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OfflineAlan RockefellerM
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Panaeolus phylogenetic tree * 3
    #14123739 - 03/15/11 05:03 AM (12 years, 11 months ago)

I just made this rough draft phylogenetic tree of Panaeolus using mega5 and information from Genbank.

http://plantobserver.org/Panaeolus.phylogenetic.tree.pdf

FASTA file:

http://plantobserver.org/Panaeolus.fasta

From this I may have learned:

* Panaeolus papilionaceus and P. sphinctrinus are likely the same species
* Panaeolus acuminatus is close to P. olivaceus
* Panaeolus cinctulus is close to Panaeolina foenisecii and the split from Panaeolus to Panaeolina is not supported by genetic evidence as Panaeolina does not seem to form a separate clade outside of Panaeolus.
* Panaeolus semiovatus is only distantly related to the rest of Panaeolus
* Copelandia forms a distinct clade within Panaeolus, and maybe should be a subgenus or section.
* Panaeolus cambodginiensis may be the same as P. cyanescens


If anyone can provide me with more Panaeolus sequences that would be great.  There are only 21 good 28s sequences in Genbank.  And the bottom three are highly questionable.


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OfflineSt. Chibes
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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #14123834 - 03/15/11 06:38 AM (12 years, 11 months ago)

Thanks for posting that Alan!
So we should call Panaeolina foeniseciiPanaeolus foenisecii ?

I never did figure out how to use that software to make these trees on my mac. I have everything I need, I just can't figure out how to open the fasta files and run the software...


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Re: Panaeolus phylogenetic tree [Re: St. Chibes]
    #14124198 - 03/15/11 09:02 AM (12 years, 11 months ago)

yeah bravo!


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OfflineTwiztidsage
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Re: Panaeolus phylogenetic tree [Re: AIRDOG]
    #14124471 - 03/15/11 10:36 AM (12 years, 11 months ago)

:popcorn:


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OfflineNeoSporen
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Re: Panaeolus phylogenetic tree [Re: Twiztidsage]
    #14124804 - 03/15/11 01:03 PM (12 years, 11 months ago)

Great work Alan :super: It's nice to see things like this. Helps me understand how different mushrooms may be related. Your work is an inspiration to us all. :cheers:


--------------------
Having lived through an existence close to nature, one accepts the small and simple things as most important in life. Sun, wind, rain and snow. The sounds birds make, smells of fresh wild flowers. Love of all kinds, from friends and family, thy self and our neighbors. Beautiful sunrises to the darkest clouds dancing above in the sky. To forgive, learn, share and express. This is the only thing a man such as myself can ask for. What comes as the result is nothing short of the core of human existence, to truly live free in body and mind.


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Re: Panaeolus phylogenetic tree [Re: NeoSporen]
    #14124983 - 03/15/11 01:40 PM (12 years, 11 months ago)

Quote:

* Panaeolus papilionaceus and P. sphinctrinus are likely the same species


i agree.

Quote:

* Panaeolus cinctulus is close to Panaeolina foenisecii and the split from Panaeolus to Panaeolina is not supported by genetic evidence as Panaeolina does not seem to form a separate clade outside of Panaeolus.




also agree. i can help you out with this if youd like.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #14125248 - 03/15/11 02:33 PM (12 years, 11 months ago)

Interesting stuff.

Quote:

Alan Rockefeller said:
If anyone can provide me with more Panaeolus sequences that would be great.  There are only 21 good 28s sequences in Genbank.  And the bottom three are highly questionable.




Could you elaborate on what you mean?  Collecting more samples to sequence?

Edit:  And are there any sequences on Panaeolopsis yet?


Edited by Byrain (03/15/11 02:33 PM)


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #14126260 - 03/15/11 05:49 PM (12 years, 11 months ago)

Alan, which inference method did you use to construct this tree?


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OfflineKaiser
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Re: Panaeolus phylogenetic tree [Re: CureCat]
    #14126388 - 03/15/11 06:17 PM (12 years, 11 months ago)

Very interesting thanks for the link.
It doesn't surprise me that Panaeolina foenisecii is close to Panaeolus cinctulus since they look very similar with the only difference being the spore colour (which I guess is the only reason why it is a separate genus).
And I wonder were the genus Panaeolopsis fits in with all this.


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OfflineAlan RockefellerM
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Re: Panaeolus phylogenetic tree [Re: Byrain]
    #14126690 - 03/15/11 07:13 PM (12 years, 11 months ago)

Quote:

CureCat said:
Alan, which inference method did you use to construct this tree?




It is a maximum likelihood tree, with the numbers being branch lengths.


Quote:

Byrain said:
And are there any sequences on Panaeolopsis yet?




There will be one very soon.


Quote:

St. Chibes said:
I never did figure out how to use that software to make these trees on my mac.





You don't need any software, you can do it all on the web using the online version of mafft


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OfflineHashishin13
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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #14127472 - 03/15/11 09:32 PM (12 years, 11 months ago)

Why are Pan foes on there like 4 times?


--------------------
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OfflineAlan RockefellerM
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Re: Panaeolus phylogenetic tree [Re: Hashishin13]
    #14127486 - 03/15/11 09:35 PM (12 years, 11 months ago)

Four collections were sequenced.


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OfflineSt. Chibes
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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #14127522 - 03/15/11 09:42 PM (12 years, 11 months ago)

Quote:

Alan Rockefeller said:
Quote:

St. Chibes said:
I never did figure out how to use that software to make these trees on my mac.




You don't need any software, you can do it all on the web using the online version of mafft





Thanks for the link.


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Re: Panaeolus phylogenetic tree [Re: St. Chibes]
    #14127739 - 03/15/11 10:13 PM (12 years, 11 months ago)

nice worky alan. :thumbup:


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OfflineSt. Chibes
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Re: Panaeolus phylogenetic tree [Re: psylosymonreturns]
    #14127776 - 03/15/11 10:21 PM (12 years, 11 months ago)

I think I'm retarded....
I don't get it...


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OfflineAlan RockefellerM
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Re: Panaeolus phylogenetic tree [Re: St. Chibes]
    #14128053 - 03/15/11 11:07 PM (12 years, 11 months ago)

Rough instructions for making phylogenetic trees with mafft using only a web browser:


1) Get the DNA from Genbank, http://www.ncbi.nlm.nih.gov/nuccore?term=Panaeolus%20large%20subunit
At the bottom right click Send to:, chose file with FASTA format
2) Go to online mafft, start from the FASTA file on your hard drive, open all plots, make sure the lines are red.
If any graphs have blue lines remove the sequence from the file, reverse them with rev_comp, then put them back in.  http://www.bioinformatics.org/sms/rev_comp.html
3) If sequences do not allign well put the best/longest one at the top
4) Click the phylogenetic tree button

Better trees could be made by trimming off unaligned parts and manually verifying that the alignments are sane.


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OfflineAlan RockefellerM
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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #14128727 - 03/16/11 01:25 AM (12 years, 11 months ago)

Here is a tree I made for Rhodocollybia

http://plantobserver.org/rhodocollybia.pdf


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OfflineNeoSporen
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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #14128825 - 03/16/11 02:02 AM (12 years, 11 months ago)

Thanks for all the links. I'll have to sit down for a few hours and figure it all out.


--------------------
Having lived through an existence close to nature, one accepts the small and simple things as most important in life. Sun, wind, rain and snow. The sounds birds make, smells of fresh wild flowers. Love of all kinds, from friends and family, thy self and our neighbors. Beautiful sunrises to the darkest clouds dancing above in the sky. To forgive, learn, share and express. This is the only thing a man such as myself can ask for. What comes as the result is nothing short of the core of human existence, to truly live free in body and mind.


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OfflineSt. Chibes
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Re: Panaeolus phylogenetic tree [Re: NeoSporen]
    #14129633 - 03/16/11 09:02 AM (12 years, 11 months ago)

I'm going to attempt again tonight.
Thanks Alan.


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Offlinewintersbefore
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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller] * 1
    #17809946 - 02/15/13 08:40 PM (11 years, 9 days ago)

:bump:


Quote:

Alan Rockefeller said:
Rough instructions for making phylogenetic trees with mafft using only a web browser:


1) Get the DNA from Genbank, http://www.ncbi.nlm.nih.gov/nuccore?term=Panaeolus%20large%20subunit
At the bottom right click Send to:, chose file with FASTA format
2) Go to online mafft, start from the FASTA file on your hard drive, open all plots, make sure the lines are red.
If any graphs have blue lines remove the sequence from the file, reverse them with rev_comp, then put them back in.  http://www.bioinformatics.org/sms/rev_comp.html
3) If sequences do not allign well put the best/longest one at the top
4) Click the phylogenetic tree button

Better trees could be made by trimming off unaligned parts and manually verifying that the alignments are sane.





Here's what I did, sort of following those directions:
1. First I found BLAST:


2. Then I clicked on the NCBI so I could search for something:


3. Then I started typing the name of the mushroom I wanted to search for:


4. Then I told it to do a Nucleotide blast:


5. It brought up these mushrooms, and I clicked on the first one:


6. Then I told it to do a blast query on that particular sequence so that I could see all of the other sequences in GenBank that were a pretty good match for the one I typed in:



7. Then I selected from those that came up:


8. Then I saved them as a Fasta file:


9. Then I went to THIS WEBSITE and uploaded the file that I just downloaded:

and hit submit


10. Then I clicked on Phylogenetic Tree:


11. It produces this tree:



I think theres problems with this tree because I didn't "root" it? or didn't root it properly? Or I didn't really chose an outgroup, just randomly selected the sequences that came up first... I will try again later by including an outgroup somehow...


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OfflineAlan RockefellerM
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Re: Panaeolus phylogenetic tree [Re: wintersbefore]
    #17809974 - 02/15/13 08:44 PM (11 years, 9 days ago)

I think that the problem with your tree is that you chose everything that Genbank thought was similar, without vetting the sequences it gave you.  You got all of Psilocybe, and all sequences that are from Psilocybe but are misidentified as something else.

Try choosing a reasonable outgroup and retrieve the sequence for the outgroup by searching by name and choosing the most believable sequence available.

The problematic sequences are the ones that don't make any sense that are clustered with the ones that do make sense.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #17810335 - 02/15/13 09:46 PM (11 years, 9 days ago)

nice work Alan, very interesting...
however, the process seems seriously flawed (not on your end), imo.

:aliendance:
peace and love
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OfflineAlan RockefellerM
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Re: Panaeolus phylogenetic tree [Re: bloodworm]
    #17811017 - 02/15/13 11:56 PM (11 years, 9 days ago)

How is it flawed?


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Offlinewintersbefore
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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #17811703 - 02/16/13 04:35 AM (11 years, 9 days ago)

Quote:

Alan Rockefeller said:
How is it flawed?




