I trimmed a full length ‘seed’ reference to the V4 region using the blog on customization as follows:
- Short perl script (yes, perl…) to pull the V4 region from one full length SSU acantharia. Output as Acantharia_V4.fasta.
- align.seqs(fasta=Acantharia_V4.fasta, reference=silva.seed_v132.align)
- pcr.seqs(fasta=silva.seed_v132.align, start=6388, end=13861, keepdots=FALSE)
- Rename and format using bash (sed)
After trimming to the V4 region, none of my reads aligned to the trimmed V4 version. I then tried aligning to the full-length reads seed and got the same results; no alignments. However, when I try assigning taxonomy using the regular full length silva database, things work just fine.
Any troubleshooting advice? My end goal is to produce a V4 reference for taxonomic assignments. Preferably out of a database that comes with a taxonomy .txt file as both are required for taxonomic assignment.