Why you don't want to use the distilled water yeast storage method.

[spareparts]

I was talking to a retired faculty member today and it turns out that he's an old yeast scientist.

I was discussing the various methods that we homebrewers use in our yeast ranching methods and getting tips and advice.

He advised against the distilled water method. He said that in that method you are relying on the yeast to sporulate. When they do this they can survive for a very long time. I don't exactly remember the terms he used but he described the yeast spore forming what under a microscope will look like a pod with four yeast spores in side. When revived they will come out of sporulation and mate. Two of the spores will mate and you will effectively get those genetic traits but loose the traits from the other two which won't mate.

Anyway, I don't know if I'm completely describing the conversation accurately but what he summed up for us homebrewers is that sporulation is the worst method we can use for keeping a strain pure.

 

[CaptainCoconut]

Very interesting, did he say what would be the best?

 

[spareparts]

Very interesting, did he say what would be the best?

Freezing at -80C was his recommendation. He said he actually thawed some out that had been frozen for 20 and 40 years and was able to revive it.

He also recommended freeze drying. Since most of us homebrewers don't have access to a freeze drying machine we toyed with some other ideas for drying. I'm going to try a method and see how it works. I'll post details down the road.

 

[CaptainCoconut]

Well, most of us homebrewers don't have access to -80C freezer temperatures either. Let us know how the drying works.

 

[RogerMcAllen]

Post redacted

 

[rico567]

(Aaaaaand so much for that method!)

We tend to coddle our brewing yeast compared to baking yeast. My sourdough starter will keep for a month in the fridge with no apparent ill effects. If I want to keep it longer, I just pour out a few ounces on wax paper on a cookie sheet and throw it in the freezer. The freezer both freezes it and dries it out. The resulting flakes will keep for quite a while, and revive very nicely. My regular dry instant baking yeast I keep in the fridge and most of it in the freezer. Never seen any ill effects. Tough stuff, yeast.

 

[enid]

After reading the different options, the distilled water don't seem that bad. I have never had a problem doing it this way, and I just used a Kolsch yeast that was over 7 months old with no problems. If it is a limited strain I always put yeast in test tubes with glycerin and freeze and have revived them after a year. Until I hear of something that is easy, time and cost effective I will stick with the old way.

 

[logdrum]

and it continues to give dependable, vigorous fementation.

-d

 

[broadbill]

and it continues to give dependable, vigorous fementation.

-d

This is my feeling as well....I appreciate the professors academic opinion and he very well may be correct, but the question is: Does it really matter for homebrewers? From everything I've read and my experience with yeast I would have to say no. Even if it did matter, is there any cost- and/or time-permissive solution that we as homebrewers could do to prevent this?...again probably not.

We know over multiple passages (i.e. be washed/stored/reused), the yeast change/mutate and at a certain point we need to go back to a pure culture. Its sounds like from the professor that maybe these changes/mutations are being caused by repeated sporulations going on in between passages/beers. Good to know, but that doesn't really help with a viable alternative.

I think the best advice is still: Treat your yeast well (make starters when necessary, don't underpitch, wash to save your yeast) and only reuse 3-4 times at most. Any more and you are gambling on the possibility that you may have fermentation issues because that yeast is changed.

What I wonder is: From my reading it sounds as if sporulation occurs in conditions of nutrient deprivation (both nitrogen and maltose). Therefore, I would expect sporulation to be happening way before you get to the yeast washing/storing point. Sporulation is probably occurring post-fermentation as the yeast are flocculating and settling out, correct? It is my impression that yeast stop fermenting and flocculate because they've run out of nutrients. Thoughts?

 

[Revvy]

After reading the different options, the distilled water don't seem that bad. I have never had a problem doing it this way, and I just used a Kolsch yeast that was over 7 months old with no problems. If it is a limited strain I always put yeast in test tubes with glycerin and freeze and have revived them after a year. Until I hear of something that is easy, time and cost effective I will stick with the old way.

and it continues to give dependable, vigorous fementation.

-d

+1 to both of you!!

I don't often store my yeast in distilled water, but if I spontaenously decide to wash and harvest a batch of yeast when I am bottling or something, and don't have want to wait to boil and cool the water I need, I HAVE grabbed sealed gallons of distilled and used that. And never had any issues either.

 

[spareparts]

This is my feeling as well....I appreciate the professors academic opinion and he very well may be correct, but the question is: Does it really matter for homebrewers?