Because my goal here was to generate a tree at finer scales than a published tree that I am using for reference in order to find P. cubensis' most closely related inactive relative.... and I was not able to do that because I would have already had to know that information in order to have a tree that makes sense.  For example, I would have to select sequences that I trust have not been misidentified to choose an "outgroup" (so I have to select something from another genus over, but how do I know what that is?!?) Is it Agrocybe praecox? If so, I've answered my question and I don't need the tree.


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OfflineAlan RockefellerM
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Re: Panaeolus phylogenetic tree [Re: wintersbefore]
    #17811712 - 02/16/13 04:40 AM (11 years, 9 days ago)

According to BLAST, Naucoria bohemica is the closest non-psychoactive mushroom to Psilocybe cubensis.

To see these results, go to this link, scroll down to the bottom of the page and click BLAST.

To get the best possible results I used a long cubensis sequence, 1997 base pairs.  But it appears that they tacked together its1, its2 and 28s.  Whether this actually gives the best results, I don't know.



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Offlinewintersbefore
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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #17811738 - 02/16/13 04:54 AM (11 years, 9 days ago)

Quote:

Alan Rockefeller said:
According to BLAST, Naucoria bohemica is the closest non-psychoactive mushroom to Psilocybe cubensis.



Would you say that Agrocybe praecox is the most closely related "common" mushroom? If not, what is (I have only seen a few pictures of Naucoria) so I guess I'd like to see, in a tree, how far away, at these finer scales, is A. praecox?

Quote:


To see these results, go to this link, scroll down to the bottom of the page and click BLAST.

To get the best possible results I used a long cubensis sequence, 1997 base pairs.  But it appears that they tacked together its1, its2 and 28s.  Whether this actually gives the best results, I don't know.







So are all those Lepistas, Tricholomas, and Hygrocybes misidentified?!? How does that even happen?? Those mushrooms don't look anything like a Psilocybe, and they are light spored!?!


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OfflineAlan RockefellerM
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Re: Panaeolus phylogenetic tree [Re: wintersbefore]
    #17811760 - 02/16/13 05:05 AM (11 years, 9 days ago)

Quote:

wintersbefore said:
Would you say that Agrocybe praecox is the most closely related "common" mushroom? If not, what is (I have only seen a few pictures of Naucoria) so I guess I'd like to see, in a tree, how far away, at these finer scales, is A. praecox?




I think that Agrocybe pediades is closer and also common, and that Panaeolus papilionaceus is closer still, and still common.


Quote:

So are all those Lepistas, Tricholomas, and Hygrocybes misidentified?!? How does that even happen?? Those mushrooms don't look anything like a Psilocybe, and they are light spored!?!




They tried to sequence a Psilocybe accidentally put the wrong dna in the well, an error caused by being distracted by people talking to them in the lab or forgetting which well is which due to smoking marijuana, or they didn't use a new pipette tip when they should have. 


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Offlinewintersbefore
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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #17811839 - 02/16/13 06:06 AM (11 years, 8 days ago)

Quote:

Alan Rockefeller said:

They tried to sequence a Psilocybe accidentally put the wrong dna in the well, an error caused by being distracted by people talking to them in the lab or forgetting which well is which due to smoking marijuana, or they didn't use a new pipette tip when they should have. 





While that does happen, it does not exactly explain what happened here.  It is possible that someone "was trying to sequence a Tricholoma" but accidentally sequenced a Psilocybe and when they got sequences back and the closest match was for Psilocybe, it should have been renamed before being submitted. Anyhow, all of those are errors in GenBank and there are far too many of them, much worse then wikipedia, because with wikipedia anyone can go in and fix the errors, but here, only the author's can right? Anyhow... unacceptable number of errors for this particular blast querry, I've not had as messy results with other things that I've querried...

Quote:

I think that Agrocybe pediades is closer and also common



Perfect! Thank you very much!


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Re: Panaeolus phylogenetic tree [Re: wintersbefore]
    #18165417 - 04/25/13 09:11 AM (10 years, 9 months ago)

Alright. Alright. I have to stir up the Hornet's Nest for a second here.

I'm equally interested in seeing an authentic phylogenetic tree of Panaeolus, but I've been experimenting with both the NCBI Nucleotide BLAST site and the MAFFT Phylogenetic Tree site, and both are currently producing highly flawed and highly incomplete results.

Let's talk Psilocybe for a second. And to make to things more accurate, let's also discuss this within the ITS1 region.

When we compare a bunch of species we don't want to randomly compare genes or regions. That doesn't make sense and is completely devoid of reason (ie compare to the purpose of DNA barcoding).

The Advanced Search at the NCBI nucleotide site is not currently functioning at it should - and that should be obvious to the NCBI Helpdesk. However, if you simply type in "Psilocybe ITS1" at http://www.ncbi.nlm.nih.gov/nuccore and hit Search, it will come up currently with 75 search results.

Issue No. 1: Almost immediately you will notice that there are old species that were once in the genus Psilocybe and have been moved into the genus Deconica, but they are still listed on NBCI as being in the genus Psilocybe. Who wants to email them the genus/species lists for these to genera? Please, someone intervene.

Issue No. 2: As most of us know, there are well over 100 species, currently, listed in the genus Psilocybe. The NCBI database is missing most of these species' sequences. If Douglas Smith or anyone else has successfully constructed an ITS1 and/or ITS2 tree that is more complete and accurate please post it here.

Issue No. 3: The MAFFT phylogenetic tree that I produced based on the 75 Psilocybes produced a tree that does not coherently display ancestry. This is the whole point of a phylogenetic tree - to make sense of the Common Ancestor theory and to show which species mutated over time to form new species.

Please - all Hornets rise! All Hornest get angry and overcome this! Email these Helpdesks! Call your senators and representatives, your teachers, your students, you colleagues, your friends......


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OfflineAlan RockefellerM
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Re: Panaeolus phylogenetic tree [Re: The Lightning]
    #18166465 - 04/25/13 01:44 PM (10 years, 9 months ago)

Quote:

Issue No. 1: Almost immediately you will notice that there are old species that were once in the genus Psilocybe and have been moved into the genus Deconica, but they are still listed on NBCI as being in the genus Psilocybe. Who wants to email them the genus/species lists for these to genera? Please, someone intervene.




They don't update their DB when new names are published.  It's up to you to cut out the sequences which are not in Psilocybe.


Quote:

Issue No. 2: As most of us know, there are well over 100 species, currently, listed in the genus Psilocybe. The NCBI database is missing most of these species' sequences.





An effort should be made to gather and sequence species that are not yet in genbank.  Cyanofibrillosa is done, baeocystis should be done soon.  What else do we have access to that isn't in there?


Quote:

Issue No. 3: The MAFFT phylogenetic tree that I produced based on the 75 Psilocybes produced a tree that does not coherently display ancestry. This is the whole point of a phylogenetic tree - to make sense of the Common Ancestor theory and to show which species mutated over time to form new species.





How well are your sequences aligning?    You might need to toss out some sequences that don't have significant overlap.  And definitely toss out the Deconica sequences.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #18166509 - 04/25/13 01:56 PM (10 years, 9 months ago)

Quote:

Alan Rockefeller said:
Quote:

Issue No. 1: Almost immediately you will notice that there are old species that were once in the genus Psilocybe and have been moved into the genus Deconica, but they are still listed on NBCI as being in the genus Psilocybe. Who wants to email them the genus/species lists for these to genera? Please, someone intervene.




They don't update their DB when new names are published.  It's up to you to cut out the sequences which are not in Psilocybe.


Quote:

Issue No. 2: As most of us know, there are well over 100 species, currently, listed in the genus Psilocybe. The NCBI database is missing most of these species' sequences.





An effort should be made to gather and sequence species that are not yet in genbank.  Cyanofibrillosa is done, baeocystis should be done soon.  What else do we have access to that isn't in there?


Quote:

Issue No. 3: The MAFFT phylogenetic tree that I produced based on the 75 Psilocybes produced a tree that does not coherently display ancestry. This is the whole point of a phylogenetic tree - to make sense of the Common Ancestor theory and to show which species mutated over time to form new species.





How well are your sequences aligning?    You might need to toss out some sequences that don't have significant overlap.  And definitely toss out the Deconica sequences.



Maybe someone (me) can make a list of the species that still need sequenced and make a thread to let other know what is needed...

Why don't they test all collections the same way? Ie.

-Psilocybe cyanescens voucher PRM 902040 18S small subunit ribosomal RNA gene, partial sequence; internal transcribed spacer 1, 5.8S ribosomal RNA gene, and internal transcribed spacer 2, complete sequence; and 28S large subunit ribosomal RNA gene, partial sequence

-Psilocybe cyanescens voucher PRM 901481 translation elongation factor 1-alpha (EF1-alpha) gene, partial cds

I am correct in thinking these are tests in difference area of the genetic sequence?


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OfflineAlan RockefellerM
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Re: Panaeolus phylogenetic tree [Re: maynardjameskeenan]
    #18166563 - 04/25/13 02:09 PM (10 years, 9 months ago)

Quote:

maynardjameskeenan said:
Why don't they test all collections the same way? Ie.

-Psilocybe cyanescens voucher PRM 902040 18S small subunit ribosomal RNA gene, partial sequence; internal transcribed spacer 1, 5.8S ribosomal RNA gene, and internal transcribed spacer 2, complete sequence; and 28S large subunit ribosomal RNA gene, partial sequence

-Psilocybe cyanescens voucher PRM 901481 translation elongation factor 1-alpha (EF1-alpha) gene, partial cds

I am correct in thinking these are tests in difference area of the genetic sequence?





Correct, those are two different loci and they can not be directly compared because there is no overlap.  ITS is often the best gene region for finding relationships between species.  LSU and ef-1 are better for determining the relationships between genera.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #18166713 - 04/25/13 02:37 PM (10 years, 9 months ago)

Quote:

What else do we have access to that isn't in there?




I'll repeat this to others because I'm sure there's important data out there: Who knows of some research that did not get into Genbank (NCBI)?

Quote:

How well are your sequences aligning?




I hate it when you make me ask you questions. I feel stupid when I ask questions. Can you tell us the difference between a sequence, a BLAST, and a sequence alignment? If you have an extra moment or two, can you also show us the difference between a poorly aligned sequence and a well-aligned sequence?

Quote:

Maybe someone (me) can make a list of the species that still need sequenced and make a thread to let other know what is needed.




Please do. I would benefit from it and it'll keep things rolling.


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Re: Panaeolus phylogenetic tree [Re: The Lightning]
    #18166939 - 04/25/13 03:03 PM (10 years, 9 months ago)

Quote:

The Lightning said:


Quote:

How well are your sequences aligning?




