Identifying adapter sequence to trim for Illumina IIx data

Question

[shpak.max]

I have two question concerning using trimmomatic to trim adapter sequences.

The first is which adapter sequences to use. My sequence data is Illumina Genome Analyzer IIx, and I'm not sure which (if any) of the sequences is applicable:

https://github.com/timflutre/trimmomatic/tree/master/adapters

The example in the user guide is TruSeq3-PE.fa, while Illumina's website suggests AGAT...sequences (though I'm not sure if this applies to the IIx sequencing platform vs something more recent):

https://support.illumina.com/bulletins/2016/12/what-sequences-do-i-use-for-adapter-trimming.html

My second question deals with how trimmomatic handles orders of operations. If I apply the following commands, will trimmomatic first trim the adapters and then crop the first 50 remaining bases after the adapters are clipped (assuming I'm using the correct one, the one there now is just a placeholder), or do I need to first trim the adapters, save the new fastq file, and then apply trimmomatic with CROP?

for infile in *_1.fastq
    do
    base=$(basename ${infile} _1.fastq)
    ./trimmomatic PE ${infile} ${base}_2.fastq \
        ${base}_1.trim.fastq ${base}_1un.trim.fastq \
        ${base}_2.trim.fastq ${base}_2un.trim.fastq \
    ILLUMINACLIP:TruSeq3-PE.fa:2:30:10:2:True LEADING:3 TRAILING:3 MINLEN:36\
   CROP:50
  done

Answer 1

[GenoMax]

See: How to figure out adapter sequence for Illumina reads?

[shpak.max]

I saw the discussion that you linked to before posting and didn't find that that the suggestions there applied in my case. I ran FastQC and the Adapter Content section shows trace frequencies (<< 1%) for SOLID Small RNA Adapter. I'm not sure if this makes sense for the type of data set that I have, nor do I know which (if any) of the adapter sequences in this list it corresponds to:

https://github.com/timflutre/trimmomatic/tree/master/adapters

[swbarnes2]

If it's just trace, why worry about it? It it makes no sense, and it's just trace, it's probably just some random sequence that just happens to kind of match an adapter in the database.