Using ESCOTT via Docker
========================

Requirements
------------

You need to have docker installed on your machine. You can consult the
following page for this: https://docs.docker.com/get-docker/

I am assuming some basic familiarity with Linux/Unix/MacOS terminal
commands.

Let’s start our favorite terminal app.

You must create a folder called docker-tutorial and go to that empty
folder:

.. code:: bash

   mkdir docker-tutorial
   cd docker-tutorial

Getting the example input data
------------------------------

Let’s download the sample data provided in the PRESCOTT repository for
this exercise. First, we will download the multiple sequence alignment
file in fasta format:

.. code:: bash

   wget http://gitlab.lcqb.upmc.fr/tekpinar/PRESCOTT/raw/8d766d4d11af0e93c9da8fc2c5cc1bfc457d2936/data/aliBLAT.fasta

If you don’t have wget, you can try the same command with curl:

.. code:: bash

   curl http://gitlab.lcqb.upmc.fr/tekpinar/PRESCOTT/raw/8d766d4d11af0e93c9da8fc2c5cc1bfc457d2936/data/aliBLAT.fasta >aliBLAT.fasta

Please verify that the aliBLAT.fasta file is in the folder.

Now, we will download the PDB (Protein Databank) file for BLAT:

.. code:: bash

   wget http://gitlab.lcqb.upmc.fr/tekpinar/PRESCOTT/raw/8d766d4d11af0e93c9da8fc2c5cc1bfc457d2936/data/blat-af2.pdb

Getting the docker image
------------------------

In order to make sure that the docker is installed:

.. code:: bash

   sudo docker -h

If it shows you a list of options, you are on a good track. On MacOS,
you may not need ‘sudo’ word before the docker command at all.

Now, let's pull the image to our computer:

.. code:: bash

   sudo docker pull tekpinar/prescott-docker:v1.6.0
   

Single point mutation calculations 
----------------------------------

.. code:: bash

   sudo docker run -ti --rm --mount type=bind,source=$PWD,target=/home/tekpinar/research/myexample \
   tekpinar/prescott-docker:v1.6.0

You are in the container (your virtual operating system) now. You
created a folder called myexample in your container with the previous
command. Let’s change to that folder.

Note: If you are running it on a recent MacOS version such as Ventura or Sonoma, you may need to add '--platform linux/amd64'
after "run" command above. 

.. code:: bash

   cd ../myexample/

When you check the data in that folder with ‘ls’ command, you are
supposed to see aliBLAT.fasta and blat-af2.pdb files. Basically, your
docker-tutorial folder on the host system and myexample folder on the
docker container are pointing to the same place.

Obtaining the entire single point mutation landscape
^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^
In this step, we will use only evolutionary information from an MSA file:

.. code:: bash

   escott aliBLAT.fasta 

After a few minutes of calculation, you must see at least two files named
BLAT_normPred_evolCombi.txt and BLAT_normPred_evolCombi.png. You have
the entire single point mutational landscape of BLAT protein in these
files.

If you want to utilize structural information (highly recommended) as well as 
evolutionary information:

.. code:: bash

   escott aliBLAT.fasta --pdbfile blat-af2.pdb

Predicting the effect of a subset of single point mutations
^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^
If you are interested in only a bunch of single point mutations, 
you have to prepare a mut file. The format is a simple text file and 
each line contains a single point mutation such as D26A....
Fortunately, we have an example mut in data folder of PRESCOTT repository.

.. code:: bash

   wget http://gitlab.lcqb.upmc.fr/tekpinar/PRESCOTT/raw/master/data/Stiffler_2015_BLAT_ECOLX_singles.mut

Mutation file is a simple text file and we use ".mut" file extension for it.

Similar to the previous step, there are two possible ways to do the calculations: with or without
structural information. First, let's do it without structural information: 

.. code:: bash
   
   escott aliBLAT.fasta -m Stiffler_2015_BLAT_ECOLX_singles.mut 

You can include structural information in the following way:

.. code:: bash
   
   escott aliBLAT.fasta --pdbfile blat-af2.pdb \
   -m Stiffler_2015_BLAT_ECOLX_singles.mut

You will have BLAT_normPred_evolCombi.txt file in your folder. However, the output 
format is completely different from the entire mutational landscape scanning file.
Each line of this file is a mutation and its predicted effect separated by a space.
In addition, you won't have a png file like in the previous case. 

Multiple point mutation calculations 
------------------------------------
Sometimes, we need to see effects of double or triple mutations. ESCOTT can 
perform calculations if you provide a mut file. Double or triple mutations should be separated 
with a colon and do not forget to add a new line after the last line of mutations. In this case, the mut file must
have the following format:

.. code:: bash

   E26D:Y44R
   E56N:A77F:H94V
   S96C:E26A:Y44C


The first line of the text file is impact of a double mutation, namely combined effect of E26D and Y44R.
The second and the third lines are the impacts of the triple mutations. As you can see, the mutations are 
separated by a colon(:) character. 
The output file will be in a similar format. Each line will contain the multiple 
mutations separated by colons. We provide the example multiple point mutation file 
for BLAT in the data folder of this repository with the file name Stiffler_2015_BLAT_ECOLX_multiples.mut. 
You can download the file in Linux/Unix operating systems with the following command:

.. code:: bash

   wget http://gitlab.lcqb.upmc.fr/tekpinar/PRESCOTT/raw/master/data/Stiffler_2015_BLAT_ECOLX_multiples.mut


Running several jobs using docker
---------------------------------
If you want to use docker in a more automated way for several proteins,
you can call docker within a bash script. 

.. code:: bash

   sudo docker run --rm -v $PWD:/home/tekpinar/research/lcqb tekpinar/prescott-docker:v1.6.0 escott aliBLAT.fasta --pdbfile blat-af2.pdb

Note: It is very important to have aliBLAT.fasta and blat-af2.pdb files in your local folder when you call docker like an executable. 
Typically, I create a folder for each protein that contain the alignment and the structure. Then, I change the path to each folder with 'cd' 
command inside bash script and execute the command above in each local folder. 