I hate it when you make me ask you questions. I feel stupid when I ask questions. Can you tell us the difference between a sequence, a BLAST, and a sequence alignment? If you have an extra moment or two, can you also show us the difference between a poorly aligned sequence and a well-aligned sequence?






I think this is what he means.


aligned meaning that the same area of the sequence are compared to one another.
If the same regions of the sequence aren't compared it gives erroneous results.
They need to have overlapping sequences to be meaningful, it determines common ancestry I think if they share common areas of their sequences IE. same genes in the same places. 
It's like asking what's the make and model of your car and I say blue, unless certain aspects of the sequence are compared it doesn't mean much at all. I'm not certain to be honest, I am still kinda new to this.

Why the hell don't they test all samples the same way?

I know what you mean about Alan making you feel dumb, he does it to me too... :dunce:

Quote:

What else do we have access to that isn't in there?




Quote:

The Lightning said:
I'll repeat this to others because I'm sure there's important data out there: Who knows of some research that did not get into Genbank (NCBI)?




This is easy to find out, just search the Genbank for any given species and if it doesn't return any results that means it doesn't have an entry, Psilocybe strictipes is a good example.



Edited by maynardjameskeenan (04/25/13 03:13 PM)


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Re: Panaeolus phylogenetic tree [Re: maynardjameskeenan]
    #18167102 - 04/25/13 03:29 PM (10 years, 9 months ago)

I think sometimes Alan is intentionally obtuse so I will find things out for myself,
It's probably more meaningful to find things out on your own then to just get the answers from somebody. 

I wish he would explain things to me like I'm 5 sometimes.


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Re: Panaeolus phylogenetic tree [Re: maynardjameskeenan]
    #18167171 - 04/25/13 03:42 PM (10 years, 9 months ago)

Quote:

Why the hell don't they test all samples the same way?




DNA sequencing is an attempt at formal species identification. Different life forms (birds vs fungi for instance) can have different regions or genes that are more accurate, more legible, or more determinative - so for birds you might want to use CO1 and for fungi you might want to use ITS1.

There have been proposals and articles written as recently as the past two years about which area is most relevant for fungi.

http://www.pnas.org/content/early/2012/03/29/1117018109.full.pdf

http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025081

http://www.springerprotocols.com/Pdf/doi/10.1007/978-1-61779-591-6_9?encCode=TkVHOjlfNi0xOTUtOTc3MTYtMS04Nzk=&tokenString=XXcLRlQOVJlxS+34hSUhsw==&access=denied

http://www.benthamscience.com/open/toainfoj/articles/V005/SI0001TOAINFOJ/45TOAINFOJ.pdf


Quote:

It's probably more meaningful to find things out on your own




Not always. Alan and others have really been helpful to me. I'm thankful for their answers.


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Re: Panaeolus phylogenetic tree [Re: The Lightning]
    #18167321 - 04/25/13 04:07 PM (10 years, 9 months ago)

Quote:

The Lightning said:
Quote:

Why the hell don't they test all samples the same way?




DNA sequencing is an attempt at formal species identification. Different life forms (birds vs fungi for instance) can have different regions or genes that are more accurate, more legible, or more determinative - so for birds you might want to use CO1 and for fungi you might want to use ITS1.

There have been proposals and articles written as recently as the past two years about which area is most relevant for fungi.

http://www.pnas.org/content/early/2012/03/29/1117018109.full.pdf

http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025081

http://www.springerprotocols.com/Pdf/doi/10.1007/978-1-61779-591-6_9?encCode=TkVHOjlfNi0xOTUtOTc3MTYtMS04Nzk=&tokenString=XXcLRlQOVJlxS+34hSUhsw==&access=denied

http://www.benthamscience.com/open/toainfoj/articles/V005/SI0001TOAINFOJ/45TOAINFOJ.pdf




Makes sense. Thank you.

Quote:

It's probably more meaningful to find things out on your own




Quote:

The Lightning said:
Not always. Alan and others have really been helpful to me. I'm thankful for their answers.




You make it sound as if I'm ungrateful...
I agree with you. I did say sometimes. He's been a great help to me over the past few years but at times he can seem intentionally vague, IMO.


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Edited by maynardjameskeenan (04/25/13 04:10 PM)


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Re: Panaeolus phylogenetic tree [Re: maynardjameskeenan]
    #18167548 - 04/25/13 04:54 PM (10 years, 9 months ago)

Quote:

You make it sound as if I'm ungrateful...




No, my friend. I didn't mean it that way at all. Sometimes my typed-tone doesn't translate the actual way I intended it to sound.


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Re: Panaeolus phylogenetic tree [Re: The Lightning]
    #18198462 - 05/01/13 06:15 PM (10 years, 9 months ago)

Quote:

Can you tell us the difference between a sequence, a BLAST, and a sequence alignment?




Sequence

A nucleic acid sequence is a succession of letters that indicate the order of nucleotides within a DNA (using GACT) or RNA (GACU) molecule.


Example of a sequence of Psilocybe ovoideocystidiata:

1 gatcattatt gaataacttt ggcgtggttg tagctggccc tctcgggggc atgtgctcgc
      61 cygtcatctt tatatctcca cctgtgcacc ttttgtagac gtctttgttg gaagctgrat
      121 aggagagaat gggtgctagt cactctttct cgagttgaag gctttctcaa ggtcgctcta
      181 tgttttcata taccccaagt atgtaacaga atgtatctat atggccttgt gcctataaaa
      241 ctatatacaa ctttcagcaa cggatctctt ggctctcgca tcgatgaaga acgcagcgaa
      301 atgcgataag taatgtgaat tgcagaattc agtgaatcat cgaatctttg aacgcacctt
      361 gcgctccttg gtattccgag gagcatgcct gtttgagtgt cattaaattc tcaaccttac
      421 cagcttttgt tagcttgtgt aatggcttgg acttgggggt tttttgccgg cttctaacaa
      481 agtcagctcc ccttaaatgc attagccggc tgcccgctgt ggaccgtcta ttggtgtgat
      541 aattatctac gccgtggatg tctgctatca atgggttttt aaagctgctt ctaaccgtct
      601 gttcattcgg acaatacaat gacaatttga cctcaaatca ggtaggacta cccgctgaac
      661 ttaagcatat ca


BLAST (Basic Local Alignment Search Tool)

BLAST finds regions of similarity between biological sequences. The program compares nucleotide or protein sequences to sequence databases and calculates the statistical significance of matches. BLAST can be used to infer functional and evolutionary relationships between sequences as well as help identify members of gene families.


Sequence Alignment

A sequence alignment is a way of arranging the sequences of DNA, RNA, or protein to identify regions of similarity that may be a consequence of functional, structural, or evolutionary relationships between the sequences. Aligned sequences of nucleotides are typically displayed in rows.



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Re: Panaeolus phylogenetic tree [Re: The Lightning]
    #18198536 - 05/01/13 06:32 PM (10 years, 9 months ago)

Quote:

The Lightning said:
Quote:

Can you tell us the difference between a sequence, a BLAST, and a sequence alignment?




Sequence

A nucleic acid sequence is a succession of letters that indicate the order of nucleotides within a DNA (using GACT) or RNA (GACU) molecule.


Example of a sequence of Psilocybe ovoideocystidiata:

1 gatcattatt gaataacttt ggcgtggttg tagctggccc tctcgggggc atgtgctcgc
      61 cygtcatctt tatatctcca cctgtgcacc ttttgtagac gtctttgttg gaagctgrat
      121 aggagagaat gggtgctagt cactctttct cgagttgaag gctttctcaa ggtcgctcta
      181 tgttttcata taccccaagt atgtaacaga atgtatctat atggccttgt gcctataaaa
      241 ctatatacaa ctttcagcaa cggatctctt ggctctcgca tcgatgaaga acgcagcgaa
      301 atgcgataag taatgtgaat tgcagaattc agtgaatcat cgaatctttg aacgcacctt
      361 gcgctccttg gtattccgag gagcatgcct gtttgagtgt cattaaattc tcaaccttac
      421 cagcttttgt tagcttgtgt aatggcttgg acttgggggt tttttgccgg cttctaacaa
      481 agtcagctcc ccttaaatgc attagccggc tgcccgctgt ggaccgtcta ttggtgtgat
      541 aattatctac gccgtggatg tctgctatca atgggttttt aaagctgctt ctaaccgtct
      601 gttcattcgg acaatacaat gacaatttga cctcaaatca ggtaggacta cccgctgaac
      661 ttaagcatat ca


BLAST (Basic Local Alignment Search Tool)

BLAST finds regions of similarity between biological sequences. The program compares nucleotide or protein sequences to sequence databases and calculates the statistical significance of matches. BLAST can be used to infer functional and evolutionary relationships between sequences as well as help identify members of gene families.


Sequence Alignment

A sequence alignment is a way of arranging the sequences of DNA, RNA, or protein to identify regions of similarity that may be a consequence of functional, structural, or evolutionary relationships between the sequences. Aligned sequences of nucleotides are typically displayed in rows.





:bow2: You are great at explaining things! Awesome, i wanna guide for DNA of mushrooms and using BLAST, ect. You should make me one for the guide :super:


