Hello,

I am a university student currently trying to use Trimmomatic for the first time and have attempted to use a command line I have formed from using the official Trimmomatic manual and some other online resources. I am very new to Bioinformatics and especially new to programming, so I would appreciate if you can try to defer from using any jargon so I can understand your points easier. Thank you.

I am using Trimmomatic-0.36 and also undertaking this command on a remote server, rather than my local drive.
The command line I have most recently used is the following:

java -jar $trimmomatic PE -phred33 /home/will/240_CTTGTA_L004_R1_001.fastq.gz /home/will/C240_CTTGTA_L004_R2_001.fastq.gz C240_CTTGTA_L004_R1.TRIM.fastq.gz C240_CTTGTA_L004_R1.UNTRIM.fastq.gz C240_CTTGTA_L004_R2.TRIM.fastq.gz C240_CTTGTA_L004_R2.UNTRIM.fastq.gz ILLUMINACLIP:TruSeq-PE.fa:2:30:10

I have also tried to use the command line without including a path to my original read files, seen below:

 java -jar $trimmomatic PE -phred33 240_CTTGTA_L004_R1_001.fastq.gz C240_CTTGTA_L004_R2_001.fastq.gz C240_CTTGTA_L004_R1.TRIM.fastq.gz C240_CTTGTA_L004_R1.UNTRIM.fastq.gz C240_CTTGTA_L004_R2.TRIM.fastq.gz C240_CTTGTA_L004_R2.UNTRIM.fastq.gz ILLUMINACLIP:TruSeq-PE.fa:2:30:10

Lastly, I have included the path to the adapters file within the Trimmomatic-0.36 file:

 java -jar $trimmomatic PE -phred33 240_CTTGTA_L004_R1_001.fastq.gz C240_CTTGTA_L004_R2_001.fastq.gz C240_CTTGTA_L004_R1.TRIM.fastq.gz C240_CTTGTA_L004_R1.UNTRIM.fastq.gz C240_CTTGTA_L004_R2.TRIM.fastq.gz C240_CTTGTA_L004_R2.UNTRIM.fastq.gz ILLUMINACLIP:/usr/local/bin/Trimmomatic-0.36/adapters/TruSeq-PE.fa:2:30:10

For all 3 of the inputs I have used, the terminal has returned this error message:

java.io.FileNotFoundException: /usr/local/bin/Trimmomatic-0.36/adapters/TruSeq-PE.fa (No such file or directory)
at java.io.FileInputStream.open0(Native Method)
at java.io.FileInputStream.open(FileInputStream.java:195)
at java.io.FileInputStream.<init>(FileInputStream.java:138)
at org.usadellab.trimmomatic.fasta.FastaParser.parse(FastaParser.java:54)
at org.usadellab.trimmomatic.trim.IlluminaClippingTrimmer.loadSequences(IlluminaClippingTrimmer.java:110)
at org.usadellab.trimmomatic.trim.IlluminaClippingTrimmer.makeIlluminaClippingTrimmer(IlluminaClippingTrimmer.java:71)
at org.usadellab.trimmomatic.trim.TrimmerFactory.makeTrimmer(TrimmerFactory.java:32)
at org.usadellab.trimmomatic.Trimmomatic.createTrimmers(Trimmomatic.java:59)
at org.usadellab.trimmomatic.TrimmomaticPE.run(TrimmomaticPE.java:536)
at org.usadellab.trimmomatic.Trimmomatic.main(Trimmomatic.java:80)
Exception in thread "main" java.io.FileNotFoundException: 240_CTTGTA_L004_R1_001.fastq.gz (No such file or directory)
at java.io.FileInputStream.open0(Native Method)
at java.io.FileInputStream.open(FileInputStream.java:195)
at java.io.FileInputStream.<init>(FileInputStream.java:138)
at org.usadellab.trimmomatic.fastq.FastqParser.parse(FastqParser.java:135)
at org.usadellab.trimmomatic.TrimmomaticPE.process(TrimmomaticPE.java:264)
at org.usadellab.trimmomatic.TrimmomaticPE.run(TrimmomaticPE.java:539

I have looked online for solutions and attempted to fix my command line, but so far nothing has worked. Any help would be greatly appreciated and would help me massively. If anymore information is needed, please do not hesitate to ask.

Many thanks,

Will.



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