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Collecting and Isolating Wild Yeast

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Zabuza

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So I've done several searches and read several threads, but none of them really outline how to capture/collect wild yeast and then isolate a singular strand (or isolate it from the other baddies in the air). For example, I've read https://www.homebrewtalk.com/entries/collecting-wild-yeast.html and although this seems close to what I'm talking about, the author never makes explicit instructions for isolating yeast strands. I'd imagine that letting a petri dish sit out or dropping a piece of local fruit into some wort would not really guarantee yeast purity (I understand there aren't really guarantees without a lab, per se, merely increasingly high probabilities of purity, but you know what I mean). I've also read https://www.homebrewtalk.com/f127/howto-capture-wild-yeast-101886/ in which the author basically makes a spontaneously fermented starter, then steps it up and pitches it into a full batch.

These are both interesting methods of obtaining wild yeast colonies, but the authors (at least according to my limited understanding) do not demonstrate how to completely isolate a wild, pure yeast strain.

I've certainly read https://www.homebrewtalk.com/f163/slanting-yeast-133103/ and the linked http://braukaiser.com/wiki/index.php?title=Making_Plates_Slants&printable=yes therein, but neither of those articles address wild yeast. They provide impeccable methods of isolating pure strains, but not from wild cultures.

So, given all of this background literature, I have a relatively simple question: could I capture wild yeast using one of the methods described in the first paragraph then apply isolation techniques from the last paragraph to obtain a pure strain? One hypothetical method would be to let the petri dish sit out, allow yeast colonies to form, grow one (but not necessarily pure) up to a small starter, then use an inoculation needle to streak some petri dishes. This would (I'm guessing from a purely hypothetical standpoint) allow for the separation of perhaps numerous wildly settled strains in the original petri dish by harvesting single cell colonies in the manually inoculated petri dish. Or, I could drop some wild fruit into some wort or allow the wort to sit for a week or two (the other two methods mentioned in the above articles) until I had a foaming starter, then manually inoculate a petri dish from that starter for a pure colony.

Is my methodology secure here, or is there something I'm missing? Would these methods work for obtaining pure strands of wild yeast? I simply couldn't find any start-to-finish guides for isolating pure strands of wild yeast when I searched.
 

Calichusetts

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Your methods seems pretty straightforward and I am sure they would work, though you would never be certain it is a single strain. BUT, if you wash the yeast after I'm sure the most dominate strain would begin to emerge. After a few generations you would be more certain its a single strain...maybe.

Consider writing to Mystic in Massachusetts. They are doing a wild yeast series and I am sure they could help you with these questions.
 
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Zabuza

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I will write to Mystic, thanks for the tip. In the articles, the authors talk about isolating single-cell colonies. Why wouldn't those be pure colonies? Did I misunderstand something?
 
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Zabuza

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Can anyone perhaps add a bit more to this? Are the techniques outlined above not effective for isolating pure strains of wild yeast? That seems odd, since it was the entire point of their existence.
 
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Zabuza

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EarlyAmateurZymurgist said:
The technique that one uses for isolating wild yeast is the same technique that one uses to isolate pure cultures of brewer's yeast. The most commonly used technique is known as a "four quadrant streak." http://en.wikipedia.org/wiki/Streaking_%28microbiology%29 Here's a web page that explains the procedure in fairly good detail: http://amrita.vlab.co.in/?sub=3&brch=73&sim=213&cnt=2
Ok, good, that was exactly what I linked to above, so...thanks. I'm aware of the method, I just got confused when I was told it wouldn't isolate a pure yeast strand.
 

EarlyAmateurZymurgist

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Ok, good, that was exactly what I linked to above, so...thanks. I'm aware of the method, I just got confused when I was told it wouldn't isolate a pure yeast strand.
Strands are not necessarily pure yeast cultures. Strands contain the offspring of multiple yeast cells.

Here's a photo of a plate that I streaked several weeks ago:



What you are after are isolated round colonies. The rectangle contains the colonies that I harvested from the plate. Round colonies are formed when isolated single yeast cells divide. Each round colony is therefore a single-cell pure culture.
 
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Zabuza

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Stands are not necessarily pure yeast cultures. Strands contain the offspring of multiple yeast cells.

Here's a photo of a plate that I streaked several weeks ago:



What you are after are isolated round colonies. The rectangle contains the colonies that I harvested from the plate. Round colonies are formed when isolated single yeast cells divide. Each round colony is therefore a single-cell pure culture.
Ok so if I'm understanding you correctly you're saying a strand does not necessarily equal a pure, single-cell colony. Fair enough - strands might contain blends of various yeast colonies. I guess my question, though, was whether using these techniques would allow me to isolate a single-cell pure culture. The original responder said no, that this was impossible. I take it you disagree? It seems as though the methods you and I linked to would allow one to isolate single-cell colonies, as I asserted in my original post.
 

EarlyAmateurZymurgist

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Well-isolated round colonies are almost always pure cultures. However, with a wild sample, being able to isolate pure cultures is just the first step in identifying one's target yeast strain, as there may be more than more yeast strain in one's sample. The best way to do that in home setting is to slant colonies that exhibit good morphology, one colony per slant. After the slants have been incubated, one can take a sample from each slant and step it up to a pitchable size using aseptic transfer technique (the remaining yeast on each slant becomes the reference culture from which other slants are made).

I wouldn't brew a full-size batch until I was sure that I had identified a useful strain. It's imperative that your small starters are absolutely sterile, which means that the starter wort needs to be autoclaved (pressure cooked) in its container at 121C (250F) for at least 15 minutes. A generally accepted first stage starter contains 5 to 10 milliliters of absolutely sterile 1.020 to 1.030 wort (I have gone as high as 30mls when taking a large sample from a slant). The first stage starter is used to inoculate 50 to 100 milliliters of absolutely sterile 1.020 to 1.030 wort. A 50 to 100 milliliter starter can be used to inoculate a 1L test batch.
 

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