--------------------
~~~~~~***Psilocybin Mushrooms***~~~~~~
_________A Practical Guide To Psilocybin Mushrooms_________

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Re: Panaeolus phylogenetic tree [Re: Joust]
    #18198628 - 05/01/13 06:50 PM (10 years, 9 months ago)

Quote:

I wanna guide for DNA of mushrooms and using BLAST




Here's the basic instructions, but more people will need to use the site and consider how it can be better designed, not that I'm one to complain:

1. Figure out what Psilocybe (or mushroom species) you want to study. Let's pick Psilocybe fasciata for this experiment.



2. What region or gene(s) do you want to study for this experiment? The proposed universal fungi DNA barcode is ITS. Let's pick ITS1 since the NCBI Advanced Search isn't working using ITS.

3. Let's go to this site: https://www.ncbi.nlm.nih.gov/

4. Type in "Psilocybe fasciata ITS1" and search. On the next screen hit Nucleotide.

5. Two sequences (as of today) appear. Let's take a moment to reflect on some of the info displayed before going further. What is an accession number? Also notice that one sequence is going to show us 1592 base pairs but the second one has 647 base pairs. What is a base pair?


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Re: Panaeolus phylogenetic tree [Re: The Lightning]
    #18200280 - 05/01/13 11:50 PM (10 years, 9 months ago)

Quote:

What is an accession number?


 

The first Psilocybe fasciata listing on NCBI has Accession: AB158635.1.

This is simply a number assigned to the sequence for reference and organization.  https://www.ncbi.nlm.nih.gov/Sequin/acc.html

Quote:

What is a base pair?




This is all about nucleotides.



Nucleotides for DNA = adenine (A), cytosine (C), guanine (G), & thymine (T)

A DNA "ladder" is made of these nucleotides. They are connected and form a spiral railroad track. Each "wood track" of the railroad so-to-speak is made of a base pair (two nucleotides connected to each other):  G-C guanine-cytosine or A-T adenine-thymine.

When we look at DNA sequences, we are looking at base pairs - or more accurately 1/2 of the base pairs (ie one half of the rail road - or one strand of the two-strand DNA).



Example of a nucleotide sequence of Psilocybe fasciata:

        1 aaggatcatt attgaatgaa cttggctcgg ttgcagctgg tcctctcgag ggcatgtgct
      61 cgccgtgtca tctttatctc tccacctgtg caccttttgt agacctggat tagttaactt
      121 tccgaggaaa ctcggtcggg aggattgctt tcacaagctc tccttgcaat taacccaggc
      181 ctacgttttc atatacccca aagaatgtaa cagaatgtat tatattggcc ttgtgcctat
      241 aaactatata caactttcag caacggatct cttggctctc gcatcgatga agaacgcagc
      301 gaaatgcgat aagtaatgtg aattgcagaa ttcagtgaat catcgaatct ttgaacgcac
      361 cttgcgctcc ttggtattcc gaggagcatg cctgtttgag tgtcattaaa ttctcaacct
      421 taccagcttt tgctgataat ggcttggatg tgggggtctt ttgctggctt cgtcaagagg
      481 tctgctcccc ttaaatgtat tagccggtgc cccgcgcaga gccgtctatt ggtgtgataa
      541 ttatctacgc cgtggacctt tgcatgaatg ggattgcgct gcttctaacc gtccttcact
      601 ggacaataca aatgacaatt tgacctcaaa tcaggtagga ctacccgctg aacttaagca
      661 tatcaataag cggaggaaaa gaaactaaca aggattcccc tagtaactgc gagtgaagcg
      721 ggaaaagctc aaatttaaaa tctggcggtc tttggccgtc cgagttgtaa tctagagaag
      781 tgttatccgc gctggaccgt gtacaagtct cctggaatgg agcgtcatag agggtgagaa
      841 tcccgtcttt gacacggact gccagggctt tgtgatgcgc tctcaaagag tcgagttgtt
      901 tgggaatgca gctcaaaatg ggtggtaaat tccatctaaa gctaaatatt ggcgagagac
      961 cgatagcgaa caagtaccgt gagggaaaga tgaaaagaac tttggaaaga gagttaaaca
    1021 gtacgtgaaa ttgctgaaag ggaaacgctt gaagtcagtc gcgttgtccg gggatcaacc
    1081 ttgcttttgc tgggtgtact ttccggttga cgggtcagca tcaattttga ccgttggata
    1141 aagtctgtgg aaatgtggct tcttcggaag tgttatagtc catggtcgca tacaacggtt
    1201 gggattgagg aactcagcac gccgaaaggc cgggtacttg taccacgttc gtgcttagga
    1261 tgctggcata atggctttaa tcgacccgtc ttgaaacacg gaccaaggag tctaacatgc
    1321 ctgcgagtgt ttgggtggaa aacccgagcg cgtaatgaaa gtgaaagttg agatccctgt
    1381 cgcggggagc atcgacgccc ggaccagacc ttttgtgacg gatccgcggt agagcatgta
    1441 tgttgggacc cgaaagatgg tgaactatgc ctgaataggg tgaagccaga ggaaactctg
    1501 gtggaggctc gtagcgattc tgacgtgcaa atcgatcgtc aaatttgggt ataggggcga
    1561 aagactaatc gaaccatcta gtagctggtt cc

So let's get an idea of what the above stuff is. We know our letters represent the four DNA nucleotides (GCAT) since this is a DNA nucleotide sequence we searched for.

What are the numbers on the left of each line? Count the letters in the first small group of the sequence. It's the same number of letters (10) for each group of letters until the very last group where the sequence ends. Each number listed on the left represents the starting number of the nucleotide as if it were a numbered list. (It is a numbered list!)

Remember ~ there were 1592 base pairs listed with this particular sequence as noted earlier. We're seeing one "side" (one strand!) and one half of the pairs throughout the sequence typed out above. Because we know one half of the two DNA "railroad sides" we also know the other half without mentioning it in the above sequence (A always connectes to T, and C always connects to G)*.

*There are actually exceptions to this but for the sake of getting on track (pun intended) let's keep it simple.


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Re: Panaeolus phylogenetic tree [Re: The Lightning]
    #18201206 - 05/02/13 08:36 AM (10 years, 9 months ago)

So, let's continue...

We searched for Psilocybe fasciata and arrived here: https://www.ncbi.nlm.nih.gov/nuccore/?term=Psilocybe%20fasciata%20ITS1 - and now we'll click the first listing which takes us here: https://www.ncbi.nlm.nih.gov/nuccore/AB158635.1

To perform a BLAST (Basic Local Alignment Search Tool) of just this DNA sequence we'll click "Run BLAST" on the right side.

BLAST finds regions of similarity between biological sequences. The program compares nucleotide sequences (or protein sequences) to sequence databases and calculates the statistical significance of matches.

After clicking "Run BLAST" a new page will appear with a title "Standard Nucleotide BLAST". Notice the Accession Number was added into the "Enter Query Sequence" box. Scroll down and click the BLAST button.



Edited by The Lightning (05/02/13 08:41 AM)


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Re: Panaeolus phylogenetic tree [Re: The Lightning]
    #18201384 - 05/02/13 09:30 AM (10 years, 9 months ago)

A new screen will appear. The first part looks like this:



The second part like this:



You can mouse-over each red line (each red line represents an individual sequence the BLAST was compared to). Reminder: Mouse-over and then click to show alignments, if you'd like to review an individual "alignment."

https://en.wikipedia.org/wiki/Sequence_alignment

Reminder:

Sequence Alignment

A sequence alignment is a way of arranging the sequences of DNA, RNA, or protein to identify regions of similarity that may be a consequence of functional, structural, or evolutionary relationships between the sequences. Aligned sequences of nucleotides are typically displayed in rows.



Let's test this idea out. (Click on the first red line). It will scroll down really fast on the same page and go to the actual sequence. The first match turned out to be the exact same Accession Number (ie the same sequence) as we BLASTed. In other words, we're now looking at the Psilocybe fasciata sequence compared to itself. This is just to get us familiar with the site so don't get frustrated.



The Query (the originally BLASTed accession number) is compared to another accession number which is listed as Sbjct.

(Again, in this case everything is identical because we're comparing a sequence to itself).


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Re: Panaeolus phylogenetic tree [Re: The Lightning] * 1
    #18201837 - 05/02/13 11:31 AM (10 years, 9 months ago)

Okay. So let's go back up near the top of that screen, below the graph where this info appears:




Notice the following categories (fields):

Max score
Total score
Query cover
E value
Max ident
Accession
- We already know what the Accession is. It's just the number assigned to this sequence.


:mushdance:


Here's my understanding of these fields and what they mean, although I think re-wording will be needed as I grasp them further:

Max score: This number is based on how well the items match. The longer the sequence and the better the match, the higher the score. I would pay far more attention to the Query Coverage field and the Maximum Identity field.

Total score: Sum of scores of all aligned sequence. Again, pay more attention to the Query Coverage and the Maximum Identify.

Query coverage: Displays the percentage of the original sequence aligned to the listed item. You want a high percentage so there is a fair comparison going on.

E Value: The E value (Expect Value) represents the number of search hits you would expect to find on NCBI - by chance - based on the quality of the alignment and the size of the actual database (ie the entire NCBI database and the databases NCBI draws data from). It decreases exponentially as the Score (S) of the match increases. For example, an E value of 1 assigned to a hit can be interpreted as meaning that in a database of the current size one might expect to see 1 match with a similar score simply by chance.

Max ident (Maximum Identity): The maximum percentage of identical identity between the accession number and another sequence.


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Re: Panaeolus phylogenetic tree [Re: The Lightning]
    #18203304 - 05/02/13 04:17 PM (10 years, 9 months ago)

:justastonishing:
You do good work, I'm am glad to see others taking interest in this subject.
This is where the the future of mycology is headed, data based analysis beats guess-work any day. 

Have you made any phylogenetic tree's yet?
Would you be interested in collaborating on a Psilocybes of the PNW tree, if you have any extra time?
I love to see how species relate to one another.


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Re: Panaeolus phylogenetic tree [Re: maynardjameskeenan]
    #18203381 - 05/02/13 04:27 PM (10 years, 9 months ago)

Quote:

Have you made any phylogenetic tree's yet?




This tree was constructed using phylogeny.fr (http://www.phylogeny.fr/version2_cgi/index.cgi):



This is based on hand-picked entries of Psilocybe species after an NCBI search for "Psilocybe ITS1." I had to leave out species which were still labelled as Psilocybe species and were actually Deconica species. It doesn't tell us much and lacks any usefulness for the time being.

Quote:

Would you be interested in collaborating on a Psilocybes of the PNW tree, if you have any extra time?




Yes. Maybe you can do the next tree and we can explore the DNA Species Concept together.


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Re: Panaeolus phylogenetic tree [Re: The Lightning]
    #18203464 - 05/02/13 04:40 PM (10 years, 9 months ago)

Did you use MAFFT for aligning the data collected from BLAST?

Here is a rough tree for the Psilocybes that occur in Oregon. There are still a few species missing from the genbank.


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Re: Panaeolus phylogenetic tree [Re: maynardjameskeenan]
    #18203537 - 05/02/13 04:52 PM (10 years, 9 months ago)

I'm not sure what MAFFT gives us that the NCBI site isn't already giving us.


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Re: Panaeolus phylogenetic tree [Re: The Lightning]
    #18203595 - 05/02/13 05:00 PM (10 years, 9 months ago)

Aesthetics, mostly. It's also helpful for comparing collection that are not genitally similar.



This tree could not be possible using the NCBI website.


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Re: Panaeolus phylogenetic tree [Re: The Lightning]
    #18203601 - 05/02/13 05:01 PM (10 years, 9 months ago)

Quote:

The Lightning said:
I'm not sure what MAFFT gives us that the NCBI site isn't already giving us.





It has some nice tools for getting rid of the sequences that don't align well.


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Re: Panaeolus phylogenetic tree [Re: maynardjameskeenan]
