DNA barcoding of mushrooms

[peter russell]

I am a complete novice but want to do some DNA barcoding of fungi around where I live in New York.  Am looking to visit my local community biolab but had a few basic questions.  I have a herbarium of dried fungi .... am I likely to have any success with these or do I really need fresh specimens?  Has anybody tried this on Waxcaps and if so what [cheap][simple] extract protocol was successful?  If I am asking this in the wrong forum I apologise... if anybody can suggest an approriate forum I would appreciate it!

[Dakota Hamill]

Check through some of my old blog posts on www.basementbiotech.org we barcoded a ton of fungi.  Dried mushrooms are fine.  We had 3 year old dessicated mushrooms we got ITS bands on with a simple ethanol prep.

On Jul 10, 2015 5:22 PM, "'peter russell' via DIYbio" <diy...@googlegroups.com> wrote:

I am a complete novice but want to do some DNA barcoding of fungi around where I live in New York.  Am looking to visit my local community biolab but had a few basic questions.  I have a herbarium of dried fungi .... am I likely to have any success with these or do I really need fresh specimens?  Has anybody tried this on Waxcaps and if so what [cheap][simple] extract protocol was successful?  If I am asking this in the wrong forum I apologise... if anybody can suggest an approriate forum I would appreciate it!

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[Nathan McCorkle]

If you don't know what you're looking at: Look up 28S or ITS2 barcode
sequencing.

Here is a preparation protocol, which would get any samples ready for
sending off for sequencing:
http://1ofdmq2n8tc36m6i46scovo2e.wpengine.netdna-cdn.com/wp-content/uploads/2015/02/iTag-Sample-Amplification-QC-v1.1.pdf

If you know what mushroom you're looking for, and there are
pre-existing primers for a unique-to-that-species gene-fragment, then
you may be able to get around sequencing and simply do a PCR.

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[Dakota Hamill]

ITS1F ITS4R I think are the primers.  Nathan's protocol is probably good.  When I'm not on my phone I'll follow up later

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[Nathan McCorkle]

On Fri, Jul 10, 2015 at 9:51 AM, Dakota Hamill <dko...@gmail.com> wrote:
> ITS1F ITS4R I think are the primers. Nathan's protocol is probably good.
> When I'm not on my phone I'll follow up later

I just went through it, and it is pretty high-level. Doesn't actually
cover extraction... only setting up the PCR pretty much.

[peter russell]

Thanks for the speedy responses.... I've a lot to learn!

[Josiah Zayner]

We have Fungal ITS primers on The ODIN for purchase http://www.the-odin.com/fungal-its-pcr-primers-for-identification-and-barcoding/
You can also purchase Taq and other things you need for the PCR. Let me know if you need some help setting up an experiment.

Josiah Zayner

[Pjrthe1st]

Thx very much!  I will look at that. I was going to attend a intro course at genspace first so will be probably be active in the fall

Sent from my iPhone

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