Using trimmomatic on multiple paired-end read files

Question

[bioinformaticssrm2011]

I need help to write a for loop to run Trimmomatic tool for quality trimming of paired end fastq files. I need to write a for loop so that I can run an executable for all multiple files.

Input PE files looks like - C1_R1.fastq

C1_R2.fastq

C2_R1.fastq

C2_R2.fastq

C3_R1.fastq

C3_R2.fastq

T1_R1.fastq

T1_R2.fastq

T2_R1.fastq

T2_R2.fastq

T3_R1.fastq

T3_R2.fastq

To run trimmomatic for the paired reads corresponding to C1_R1.fastq and C1_R2.fastq, the following command works:

java -jar ~/Trimmomatic-0.36/trimmomatic-0.36.jar PE -phred33 C1_R1.fastq C1_R2.fastq C1_R1_paired.fastq C1_R1_unpaired.fastq C1_R2_paired.fastq C1_R2_unpaired.fastq LEADING:3 TRAILING:3 SLIDINGWINDOW:4:15 MINLEN:35

I have tried using this thread, but I am unable to understand.

Any help please! Thanks!

[WouterDeCoster]

Hi bioinformaticssrm2011,

The second command I posted on seqanswers (with an echo in front) should show you the command how it would be executed. That way you can figure out what is wrong.

Shouldn't you use this command in paired end mode?

java -jar <path to trimmomatic.jar> PE [-threads <threads] [-phred33 | -phred64] [-trimlog <logFile>] >] [-basein inputBase> | <input 1> <input 2>] [-baseout <outputBase> | <unpaired output 1> <paired output 2> <unpaired output 2> <step 1> ...

[bioinformaticssrm2011]

I got an error, if you see that thread

[WouterDeCoster]

I've seen that error. Now read what I wrote about using the echo statement and figuring out what's wrong.

[bioinformaticssrm2011]

About echo, I am new to this, so unable to understand it. Sorry

[WouterDeCoster]

If you don't know the echo command you need to start with following a command line tutorial, that will make everything less painful.

[shengweima]

this is my python script

#!/usr/bin/env python
# -*- coding: utf-8 -*-

import subprocess

with open('/data2/masw_data/Nanda/5A_and_5D/bbmap_config.txt', 'r') as f:
    for line in f:
        line = line.strip().split()
        ref, in1, in2, outm = line
        proc = subprocess.Popen(['/data1/masw/bbmap/bbmap.sh', ref, in1, in2, outm, 'threads=20', 'minid=0.70', 'nodisk'], shell=False)
        proc.wait()
f.writelines()

[GenoMax]

shengweima : Did you post in the wrong thread or were you presenting an example?

Answer 1

[lakhujanivijay]

Answer 2

[agata88]

You need to specify the directories to your input/output files :

"inputdirectory" directory "processed" directory (as output) "log" directory for log information "program" directory with Trimmomatic jar file

#! python
import os
import sys
import subprocess 

for fileR1 in os.listdir(inputdirectory): 
    dividing = fileR1.split(".")
    if ("R1" in fileR1) :
        fileR2 = fileR1.replace('R1', 'R2')
        if os.path.isfile(inputdirectory + fileR2) :
            dividing1 = fileR2.split(".")
            log1 = dividing[0]
            output1 = dividing[0]
            output2 = dividing1[0]
            subprocess.call("java -jar " + program + "trimmomatic-0.35.jar PE -threads 12 -phred33 " + inputdirectory + fileR1 + " " + inputdirectory + fileR2 + " " + processed + output1 +"_trimmed.fastq.gz " +  "output_forward_unpaired.fq.gz "  + processed + output2 + "_trimmed.fastq.gz " + "output_reverse_unpaired.fq.gz " + " LEADING:3 TRAILING:3 SLIDINGWINDOW:4:15 MINLEN:35" +" >" + log + log1 + "_trimmomatic.txt" + " 2>&1", shell=True)

Hope it helps :) Best,

Agata

[bioinformaticssrm2011]

hi, When I was trying to use above script, I got an error.

    #! python
    import os
    import sys
    import subprocess 
    inputdirectory = "/home/shashank/Desktop/patel_chest_NSC/inputdirectory"
   processed = "/home/shashank/Desktop/patel_chest_NSC/processed"
   log="/home/shashank/Desktop/patel_chest_NSC/log"
   for fileR1 in os.listdir(inputdirectory): 
        dividing = fileR1.split(".")
        if ("R1" in fileR1) :
            fileR2 = fileR1.replace('R1', 'R2')
            if os.path.isfile(inputdirectory + plikR2) :
                dividing1 = fileR2.split(".")
                log1 = dividing[0]
                output1 = dividing[0]
                output2 = dividing1[0]
                subprocess.call("java -jar " + program + "~/Trimmomatic-0.36/trimmomatic-0.36.jar PE -threads 12 -phred33 " + inputdirectory + fileR1 + " " + inputdirectory + fileR2 + " " + processed + output1 +"_trimmed.fastq.gz " +  "output_forward_unpaired.fq.gz "  + processed + output2 + "_trimmed.fastq.gz " + "output_reverse_unpaired.fq.gz " + " LEADING:3 TRAILING:3 SLIDINGWINDOW:4:15 MINLEN:35" +" >" + log + log1 + "_trimmomatic.txt" + " 2>&1", shell=True)

Error is-

   Traceback (most recent call last):
  File "python.py", line 10, in <module>
    if os.path.isfile(inputdirectory + plikR2) :
NameError: name 'plikR2' is not defined

[agata88]

Please replace 'plikR2' to 'fileR2'.

[bioinformaticssrm2011]

Hi, thanks for your help. But I got an error

File "python.py", line 8 if ("R1" in fileR1): ^ IndentationError: unindent does not match any outer indentation level

than I checked the indent and tried to solve it, and later I got another error-

Traceback (most recent call last): File "python.py", line 6, in <module> for fileR1 in os.listdir(input): TypeError: coercing to Unicode: need string or buffer, builtin_function_or_method found

[WouterDeCoster]

input shouldn't be used as a variable name in python3. Make it inputfile or something like that.

[bioinformaticssrm2011]

Thats helpful.

but another error pop up-

Traceback (most recent call last): File "python.py", line 6, in <module> for fileR1 in os.listdir(inputfile): NameError: name 'inputfile' is not defined

[WouterDeCoster]

That's because you didn't define the variable.

[agata88]

Yes, that is correct it shouldn’t be 'input' , I forgot about that when I was changing names... sorry ... going to change that right now.

[agata88]

Changed, now it should be ok, also the indentation was repaired.

[Safa.A]

Hi, Thanks for you python script. I am trying to run this script but I have it not running or giving me any output. and also, I have no error come at any step. when I run it the result is nothing. Can you help me, please! Thanks

[agata88]

Hi, check the indentations. maybe during copy/paste something was shifted, best, Agata