    #18292207 - 05/20/13 11:04 AM (10 years, 9 months ago)

Quote:

maynardjameskeenan said:
Aesthetics, mostly. It's also helpful for comparing collection that are not genitally similar.





Were you able to properly compare different genes or regions between species?

For example, taking one sequence of Psilocybe semilanceata that has a standard ITS region sequence (ITS1, 5.8S, and ITS2) complete, and comparing it to a sequence of Psilocybe fasciata that only has a CO1 sequence?

This does not seem logical but maybe I'm missing something here.


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Re: Panaeolus phylogenetic tree [Re: The Lightning]
    #18293722 - 05/20/13 04:56 PM (10 years, 9 months ago)

Quote:

The Lightning said:
Quote:

maynardjameskeenan said:
Aesthetics, mostly. It's also helpful for comparing collection that are not genitally similar.





Were you able to properly compare different genes or regions between species?

For example, taking one sequence of Psilocybe semilanceata that has a standard ITS region sequence (ITS1, 5.8S, and ITS2) complete, and comparing it to a sequence of Psilocybe fasciata that only has a CO1 sequence?

This does not seem logical but maybe I'm missing something here.




Hmmmm....
I not sure how to answer this.

I think that under the summery of any mushroom you look up it lists multiply gene regions sequenced.
ie;

Psilocybe fasciata
[18S ribosomal RNA gene, partial sequence;]
[internal transcribed spacer 1, 5.8S ribosomal RNA gene,[ and internal transcribed spacer 2, complete sequence;]]
[28S ribosomal RNA gene, partial sequence]

Psilocybe azurescens
[voucher PRM 901174 18S small subunit ribosomal RNA gene, partial sequence;]
[internal transcribed spacer 1, 5.8S ribosomal RNA gene,[ and internal transcribed spacer 2, complete sequence; and 28S large]] [subunit ribosomal RNA gene, partial sequence]

I think that maybe the brackets separate gene regions and just as long as the two samples you are comparing test the same regions you should get meaningful data.

I'm still kinda new to this stuff, so please be patient with me and forgive my mistakes.


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Re: Panaeolus phylogenetic tree [Re: maynardjameskeenan]
    #18293815 - 05/20/13 05:22 PM (10 years, 9 months ago)

Quote:

I think that maybe the brackets separate gene regions and just as long as the two samples you are comparing test the same regions you should get meaningful data.




This is what I'm wondering, too.

I get the feeling (when reading scientific papers) that mycologists are accidentally "aligning" regions that are dissimilar.

The ITS region (which is made up of ITS1, 5.8S, and ITS2 - as a complete sequence) is the DNA barcode for fungi including Psilocybe - at least for now.

To compare an ITS region barcode to a CO1 gene or the Small SubUnit region seems like comparing my arm to your foot. But again, there may be a way to align these regions that hasn't been explained to me yet.

Knock, knock, knockin' on Alan's door....Oh yeah....


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Re: Panaeolus phylogenetic tree [Re: The Lightning]
    #18294181 - 05/20/13 06:37 PM (10 years, 9 months ago)

You need to compare the same region to get any meaningful results.

If you are comparing different regions they won't align at all.

You can view the alignments with a tool like mafft, and drop any sequences that don't align well.  Then make the tree.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #20060495 - 05/30/14 10:27 AM (9 years, 8 months ago)



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Re: Panaeolus phylogenetic tree [Re: nomendubium]
    #20061981 - 05/30/14 05:10 PM (9 years, 8 months ago)

Why do you think that one is Panaeolina foenisecii?

I got the first results of sequencing back from the lab work I have been doing recently, here is a sequence for Panaeolus papilionaceus from Pico de Orizaba, the tallest volcano in North America.    I found this at over 4000 meters elevation, well above the tree line. 


http://mushroomobserver.org/114447

ITS sequence: AATCGTCATTATCGAATAAACTGGGTGGGTTGTTGCTGTCCCTCTCGGGGGAACTGTGCACGCCTTACCTTTTTTGTTTTTCCACCTGTGAACACACTGTAGGTCTGGAGGGAAGGGAAGCAATTCCTTGACGTTCAGGTCCTATGCTTATACATATACACTATTGAAAGTAACAGAACGATTCAATGGGCTCTAAGCCTATAAACTATATACAACTTTCAGCAACGGATCTCTTGGCTCTCGCATCGATGAAGAACGCAGCGAAATGCGATAAGTAATGTGAATTGCAGAATTCAGTGAATCATCGAATCTTTGAACGCACCTTGCGCTCTTTGGTATTCCGAAGAGCATGCCTGTTTGAGTGTCATTAAATTCTCAACCTCATCGCTTTTGTGATGATGGCTTGGATGTGGAGGTTTCTGCAGGCCGAAAGGTCTGCTCCTCTCAAATGAATTAGTGGGTGCCCCGCGCAAACCTATCTATTGGTGTGATAATTATCTACGCCGTGGTCTAGGATTGCTGTAAAAAGGTGTTTGCCCTGCTTCTAACCGTCCATTGACTTGGACAACTTGAACCATTTGACCTCAAATCAGGTAGGACTACCCGCTGAACTTAAGCATATCAATAAGCGGAGGAAAAGAAACTAACAAGGATTCCCCTAGTAACTGCGAGTGAAGCGGGAAAAGCTCAAATTTAAAATCTGGCAGTCTTCGATTGTCCGAGTTGTAATCTAGAGAAGCATTATCCGCGCTGGACCGTGTACAAGTCTCCTGA

It's a good match for Panaeolus papilionaceus from Italy and Germany.  Makes sense that it is an imported species since it grows on cow dung and they didn't have cows in North America until they were brought over here.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #20062006 - 05/30/14 05:22 PM (9 years, 8 months ago)

because
Nomendubium said
Quote:

The strangely positioned "Paneaolus subbalteatus" AB104674.2
    If you click on the pubmed entry 12808251 This is a DNA sequence of a Powdered mushroom! from 2003 I might add. At the time of publication they probably didn't have a lot of sequences to compare to and elected to call it Panaeolus subbalteatus because that species was the closest thing yet sequenced at the time. Now 12 years later it is clear that the correct identity for the entry would have been/is Panaeolina foenisecii



and it happens to be on your tree with 3 foes


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Re: Panaeolus phylogenetic tree [Re: nomendubium]
    #20062034 - 05/30/14 05:28 PM (9 years, 8 months ago)

here is the paper, it is j2, collected in 1983, sequenced in 2003 https://www.jstage.jst.go.jp/article/cpb/51/6/51_6_710/_pdf


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Re: Panaeolus phylogenetic tree [Re: nomendubium]
    #20062634 - 05/30/14 08:52 PM (9 years, 8 months ago)

I believe that it is a species of Panaeolus which is not in genbank yet.  The closest Panaeolina foenisecii sequence is 9 base pairs away.  BLAST seems to think that it is closer to Panaeolus papilionaceus.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #20062655 - 05/30/14 08:57 PM (9 years, 8 months ago)

Great job Alan, doing some real research now!


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Re: Panaeolus phylogenetic tree [Re: Joust]
    #20062866 - 05/30/14 09:59 PM (9 years, 8 months ago)

ok. I'll agree with that. The other 2 specimens I found labelled Panaeolus subbalteatus don't even make the list. I'm not trying to critisize you, I'm improving the tree. I think it being labelled subbalteatus when it is clearly not causes a misleading conclusion about the position of Panaeolina on the tree. Taking that into consideration along with this post and the next one which show that there is 100% match between the Pan foe on the branch with the semiovatus and that Panaeolus foenisecii in the other branch of the tree (with the other foes) then Paneaolina would make it's own little branch. That would
A) Make a lot of sense
B) Allow us to move forward in the science of understanding the relationship of the species


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Re: Panaeolus phylogenetic tree [Re: nomendubium]
    #20129724 - 06/14/14 03:33 PM (9 years, 8 months ago)
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I just made a new Panaeolus tree, this time using the ITS region.  This is a maximum liklihood tree using the Tamura 3 parameter model w/ 500 rounds of bootstrapping, gamma distributed rates among sites and the gap treatment set to partial deletion. 

Sequences were gathered from genbank based on the name of the sequence, and one sequence that I generated from Mexican material is included.  This pretty much follows the recommendations in http://mbe.oxfordjournals.org/content/early/2013/03/13/molbev.mst012.full, except I didn't use BLAST to get the sequences.

I have a tree calculating right now for the rpb1 region, however there aren't many sequences in genbank for that locus, and I don't have the primers to generate my own yet.

The ITS tree is attached to this post as a PDF.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #20129967 - 06/14/14 04:33 PM (9 years, 8 months ago)
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Here is another tree I just made, this one is with the rpb1 gene.  There weren't too many RPB1 sequences in Genbank, however what there is does seem to work pretty well to make a tree that comes out the way I think things evolved.  Since there weren't too many Panaeolus species, I included some Psilocybes as well.  The Psilocybe sequences are Vicky's, so I have high confidence that they are properly identified.

I used Pholiota flammans as an outgroup and made a maximum likelihood tree using the Kimura two parameter model, gamma distributed rates among sites, and gaps treated with the partial deletion algorithm.  I used the free Mega6 software to do the analysis, following the protocol outlined in Hall 2013.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #20149493 - 06/18/14 12:05 PM (9 years, 8 months ago)

3 days and 20 hours. 2 downloads. Either no one is the least bit interested in your tree or there is some other reason no one wants to look at it. Why don't you go ahead and just post the tree? If you were a noob, I could understand making a pdf, but you are not, so just post the tree and I'll go ahead and peer review it for you  :thisisheavyman:


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Re: Panaeolus phylogenetic tree [Re: nomendubium]
    #20149576 - 06/18/14 12:37 PM (9 years, 8 months ago)



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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #20149617 - 06/18/14 12:51 PM (9 years, 8 months ago)

:smile: I meant the Panaeolus tree


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Re: Panaeolus phylogenetic tree [Re: nomendubium]
    #20149672 - 06/18/14 01:08 PM (9 years, 8 months ago)

I must have been one of the two people who downloaded it...
I think he might have posted the links and another thread as well.
I hope you don't mind me posting this Alan


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Re: Panaeolus phylogenetic tree [Re: maynardjameskeenan]
    #20153344 - 06/19/14 03:52 AM (9 years, 8 months ago)

Interesting that campanulatus, sphinctrinus, papilionaceus, retirugis are not more closer, since they are considered to be the same species...


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Re: Panaeolus phylogenetic tree [Re: knarkkorven]
    #20154215 - 06/19/14 09:31 AM (9 years, 8 months ago)

That is why you can't completely count on the names associated with DNA sequences unless you can also examine the associated collection.


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Re: Panaeolus phylogenetic tree [Re: knarkkorven]
    #20156003 - 06/19/14 04:21 PM (9 years, 8 months ago)

Quote:

knarkkorven said:
Interesting that campanulatus, sphinctrinus, papilionaceus, retirugis are not more closer, since they are considered to be the same species...





I think they are close enough to be the same species, if you discount the couple obviously mislabeled ones.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #20156381 - 06/19/14 05:57 PM (9 years, 8 months ago)

Yeah, I guess there's a lot of mislabeled mushrooms in the database.
It would be interesting to see a tree with only correct ID's.


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Re: Panaeolus phylogenetic tree [Re: knarkkorven]
    #20157728 - 06/20/14 12:25 AM (9 years, 8 months ago)

Quote:

knarkkorven said:
Yeah, I guess there's a lot of mislabeled mushrooms in the database.
It would be interesting to see a tree with only correct ID's.





