Collection location: Franklin Parker Preserve, Woodland Twp., New Jersey, USA [Click for map]
> The pictured specimens probably represent two separate collections, both made by J. & N. Burghardt.
> Note the bulb of the larger fruiting body being parasitized by a turquoise-colored mold, something that RET has often observed in this species more often that other Amanitas — see the ecology section at www.amanitaceae.org/?Amanita%20subcokeri
> Both fruitbodies had a faint pleasant sweet odor
> Both specimens dropped spores — spore measurements & pix will be provided later
> The younger basidiome has been dried
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I heard of the 200 bps restriction for posting sequences to GenBank, but that must be a relatively recent policy (?), for I frequently see much shorter fragments when running BLASTn searches for matching a sequence in hand to a particular taxon ID in GenBank.
I am curious if the problem you saw with 175355 [i.e., (1) not being able to connect the right-hand end of ITS2 with the opening motif of nrLSU and (2) having a break in the LSU gene precluding a contiguous read] is exactly the same as in 179593… Perhaps that question can be partially answered by GenBank once the LSU sequences become available.
First, the two collections are 3 weeks apart. Second, as my notes indicate, the latter collection was found at a specific specific location in the Speedwell section of FPP, whereas this one is likely to have come from the Chatsworth section. If the heterogeneity problem is of exactly the same kind or even identical, then one might conclude (granted it’s only n=2) that there is one large population of A. subcokeri across a large swath of FPP. Since you have at least one example of a contiguous nrLSU read for subcokeri (HQ539747), the heterogeneity seen in this FPP material is not representative of the whole species. I wonder if you’ve ever come across a case in your study of Amanita wherein all sequenced examples of a particular species would repeatedly suffer from having non-homogenized nrDNA.
So far, my experience with LSU both as a barcode gene and as a phylogenetic tool has been a very positive one for boletes. It’s much cleaner than ITS when it comes to sequence scrambling problem, and in most cases wherein I suspected lack of a barcode gap for apparently different morphological species, the gene was eventually vindicated by a TEF-1 follow-up.
I was not aware that LSU is “a treacherous gene for tree building”. It would be interesting to know why given that for starters LSU doesn’t have the alignment problem associated with ITS.
First of all, it would have been helpful if 150 character sequences could be submitted to GenBank; however, they have to set a limit somewhere; and it is 200 characters.
It would have been very satisfying if I could have attached the 3’ end of ITS2 to the 5’ end of LSU; but the circumstances did not permit it.
It would be nice if something that doesn’t suffer from being part of the nrDNA repeat were the barcode. It might serve as a better barcode; however the investment in ITS is enormous and growing.
I have seen discussions of LSU as a “secondary barcode”; however, while usually less of a problem than ITS, it sometimes suffers from heterogeneity in a manner and to a degree that is undesirable and, as I’ve mentioned before, it is a treacherous gene for tree building.
Then there is bacterial cloning. I have run into a lab that can do it, but not to the successful extent that Dr. Hughes has done (e.g., the infraspecific hybrid that is a lavendula look-alike).
Linas has found value in adding rpb2 to his repertoire, at least in some cases where heterogeneity or failure to segregate probable species is/are problems. There are many problems, but it would be ridiculous to imagine we would not find some.
I very much appreciate your post.
Thank you for posting the news regarding the sequences for this obsie and that of 179593. I see that you have already added the two GB accession numbers to the subcokeri page of your website.
The heterogeneity problem frequently seen in nrDNA, particularly the ITS “gene”, is very annoying. I am sure that after having seen it many times over the years you eventually grew to accept it with total equanimity. Me… not so much. :-)
I think if the ITS locus is to eventually lose its proposed barcode gene “crown” to a better alternative, this issue (i.e., the state of being too variable for its own good) is going to be one of the major decisive factors to necessitate such a move. There is no reason why one shouldn’t expect getting consistently good results from different collections of the same species! On the other hand, crowning the new barcode gene will somehow need to address the immense complication and expense of retesting a large volume of taxa/vouchers on the global scale to build up its sequence database, so this may never happen.
This material has been received and accessioned to Rod’s herbarium. We have scheduled it for DNA sequencing.