Trimming adaptors with trimmomatic
0
0
Entering edit mode
4.8 years ago
Rimma ▴ 30

I checked my reads with fastQC and found an overrepresentation of Illumina adpaters: https://ibb.co/ftDW6sK enter image description here So I trimmed it using trimmomatic and following command (75 bases Single end reads):

java -jar <path>trimmomatic-0.39.jar SE input.fastq output.fastq ILLUMINACLIP:TruSeq2-SE:2:30:10

and the same with TruSeq3 adapters:

java -jar <path>trimmomatic-0.39.jar SE input.fastq output.fastq ILLUMINACLIP:TruSeq3-SE:2:30:10

However, the output I'm getting is always the same:

Input Reads: 48046117 Surviving: 48046117 (100.00%) Dropped: 0 (0.00%)    
TrimmomaticSE: Completed successfully
  

I'm new to bioinformatics so could you please tell me if there is smth I'm missing why trimmomatic doesn't cut off any of the reads?

RNA-Seq trimming • 2.4k views
ADD COMMENT
0
Entering edit mode

Have the reads been trimmed/ cleaned before, or was this the first time?

Did you run fastqc after to see if the adapters were removed?

ADD REPLY
0
Entering edit mode

I took these files from SRA so I don't know if they were trimmed before. Yes, I checked it looks the same as before trimming...

ADD REPLY
0
Entering edit mode

It may have to do with the CROP and MINLEN parameters. How long are your reads?

ADD REPLY
0
Entering edit mode

They are 75 bases long, the same before and after trimming

ADD REPLY
0
Entering edit mode

The adaptors from illumina have different sequences depending on the version of the kit used for the library preparation. It is possible you are just trimming the wrong sequence. As they have mentioned before, check out the "overrepresented sequences" found at the bottom of the FastQC report and you will know the version of the kit used. Then check for the right sequences and include them in the "TruSeq3-SE" you are using. And include a minim length to drop the reads that are too short after trimming. It should fix your problem.

ADD REPLY

Login before adding your answer.

Traffic: 1969 users visited in the last hour
Help About
FAQ
Access RSS
API
Stats

Use of this site constitutes acceptance of our User Agreement and Privacy Policy.

Powered by the version 2.3.6