The most important part of making a phylogenetic tree is selecting which other species to include.  This is usually done by looking at who made the sequence, and deciding if they really knew what they were talking about when they ID'd it.

You could also get the samples and ID them microscopically, but that takes a lot of time.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #20680650 - 10/09/14 04:28 PM (9 years, 4 months ago)

Here is a new Panaeolus phylogenetic tree that I made today using ITS sequences from genbank and to answer the question "Should Panaeolina foenisecii be included in Panaeolus sensu stricto?" It appears as though Maire errored when he moved Panaeolus foenisecii into Panaeolina in 1933, and we should be using the older name Panaeolus foenisecii.




The tree was built with the website phylogeny.fr, which allows you to do phylogenetic analysis from your web browser without installing any software.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #20680686 - 10/09/14 04:36 PM (9 years, 4 months ago)

Why is Panaeolus rickenii in two separate locations? Misidentification?


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #20680764 - 10/09/14 04:53 PM (9 years, 4 months ago)

It would be nice if collections of Panaeolus papilionaceus / spinchtrinus / campanulatus / etc. could be analyzed when found at the same time and place, I think confusion and misidentifications could have caused a lot of mixed samples in the collections, perhaps even leading to the false misconception of these being all the same species.

I mean, DNA-samples based on mushrooms with visual differences, like this (ignore the cinctulus at the left):


Edited by knarkkorven (10/09/14 05:08 PM)


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Re: Panaeolus phylogenetic tree [Re: knarkkorven]
    #20680774 - 10/09/14 04:56 PM (9 years, 4 months ago)

I think that you are right that there are a multiple species hiding under the name Panaeolus papilionaceus.

Whether any of those other species are the type collections of Panaeolus sphinctrinus, P. retirugus or P. campanulatus is the question that needs to be answered.


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Re: Panaeolus phylogenetic tree [Re: knarkkorven]
    #20680800 - 10/09/14 05:03 PM (9 years, 4 months ago)

Quote:

knarkkorven said:
I think confusion and miss identifications could have caused a lot of mixed samples in the collections, perhaps even leading to the false misconception of these being all the same species.






The same problem is widespread in Ganoderma as well.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #20680821 - 10/09/14 05:06 PM (9 years, 4 months ago)

It might be an idea to start from scratch and make your own collections, not give them name but assign them numbers so the testing results stay objective. Where do the samples for this phylogenetic come from?


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Re: Panaeolus phylogenetic tree [Re: maynardjameskeenan]
    #20680851 - 10/09/14 05:15 PM (9 years, 4 months ago)

I noticed that I wrote almost the same thing 3 months back, but this is a thing that keep nagging me when I'm out picking mushrooms... the color difference (one is more brown, the other gray) and the gray young grays have the saw tooth edge. This is the most apparent signs I have noticed about these two. Maybe I should collect some samples if anyone anytime is interested in writing a paper about it or something in the future... :wink:

Here's another couple of photos:


and the two groups to the left here:


Edited by knarkkorven (10/09/14 05:19 PM)


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Re: Panaeolus phylogenetic tree [Re: knarkkorven]
    #20680914 - 10/09/14 05:27 PM (9 years, 4 months ago)

I found some Panaeolus that look just like the ones on the left side of the top picture.
I thought they were just Panaeolus acuminatus :shrug:

http://mushroomobserver.org/162977?q=2GACl ( a couple of these mushrooms don't look like acuminatus to me)
http://mushroomobserver.org/160303?q=2GACl


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Re: Panaeolus phylogenetic tree [Re: maynardjameskeenan]
    #20680970 - 10/09/14 05:37 PM (9 years, 4 months ago)

Quote:

maynardjameskeenan said:
It might be an idea to start from scratch and make your own collections, not give them name but assign them numbers so the testing results stay objective. Where do the samples for this phylogenetic come from?





That's a great idea, and I look forward to doing that.  People should definitely start saving well documented Panaeolus collections.

All of the sequences in my analysis were downloaded from genbank.  Some of them are better quality than others.  Some of them have been well studied and named correctly, and some of them have names that were just guesses.   

One way to sort it out is to only include sequences from the people that you trust to accurately ID Panaeolus in your analysis.  Another is to request the herbarium collections and do your own microscopic analysis. 

Even genbank records with incorrect names are still valuable information.  Just not as valuable as it would be if we had good photos and microscopy to back up the names.


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Re: Panaeolus phylogenetic tree [Re: maynardjameskeenan]
    #20680987 - 10/09/14 05:40 PM (9 years, 4 months ago)

Sorry to go off topic, but this prooves how hard it is to distinguish between some of the Panaeolus...

If that's acuminatus, what's this? :



I believe the acuminatus is the ones with very dark cap/stipes, growing late at the season.
If I'm wrong I'm contributing to the confusion since I have this on my website :laugh: :

http://knarkkorven.magiskamolekyler.org/svampinfo/foton/panaeolusacuminatus.jpg


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller] * 1
    #20681383 - 10/09/14 07:06 PM (9 years, 4 months ago)

I'll help with the Panaeolus collections. I have a few well documented ones I can contribute.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #20681900 - 10/09/14 09:03 PM (9 years, 4 months ago)

I will do that too.
I'll post them on mushroom observer and use the observation number as its name, I'll also keep a few of them in an organized herbarium so they can be tested. I need to do a lot of work on my micrographs.


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Re: Panaeolus phylogenetic tree [Re: knarkkorven] * 2
    #20682147 - 10/09/14 10:36 PM (9 years, 4 months ago)

Thanks for posting in this thread knarkkorven, that reminded me that I have sequences from the Panaeolus alcis that you found.  I made a new tree with that sequence, and also a Panaeolus papilionaceus that I found in Mexico.

Here is the new tree:



The collections that I made sequences for have mushroomobserver.org observation numbers so you can see the photos.  My Panaeolus papilionaceus is exactly the same sequence as two sequences from Europe.  The Panaeolus alcis sequence is only three base pairs different from the Panaeolus papilionaceus sequence, and that isn't enough difference for it to show up somewhere else in the tree.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller] * 2
    #20682711 - 10/10/14 02:18 AM (9 years, 4 months ago)

Here is the newest version - this has 8 sequences I got from UNITe, plus sequences from other genera in the Psathyrellaceae.




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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller] * 3
    #20690804 - 10/11/14 09:48 PM (9 years, 4 months ago)

The above version sucked because I included sequences that were too far away from Panaeolus.  The truth is that Panaeolus is a very unique genus - not closely related to anything else.  The closest thing it is related to is Copelandia, however it isn't very close to that.  Copelandia is most closely related to Psilocybe, however Copelandia is far away from Psilocybe.

To make an accurate tree of Panaeolus and Copelandia I used Psilocybe semilanceata as an outgroup to root the tree.  P. semilanceata was chosen because it is difficult to misidentify, and is the closest relative to Panaeolus and Copelandia outside of those genera.

This tree was made by gathering sequences from Genbank and UNITe that were entered under the names Panaeolus and Copelandia, and also by BLASTing Panaeolus and Copelandia sequences to discover others that were in these genera but were entered under different names.  I added two Panaeolus sequences from my collections that I made in the lab, and one Copelandia sequence that was sent to me via facebook message.

The sequences were automatically aligned with kalign.  The FASTA was downloaded and loaded into mega6, where I went over the alignment manually and deleted any base pairs that were obviously in error.  Care was taken to not delete any phylogenetically informative data.  In borderline cases where I couldn't tell if the data was phylogenetically informative, I kept the data.

The FASTA was saved from mega6 and uploaded to phylogeny.fr, and the final tree was generated with the maximum liklihood algorithm using PhyML.  Gblocks was disabled and bootstrapping was run with 100 iterations. 

Here is the final result:



From this I learned that the traditional classification of Panaeolus into sections is not supported by DNA evidence.  Panaeolus and Copelandia deserve their own family.  In the most recent Panaeolus monograph, Gerhardt 1996, he moved Copelandia back into Panaeolus and kept Panaeolina as a separate genus.  The truth is the opposite - Copelandia is a separate genus, and Panaeolina is a clade within Panaeolus.

Panaeolus contains the following clades:

/cinctulus - One species occurs in Estonia, China and Japan.  Another species in this group occurs in China.  Panaeolus subfirmus is in the P. cinctulus clade, but is a separate species.
/antillarum - Contains Panaeolus antillarum and P. semiovatus
/fimicola - A separate clade, closely related to /antillarum
/papilionaceus clade 1 - Contains Galeropsis desetorum, in addition to Panaeolus papilionaceus collections from Florida and Italy
/papilionaceus clade 2 - Contains P. papilionaceus-like species from Washington, Italy and Mexico
/acuminatus - known from Estonia, Italy and China.  The Chinese species is slightly different.
/foenisecii - sequences from Canada, Estonia, Italy, Illinois and Finland are extremely similar
/Copelandia genus - Not closely related to the rest of Panaeolus, its placement is between Panaeolus and Psilocybe.  It has no other close relatives.

To continue this work, I need more collections that are documented with good photos


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #20690823 - 10/11/14 09:53 PM (9 years, 4 months ago)

Quote:

The truth is the opposite - Copelandia is a separate genus, and Panaeolina is a clade within Panaeolus.





Ain't that some shit. Can you use this site without having Java? Have you tried putting any Panaeolopsis spp. in there?


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OfflineAlan RockefellerM
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Re: Panaeolus phylogenetic tree [Re: o8u]
    #20690841 - 10/11/14 09:58 PM (9 years, 4 months ago)

I haven't tried it with java disabled.  I think it would work just fine.  I don't have any sequences of "Panaeolopsis", however I am pretty sure they would fall within Panaeolus sensu stricto.  That's what Christine told me, and she sequenced a few.

I think they would fall within the /fimicola clade.




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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #20691080 - 10/11/14 11:06 PM (9 years, 4 months ago)

Quote:

Alan Rockefeller said:

  In the most recent Panaeolus monograph, Gerhardt 1996, he moved Copelandia back into Panaeolus and kept Panaeolina as a separate genus.  The truth is the opposite - Copelandia is a separate genus, and Panaeolina is a clade within Panaeolus.





And yet we still say 'Panaeolus cyanescens and Panaeolina foenisecii'...

Cool stuff Alan! :thumbup:


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Re: Panaeolus phylogenetic tree [Re: maynardjameskeenan]
    #20691121 - 10/11/14 11:18 PM (9 years, 4 months ago)