It will over time if you're trying to keep a special strain of yeast. After successive generations you will be selecting for traits in the yeast that allow it to survive washing and sporulation and deselecting for the traits that made you want to use that strain of yeast in the first place. For my White Labs strains wouldn't be concerned. But for my strains that were brought back from Europe, I want to keep them as close to the original as possible.

 

[enid]

But for my strains that were brought back from Europe, I want to keep them as close to the original as possible.

I agree. If it is something that is not readily available you would want to take different procedures. I am curious on thoughts about freezing in glycerin. I do not believe this would have the same sporulation effect you are talking about. But if it is a yeast that you use regularly and after 3-4 times washing you can simply go buy a new one, distilled water seems the easiest and most cost effective solution.

 

[broadbill]

It will over time if you're trying to keep a special strain of yeast. After successive generations you will be selecting for traits in the yeast that allow it to survive washing and sporulation and deselecting for the traits that made you want to use that strain of yeast in the first place. For my White Labs strains wouldn't be concerned. But for my strains that were brought back from Europe, I want to keep them as close to the original as possible.

If you wanted to preserve the strain characteristics of your special yeasts then you should also have a storage system in place to preserve the original strain (glycerol stocks of the original, yeast slants, etc). Whether or not you are using water to wash/store is irrelevant...that yeast is going to change/mutate over time regardless. Its not like you can stop that process.

 

[Boerderij_Kabouter]

If you wanted to preserve the strain characteristics of your special yeasts then you should also have a storage system in place to preserve the original strain (glycerol stocks of the original, yeast slants, etc). Whether or not you are using water to wash/store is irrelevant...that yeast is going to change/mutate over time regardless. Its not like you can stop that process.

I agree with this. Unless you have a way to maintain the original strain, no matter what technique you use to propagate and store, you will have mutation. You have mutation everything the yeast reproduces, regardless of the storage method. Unless you have a crazy science lab in your house. And by crazy, I mean you could start a lab business out of your house.

 

[CaptainCoconut]

Well, from the OP, it sounds like each cell will keep half and lose half of its traits. If you were trying to keep a single cell to revive later, that might be something to worry about, but with billions of cells, I can't see the whole quantity losing a trait, maybe just more vulnerable to mutation later on.

 

[JohnMc]

The sack of four spores is an ascus; Saccharomyces is an ascomycete (ascus forming fungus). 'Ascus' means bag or sack or something like that in Latin or Greek.
Each spore is haploid; like sperm and egg cells. When cut loose, they'll mate and form a diploid. In this process, the genetic stuff gets rearranged.
Somewhere in my memory, there's a recollection that brewing yeast tend to be aneuploid (an=not, eu=true, ploid~chromosome number), which makes them sporulate less. I wouldn't bank on that, though.

+1 on what others have said about genetic drift; it happens and you get what you select for, be it the ability to sporulate, higher or lower flocculation, survive washing, etc. Mutation is a part of life.

 

[broadbill]

Somewhere in my memory, there's a recollection that brewing yeast tend to be aneuploid (an=not, eu=true, ploid~chromosome number), which makes them sporulate less. I wouldn't bank on that, though.

You got it...I looked it up and that's what wikipedia said, so it must be right!

 

[evanos]

If you want to retain the yeast line's genetic characteristics 100%, it's true that you need to prevent sporulation and mating (even though if they did mate, theoretically they would be selfing, so the progeny would have the same genes, but in different combinations). As mentioned, this requires preserving the yeast in their current state. Roger McAllen's protocol would work, however, I would suggest using rubbing alcohol instead of acetone as it is safer (relatively) and will evaporate off slower than acetone during dessication.

 

[spareparts]

I agree with this. Unless you have a way to maintain the original strain, no matter what technique you use to propagate and store, you will have mutation.

This particular scientist I was talking to mentioned this as well. He said that even the big breweries with expensive labs have this problem and most likely even their strains have mutated over time.

I agree. If it is something that is not readily available you would want to take different procedures. I am curious on thoughts about freezing in glycerin. I do not believe this would have the same sporulation effect you are talking about.

Glycerine stocks and -80 freezers are what all of our yeast labs use for long term preservation. All metobolic function ceases at this temperature and they yeast don't sporulate.

 

[JohnMc]

You got it...I looked it up and that's what wikipedia said, so it must be right!

I guess my memory's as reliable as wikipedia

 

[JohnMc]

mating (even though if they did mate, theoretically they would be selfing,

evanos, even when selfing, there still can be segregation at any particular gene or locus if the parent was heterozygous at that locus, the progeny would segregate 1:2:1 for the combinations, where there's one homozygote of the first allele, two hets (like the parent), and one more homozygote of the second allele. Given about 6,000+ genes in yeast, that's a lot of possibilities.
Also, cross over during meiosis (making the spores) can create new alleles and new nearest neighbor effects.
Stuff like that.

