What the fungi do I do with my ITS library? (Pt. 2)

Previously, I expressed some concern about size variation in my environmental fungal ITS PCR libraries. I'm still concerned about that, but I have an additional concern. The ITS region can't be aligned, and I'm partial to phylogenetic approaches to pretty much everything. So maybe ITS is not for me?

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  1. So, still two reasons to want a multiple sequence alignment of my "barcode" gene. 1) evolution! and 2) placement (even with uncertainty) of sequences that do not have an exact match in the database. It seems as though I won't be judged negatively should I switch to 28S (or LSU), but that I will have issues with taxonomic resolution and an inferior reference database, relative to ITS.

    Pat brings up another concern. Isn't there a lot of intra-genomic size variation in ITS?
  2. So, it seems like intragenomic ITS variation is no more of a concern with ITS than it is with 16S. So. maybe I will bring my ITS data to Pat's R workshop after all.

    So, here's my final take on the issue:
  3. We can't turn ITS into a good phylogenetic marker, but we can fill up the 28S database. Maybe taxonomic resolution isn't perfect, but what is?
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