Trim Galore Bug?

Question

[datascientist28]

I'm having some trouble with my output files after I use trim galore. I think I might have found the issue but I'm curious if anyone has had a similar problem. The command I give trim galore specifies paired end mode but for some reason cut adapt is trimming in single end mode. Anyone know if this is a bug or am I thinking about this wrong?

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Below is my trim_galore report . I highlighted in bold my what I mean...

SUMMARISING RUN PARAMETERS

Input filename: Regeneration_ATAC_1hpa-1_R1.fastq.gz

Trimming mode: paired-end

Trim Galore version: 0.4.1

Cutadapt version: 1.10

Quality Phred score cutoff: 20

Quality encoding type selected: ASCII+33

Adapter sequence: 'CTGTCTCTTATA' (Nextera Transposase sequence; user defined)

Maximum trimming error rate: 0.1 (default)

Minimum required adapter overlap (stringency): 1 bp

Minimum required sequence length for both reads before a sequence pair gets removed: 20 bp

All sequences will be trimmed by 1 bp on their 3' end to avoid problems with invalid paired-end alignments with Bowtie 1

Running FastQC on the data once trimming has completed

Output file will be GZIP compressed

.

This is cutadapt 1.10 with Python 2.7.11

Command line parameters: -f fastq -e 0.1 -q 20 -O 1 -a CTGTCTCTTATA Regeneration_ATAC_1hpa-1_R1.fastq.gz

Trimming 1 adapter with at most 10.0% errors in single-end mode ...

Finished in 1463.06 s (25 us/read; 2.45 M reads/minute).

[GenoMax]

Can you post the full command line? Based on the log above it appears that you only have one fastq file.

[datascientist28]

trim_galore --paired --phred33 --fastqc --nextera --trim1 Regeneration_ATAC_1hpa-1_R1.fastq.gz Regeneration_ATAC_1hpa-1_R2.fastq.gz

[GenoMax]

I am not a trim galore user but looking at the manual it does not appear that --paired option is used unless you also specify --length option.

--paired This option performs length trimming of quality/adapter/RRBS trimmed reads for paired-end files. To pass the validation test, both sequences of a sequence pair are
required to have a certain minimum length which is governed by the option --length (see above).

[igor]

I think it's supposed to be --trim-1, not --trim1.

[ofonov]

I have the same issue, does anyone have any idea what is going on?

[datascientist28]

Read @dariber's comment. It's all fine. That's just how trim galore runs

Answer 1

[dariober]

I think it's all fine. I think trim_galore in paired end mode runs trim_galore/cutadapt in single end mode for read 1 and read 2 independently and without discarding any read (hence the single-end log you see). Then, in a second pass, it removes read pairs where either read 1 or read 2 fail to pass the length threshold.

You might want to check with Felix, the developer, he's usually very quick and helpful answering questions.