 

[Randar]

I agree with this. Unless you have a way to maintain the original strain, no matter what technique you use to propagate and store, you will have mutation. You have mutation everything the yeast reproduces, regardless of the storage method. Unless you have a crazy science lab in your house. And by crazy, I mean you could start a lab business out of your house.

Most (by most I mean an astronomical number) or mutations will not propagate (programmed cell death, aka apoptosis) or the mutation in some small area of the genetic sequence does not manifest in an observable manner.

That being said, I still contend you can freeze and store in a typical chest freezer given the appropriate precaution against auto-defrost or short power outages (biomed type styrofoam and some gel ice packs). Storing a single effective glycerol-based yeast sample in this manner and building a starter from some scrapings should last you countless batches and provide reliable results if basic sanitation is maintained. In this manner, you are still growing yeast from the original culture/generation and the likelihood of mutation or compounded mutations impacting the finished product will be reduced vs a method employing successive generations.

Having a -80 on constant back-up power is the best, no doubt. But I think for the practical homebrewer, you can still store a vast yeast library in a chest freezer given good practices.

 

[spareparts]

Having a -80 on constant back-up power is the best, no doubt. But I think for the practical homebrewer, you can still store a vast yeast library in a chest freezer given good practices.

My yeast people tell me that even -20 would be fine. The yeast may continue to metabolize at a very slow rate so you may not get indefinate storage potential like you would at -80 but it should be ok. One professor said that if this were his only option in the lab he would reculture annually just to be safe.

 

[HighGravity]

About the distilled water method. Why wouldn't you just use regular tap water like I use in every batch I've ever made? If distilled doesn't have the nutrients it would make sense to use natural water.

 

[JohnMc]

But I think for the practical homebrewer, you can still store a vast yeast library in a chest freezer given good practices.

+1; Sounds right. I'd venture that would also hold for slant storage under mineral oil as well.

 

[chicagobrew]

I'm disappointed to hear that sterile water causes sporulation, but it makes sense. I had hoped to use this method to store archival stocks.

One method that I haven't seen kicked around the forum is using a 10% sucrose solution like is described in the book First Steps in Yeast Culture. Does anyone have an opinion on this method? Would this also cause sporulation of the sample? I guess I'm just trying to find a method that would allow for long term storage of yeast without the use of a freezer. I have only a tiny freezer in my apartment and its jam packed with food. SWMBO puts up with a lot with the brewing, but she's going to draw the line if I start taking up the freezer too.

 

[Randar]

My yeast people tell me that even -20 would be fine. The yeast may continue to metabolize at a very slow rate so you may not get indefinate storage potential like you would at -80 but it should be ok. One professor said that if this were his only option in the lab he would reculture annually just to be safe.

Yes, this is true. Longevity will be the only potential issue with using the -20. I think re-culturing could be done every 3-5 years and still be plenty safe. (I suppose you could do it if you started noticing any serious lags in start time on your small initial step-up?). The -80 is indefinite, as you noted.

 

[broadbill]

I'm disappointed to hear that sterile water causes sporulation, but it makes sense. I had hoped to use this method to store archival stocks.

What's your definition of "archival"?
The issue of sporulation aside, storing yeast under water isn't a preferred method of storing archival stocks anyway. If by archival you are talking about months, you might be OK...years? probably not.

Look into yeast slants if you are interested in a long-term storage solution that can go in your refrigerator. More knowledgable people than myself can probably help you out.

 

[CaptainCoconut]

About the distilled water method. Why wouldn't you just use regular tap water like I use in every batch I've ever made? If distilled doesn't have the nutrients it would make sense to use natural water.

According to Palmer, it's the lack of nutrients that causes them to go into hibernation mode, which is what you want.

 

[chicagobrew]

By archival, I did mean years. I was hoping that sterile water would work for that. According to some of the research out there (Hansen, I believe) the viability remains for that long, but perhaps we're risking mutation/drift by allowing sporulation.

I do use slants for regularly used strains, but I was looking for a longer-term storage option for those strains that are only used occasionally and a back up for those on slants. I've got about 15 strains in my bank right now and I don't look forward to having to reslant every few months. I'm happy to do it for stuff that gets used every few weeks, but some of my belgians only get used once a year or less.

 

[JohnMc]

chicagobrew, I'd think about slants plus sterile mineral oil. You'll need the fridge, not the freezer for best results.