I just simply wish I as close to as smart as you guys.  I have been studying my ass off for 2 straight yeasr and mostly keeping my mouth shut ( as i should be) and I am a lover of science with with biology degree from the university of Oregon but gosh darn it i am always many steps behind on these threads as I try to catch up. sad sad sad sad , eager eager eager, one in a million, so you are saying there is a chance...??? Will one of you please spend an afternoon with me at the location of your choosing??  maybe Hawaii and I will provide a condo near the beach.......


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Re: Panaeolus phylogenetic tree [Re: rlmich] * 1
    #20691149 - 10/11/14 11:26 PM (9 years, 4 months ago)

Quote:

rlmich said:
I just simply wish I as close to as smart as you guys.





I will teach you everything I know, if you will listen.

Quote:

I have been studying my ass off for 2 straight yeasr and mostly keeping my mouth shut ( as i should be) and I am a lover of science with with biology degree from the university of Oregon but gosh darn it i am always many steps behind on these threads as I try to catch up. sad sad sad sad




The more time you spend in school the less you know.  There is no substitute for real world experience!



Quote:

Will one of you please spend an afternoon with me at the location of your choosing??  maybe Hawaii and I will provide a condo near the beach.......





I am in Jalpan de Serra, Queretaro, Mexico.  Everyone is invited.  It's raining right now.  It's a good time to come.  I'll be here for six more weeks.

But if you want to fly me out to Hawaii that would be cool too.  Never been there.

If there is a plane ticket and a place to stay I'll show up anywhere on planet earth and teach mycology until you are tired of listening.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #20691244 - 10/12/14 12:06 AM (9 years, 4 months ago)

Excellent work Alan:thumbup:


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Re: Panaeolus phylogenetic tree [Re: Mrcloudy]
    #20691488 - 10/12/14 03:27 AM (9 years, 4 months ago)

Quote:

Excellent work Alan




I Agree! :thumbup: :thumbup:

It would have been great if there was a DNA sequence of P. olivaceus included in the tree, but I guess there are none made yet?


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Re: Panaeolus phylogenetic tree [Re: knarkkorven]
    #20691872 - 10/12/14 09:05 AM (9 years, 4 months ago)



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Re: Panaeolus phylogenetic tree [Re: knarkkorven] * 1
    #20692257 - 10/12/14 11:00 AM (9 years, 4 months ago)

Quote:

knarkkorven said:
Quote:

Excellent work Alan




I Agree! :thumbup: :thumbup:

It would have been great if there was a DNA sequence of P. olivaceus included in the tree, but I guess there are none made yet?




Finally, a tree that makes sense. This is outstanding work. I would bet that P. olivaceus will fall into the fimicola clade based on microscopic features and psilocybin content.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller] * 1
    #20692321 - 10/12/14 11:20 AM (9 years, 4 months ago)

Quote:

Alan Rockefeller said:
The above version sucked because I included sequences that were too far away from Panaeolus.  The truth is that Panaeolus is a very unique genus - not closely related to anything else.  The closest thing it is related to is Copelandia, however it isn't very close to that.  Copelandia is most closely related to Psilocybe, however Copelandia is far away from Psilocybe.



Do you have a cladogram that supports these claims? Yours show Copelandia as basal to Panaeolus/the rest of Panaeolus; as close to Psilocybe as the rest of Panaeolus is. While Panaeolus might not be closely related to Psathyrellaceae (which your cladogram doesn't show), that's not particularly informative when other cladograms have found:

Moncalvo et al. 2002. One hundred and seventeen clades of euagarics.
--+--Bolbitiaceae
  |--Descolea gunnii
  `--+--Galerina nana
      `--Panaeolus spp. (="Panaeoloideae" in this tree, but only Panaeolus is included in this tree.

Matheny et al. 2006. Major clades of Agaricales: a multilocus phylogenetic overview.
--+--Panaeolus spp.(="Panaeolae": Panaeolus+Panaeolina)
  |--Tubariae
  `--+--Crepidotaceae
      `--Inocybaceae

Neither of these trees found Paneolus to be all that close to Psilocybe. You'd need to make a pretty large cladogram including members of many different clades within Euagaricales to resolve the placement of Panaeolus. It'd be cool if there was a mycological equivalent of the Angiosperm Phylogeny Group.


By how much do Copelandia and the rest of Panaeolus actually differ? Whether or not this difference is significant, splitting Copelandia from Panaeolus is still just a matter of personal preference since Panaeolus is monophyletic whether or not it includes Copelandia. Plus, species of Panaeolus that aren't in your cladogram might end up being basal to Copelandia+Panaeolus

Quote:

From this I learned that the traditional classification of Panaeolus into sections is not supported by DNA evidence.  Panaeolus and Copelandia deserve their own family.  In the most recent Panaeolus monograph, Gerhardt 1996, he moved Copelandia back into Panaeolus and kept Panaeolina as a separate genus.  The truth is the opposite - Copelandia is a separate genus, and Panaeolina is a clade within Panaeolus.



Panaeolae has already been named. You could name that clade "Panaeolaceae" to give it a family-level, but that clade would be synonymous with Panaeolae, and Panaeolae is synonymous with Panaeolus unless you split Copelandia from it. Cool results overall though, especially with the clades of Panaeolus.


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Re: Panaeolus phylogenetic tree [Re: heelsplitter]
    #20692330 - 10/12/14 11:23 AM (9 years, 4 months ago)

Quote:

Alan Rockefeller said:
/cinctulus - One species occurs in Estonia, China and Japan.  Another species in this group occurs in China.  Panaeolus subfirmus is in the P. cinctulus clade, but is a separate species.




Makes sense, P. subfirmus has bigger spores and looks similar to P. cinctulus.

Quote:

[/antillarum - Contains Panaeolus antillarum and P. semiovatus
/fimicola - A separate clade, closely related to /antillarum



Both have sulphidia.

Quote:

/foenisecii - sequences from Canada, Estonia, Italy, Illinois and Finland are extremely similar




I wonder if this will hold up if we added several more collections?  Collections from India that match P. indica would be especially interesting.  Everyone should start photographing and saving Copelandia/Panaeolus/Panaeolina collections.

Quote:

heelsplitter said:
By how much do Copelandia and the rest of Panaeolus actually differ? Whether or not this difference is significant, splitting Copelandia from Panaeolus is still just a matter of personal preference since Panaeolus is monophyletic whether or not it includes Copelandia. Plus, species of Panaeolus that aren't in your cladogram might end up being basal to Copelandia+Panaeolus




Copelandia has metuloid cystidia, Panaeolus including Panaeolina don't.


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OfflineAlan RockefellerM
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Re: Panaeolus phylogenetic tree [Re: heelsplitter]
    #20695113 - 10/12/14 10:27 PM (9 years, 4 months ago)

Quote:

knarkkorven said:
It would have been great if there was a DNA sequence of P. olivaceus included in the tree, but I guess there are none made yet?




There is a Panaeolus olivaceus sequence in UNITe, but it's actually a Deconica.

Quote:

heelsplitter said:
Do you have a cladogram that supports these claims?




The tree I posted above with the labeled clades supports these claims.

Quote:

Yours show Copelandia as basal to Panaeolus/the rest of Panaeolus; as close to Psilocybe as the rest of Panaeolus is.




It doesn't say that at all.  Look at the numbers - they are super important!  Copelandia is 1.2 units away from Psilocybe semilanceata, and the closest Panaeolus is 2.5 units away.  You need to add up all the numbers in the branches to see how far away from P. semilanceata each branch is.  That's a huge difference, and shows that Copelandia is much closer to Psilocybe than anything in Panaeolus.

The numbers are calculated from the number of base pair differences.

Quote:

While Panaeolus might not be closely related to Psathyrellaceae (which your cladogram doesn't show), that's not particularly informative when other cladograms have found:




I read the Moncalvo paper and tried all of the things he said were close as outgroups.  All of them flattened the tree to a level where it was nearly useless because none of them were actually close.  Once I deleted the stuff he claimed were close, the tree became useful.

Following the Matheny paper, I added a bunch of Inocybe and Tubaria species, and the same thing happened, but even worse.

I also tried adding various species from the Psathyrellaceae, same result.  None of the stuff that was supposed to be in the same family was at all close.

I have no idea how those guys got these results, but in my time looking at the data I haven't seen anything to suggest that either is even close to being correct.

If either of these papers were correct, when I do a BLAST search on Panaeolus or Copelandia species, I would expect to see those genera start showing up after I scroll down past the Panaeolus/Copelandias.  I don't.  What I see is a bunch of Panaeolus/Copelandia, then Psilocybes start to show up.  That indicates that Psilocybe is closest, and when I chose a Psilocybe outgroup the tree looked great because the sequence wasn't so distant that everything else was almost identical by comparison.


Quote:

By how much do Copelandia and the rest of Panaeolus actually differ?




A lot.  Like night and day.  As I was manually aligning sequences, I noticed many areas that the Copelandias had in common, but none of the Panaeolus had.

Quote:

Whether or not this difference is significant, splitting Copelandia from Panaeolus is still just a matter of personal preference since Panaeolus is monophyletic whether or not it includes Copelandia.




Panaeolus and Copelandia are super different.  I'll post the final alignment I used after I cleaned it up, and you can visually scroll through it and see how different they are.  It's a lot easier to see the actual differences that way rather than just trusting numeric calculations. 

Quote:

Plus, species of Panaeolus that aren't in your cladogram might end up being basal to Copelandia+Panaeolus




All the Panaeolus had only minor differences, and all the Copelandia had only minor differences, compared to the difference between Panaeolus and Copelandia.


Quote:

Panaeolae has already been named. You could name that clade "Panaeolaceae" to give it a family-level, but that clade would be synonymous with Panaeolae, and Panaeolae is synonymous with Panaeolus unless you split Copelandia from it. Cool results overall though, especially with the clades of Panaeolus.




Copelandia should be split off, and Panaeolaceae sounds like a great family name to put those two genera in.


Thanks for taking a critical look at this.  You asked all of the hard questions that I just spent the last two days wrestling with.  I posted this in about 25 mushroom forums and thousands of people saw it, and everyone else just said "whoa, cool" or "I wish I understood this stuff".  A dozen people contacted me to ask how they could become a mycologist.

Please let me know if my arguments are convincing at all, or if I got something wrong somewhere.



Here is the FASTA file of the final alignment, after I cleaned it up and deleted the noise
http://plantobserver.org/Panaeolus-kalign-mega6manualalign.fas

mega6 session file. 
http://plantobserver.org/Panaeolus.kalign.mas

The sequences after being aligned with kalign, but before I changed them in any other way
http://plantobserver.org/Panaeolus.kalign.fas

How to do your own analysis using these files:  Go to http://phylogeny.lirmm.fr, click on Phylogeny Analysis.  Upload one of the .fas files.  Use the default settings or change them around.  Click go and wait about 3 minutes.

