -n $gr defocus.dat | tail -1 | awk '{print int($4)}'`
 set def=`head -n $gr defocus.dat | tail -1 | awk '{print $5}'`
 head -n ${tot} imagelist | tail -n ${nn} |awk -v"def=$def" -v"gr=$gr" '{print $1, gr, def, 200, 2, 0.1}' >> all_images.star
end

And execute it to generate the input STAR file with all image names and CTF information, using the command:

csh make_star.csh