Or load any of the three files into mega6 and try the various tree building options.  I tried this and got the same results, but the trees didn't look as nice.

Note that these sequences have been renamed - to add the country when available, and to fix sequences that were uploaded with the wrong names.  I think I got the names right, but I could be wrong.  You can use the accession numbers to get the original and likely less accurate names from Genbank and UNITe.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #20695217 - 10/12/14 11:02 PM (9 years, 4 months ago)

I wonder where Panaeolus guttulatus would fit into this...

Quote:

5 Gill edge (pocket-lens) with yellowish dropiets by
sticky secretes of cheilocystidia (fig. 3 f); spores with
oildrops; sulphidia absent    . .  .  .        P. sect. Guttulati
Only species:    Panaeolus guttulatus (29)




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Re: Panaeolus phylogenetic tree [Re: Byrain]
    #20695250 - 10/12/14 11:15 PM (9 years, 4 months ago)

Quote:

Byrain said:
I wonder where Panaeolus guttulatus would fit into this...





Me too.  Luckily, someone in Europe is mailing me some this week.


If anyone else has Panaeolus, Panaeolina or Copelandia collections, I would like to add them to this tree.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #20695263 - 10/12/14 11:20 PM (9 years, 4 months ago)

Cool!  Do they have photos and/or microscopy?  When and where do you want more collections?


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Re: Panaeolus phylogenetic tree [Re: Byrain]
    #20695269 - 10/12/14 11:22 PM (9 years, 4 months ago)

Quote:

Byrain said:
Cool!  Do they have photos and/or microscopy?  When and where do you want more collections?





He promised P. guttulatus and P. olivaceus and unspecified others, he didn't say if he had photos or microscopy. 

You can drop the collections in the mail any time in October or November.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #20695482 - 10/13/14 12:56 AM (9 years, 4 months ago)

Quote:

It doesn't say that at all.  Look at the numbers - they are super important!  Copelandia is 1.2 units away from Psilocybe semilanceata, and the closest Panaeolus is 2.5 units away.  You need to add up all the numbers in the branches to see how far away from P. semilanceata each branch is.  That's a huge difference, and shows that Copelandia is much closer to Psilocybe than anything in Panaeolus.



That's a pretty good reason for splitting off Copelandia. My issue with splitting off Copelandia was just that Panaeolus would be monophyletic whether or not it was included. A subgenus basal to the rest of a genus and a separate genus for that subgenus would still be the same clade, so the Linnean rank it's given doesn't really change anything. Uncertainty in the monophyly of that genus does.

Quote:

If either of these papers were correct, when I do a BLAST search on Panaeolus or Copelandia species, I would expect to see those genera start showing up after I scroll down past the Panaeolus/Copelandias.  I don't.  What I see is a bunch of Panaeolus/Copelandia, then Psilocybes start to show up.  That indicates that Psilocybe is closest, and when I chose a Psilocybe outgroup the tree looked great because the sequence wasn't so distant that everything else was almost identical by comparison.



That's pretty exciting! Do other members of Hymenogastraceae start showing up after Psilocybe?

Quote:

Panaeolae has already been named. You could name that clade "Panaeolaceae" to give it a family-level, but that clade would be synonymous with Panaeolae, and Panaeolae is synonymous with Panaeolus unless you split Copelandia from it. Cool results overall though, especially with the clades of Panaeolus.




Quote:

Copelandia should be split off, and Panaeolaceae sounds like a great family name to put those two genera in.



I checked and Panaeolaceae has already been named according to mycobank. Mycobank lists it as "invalid" but doesn't give a reason. I'm thinking that it's probably because Panaeolaceae is a junior synonym of Panaeoleae (Panaeolae was a typo on my part). Panaeoloideae is also a junior synonym of Panaeoleae. Panaeolaceae looks nicer, but it's the same thing as Panaeoleae.I don't know what rank Panaeoleae is supposed to have, but whether that rank is above or below family, Panaeoleae and Panaeolaceae still refer to the same clade.

I have some Copelandia specimens that I've been planning to send you for a while but haven't gotten around to yet due to being busy with school work. I'll send those out pretty soon and add some Panaeolus (cf. cinctulus and cf. foenisecii) specimens too.


Edited by heelsplitter (10/13/14 12:52 PM)


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Re: Panaeolus phylogenetic tree [Re: heelsplitter]
    #20695497 - 10/13/14 01:02 AM (9 years, 4 months ago)

Quote:

heelsplitter said:
That's pretty exciting! Do other members of Hymenogastraceae start showing up after Psilocybe?




Yes, there's quite a bit of Galerina and Flammula showing up as well.  Try NCBI blast on a few Panaeolus and Copelandia and see what you think.


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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #20890328 - 11/25/14 06:48 PM (9 years, 2 months ago)

Hey I was going to send you that blueing Flammulina at some point do you still want Panaeolus species for the tree? I could throw in a cinctulus if it is helpful.

I also wanted to ask is there any updates on whether is Panaeolus foenisecii again or still Panaeolina? Want to call it by the proper name I remember you saying they might or did change it back. Maybe that was an old thread.


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Edited by d0urd3n (11/25/14 06:49 PM)


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Re: Panaeolus phylogenetic tree [Re: d0urd3n]
    #20890415 - 11/25/14 07:02 PM (9 years, 2 months ago)

I believe I still have P. fimicola collection somewhere. Its about two years old. Is it still any value to your research?


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Species List (Georgia): Psilocybe caerulescens/weilii, Psilocybe atlantis/galindoi, Psilocybe cubensis, Psilocybe ovoideocystidiata, Psilocybe caerulipes, Psilocybe semilanceata, Psilocybe fagicola, Copelandia cyanescens, Panaeolus cinctulus, Panaeolus fimicola, Panaeolus olivaceus, Gymnopilus luteofolius, Gymnopilus aeruginosus, Gymnopilus junonius, Pluteus salicinus

(Ohio): Psilocybe ovoideocystidiata, Psilocybe caerulipes, Pluteus cyanopus, Pluteus salicinus sensu lato..., Panaeolus cinctulus, Gymnopilus luteus, Gymnopilus luteofolius, Gymnopilus junonius, Gymnopilus aeruginosus


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OfflineAlan RockefellerM
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Re: Panaeolus phylogenetic tree [Re: d0urd3n]
    #20891451 - 11/25/14 11:26 PM (9 years, 2 months ago)

Quote:

d0urd3n said:
Hey I was going to send you that blueing Flammulina at some point do you still want Panaeolus species for the tree? I could throw in a cinctulus if it is helpful.




Yes, I am interested in the P. cinctulus group.

Quote:

I also wanted to ask is there any updates on whether is Panaeolus foenisecii again or still Panaeolina? Want to call it by the proper name I remember you saying they might or did change it back. Maybe that was an old thread.





I think Panaeolus foenisecii is the best name for it, but I would like to check the LSU sequences as well since that locus is better for determining genus level differences.


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Offlineheelsplitter
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Re: Panaeolus phylogenetic tree [Re: Alan Rockefeller]
    #22452045 - 10/30/15 01:59 AM (8 years, 3 months ago)
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Quote:

Alan Rockefeller said:
Quote:

Yours show Copelandia as basal to Panaeolus/the rest of Panaeolus; as close to Psilocybe as the rest of Panaeolus is.




It doesn't say that at all.  Look at the numbers - they are super important!  Copelandia is 1.2 units away from Psilocybe semilanceata, and the closest Panaeolus is 2.5 units away.  You need to add up all the numbers in the branches to see how far away from P. semilanceata each branch is.  That's a huge difference, and shows that Copelandia is much closer to Psilocybe than anything in Panaeolus.

The numbers are calculated from the number of base pair differences.




This just means that Panaeolus species are more divergent from the common ancestor of Panaeolus, Copelandia and Psilocybe than Copelandia species are AFAIK. Copelandia isn't more closely related to Psilocybe since Copelandia and Panaeolus share a more recent common ancestor, which the cladogram shows.

Quote:

Alan Rockefeller said:
Quote:

While Panaeolus might not be closely related to Psathyrellaceae (which your cladogram doesn't show), that's not particularly informative when other cladograms have found:




I read the Moncalvo paper and tried all of the things he said were close as outgroups.  All of them flattened the tree to a level where it was nearly useless because none of them were actually close.  Once I deleted the stuff he claimed were close, the tree became useful.

Following the Matheny paper, I added a bunch of Inocybe and Tubaria species, and the same thing happened, but even worse.

I also tried adding various species from the Psathyrellaceae, same result.  None of the stuff that was supposed to be in the same family was at all close.

I have no idea how those guys got these results, but in my time looking at the data I haven't seen anything to suggest that either is even close to being correct.

If either of these papers were correct, when I do a BLAST search on Panaeolus or Copelandia species, I would expect to see those genera start showing up after I scroll down past the Panaeolus/Copelandias.  I don't.  What I see is a bunch of Panaeolus/Copelandia, then Psilocybes start to show up.  That indicates that Psilocybe is closest, and when I chose a Psilocybe outgroup the tree looked great because the sequence wasn't so distant that everything else was almost identical by comparison.




The issue there is just that you're comparing ITS results to some (not super great) multilocus analyses. Dentinger et al. (2015) (see the link) proved that Moncalvo's and Matheny's analyses weren't very accurate, but an ITS tree is likely to be even less accurate. 

Quote:

Alan Rockefeller said:

Yes, there's quite a bit of Galerina and Flammula showing up as well.  Try NCBI blast on a few Panaeolus and Copelandia and see what you think.




I ran BLAST on a couple of these a while back and didn't get around to posting about it until now. I mostly got Psilocybe, Alnicola, Naucoria and Galerina showing up. That would place Panaeolus in Hymenogastraceae or just somewhere in the "stropharioid clade", but the ITS region alone isn't super great for determining intergeneric relationships. It would be pretty sweet if someone sequenced the whole genome of a Panaeolus species so that it could be compared with Dentinger et al. (2015) (link attached, it's pretty exciting stuff) so its position could be resolved, but they also didn't include Hymenogaster citrinus, so that would have to be done too. I know it's possible to make a meta-tree somehow that would just use Dentinger et al.'s analysis as a framework to compare one locus or a couple loci from a Panaeolus species to, but it seems really intimidating and difficult.


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InvisibleJoust
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Re: Panaeolus phylogenetic tree [Re: heelsplitter]
    #22452271 - 10/30/15 05:15 AM (8 years, 3 months ago)

Quote:

The issue there is just that you're comparing ITS results to some (not super great) multilocus analyses. Dentinger et al. (2015) (see the link) proved that Moncalvo's and Matheny's analyses weren't very accurate, but an ITS tree is likely to be even less accurate. 




Ive found this to be true in my studies as well.


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~~~~~~***Psilocybin Mushrooms***~~~~~~
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