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Old 12-03-2011, 12:57 AM   #1
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Default A nice study - re-pitching yeast

Here is a nice study which suggests some strains resist genetic variation for hundreds of generations when there is little selective pressure. It also suggests the number of times that a commercial brewery will re-pitch the same yeast before re-propagation will depend on the susceptibility to genetic drift.



http://www.scientificsocieties.org/j...7-0420-478.pdf

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Old 12-09-2011, 01:55 AM   #2
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Unfortunately we (brewers) provide selective pressure:

- Top cropping
- Harvesting early from Primary
- Harvesting from Secondary
- Leaving under pressure (height of liquid above it)
- Alcohol environment
- Sugar environment
- Storage conditions.
- ....... and probably many others.

Each one will select a certain part of the yeast population to survive/harvest, accelerating population mutation.

Every time we handle the yeast, and every time we use yeast, we select the cells that are more capable of withstanding that environment.

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Old 12-09-2011, 03:58 AM   #3
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Quote:
Originally Posted by Calder View Post
Unfortunately we (brewers) provide selective pressure:

- Top cropping
- Harvesting early from Primary
- Harvesting from Secondary
- Leaving under pressure (height of liquid above it)
- Alcohol environment
- Sugar environment
- Storage conditions.
- ....... and probably many others.

Each one will select a certain part of the yeast population to survive/harvest, accelerating population mutation.

Every time we handle the yeast, and every time we use yeast, we select the cells that are more capable of withstanding that environment.
Right...that's why people are interested in quantifying the effects of this selection.

How many times can you re-pitch before you see genetic changes or characteristic changes from the master colony? This study suggests a lot more than I would have guessed. No detectable genetic changes after hundreds of re-pitches with deliberate selection.

I saw on a thread on this site where there was a weak consensus that "5 generations" should be the maximum number of times one should re-pitch. Based on what? This study is the first data I have seen, and it doesn't corroborate that at all.
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Old 12-09-2011, 08:10 AM   #4
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The yeast and brewery practices differ so it is just a rule of thumb, probably a very conservative one to account for slack home-brewing practices. It not only has to do with genetic drift, but inevitable contamination. I imagine the number of repitches is based on desired fermentation characteristics that each brewery determines through experience. I've read wheat beer yeast in particular have to be renewed fairly quickly. I've also read that many brewers consider a yeast to perform best at around the third repitch. More technical brewing textbooks should have more information if you are interested. I recommend Brewing Science and Practice by Dennis Briggs.

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Old 12-09-2011, 08:22 AM   #5
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Well the sheer odds of genetic mutations reaching any measurable (read: non-negligible) frequency in a huge population like active-fermenting yeast are pretty dang low.

Let's say your beer requires you pitch 200 billion yeast cells. That means that within that population size, the frequency of a new mutation would be 1/200 billion (1/400 billion if yeast are diploids... not sure though.) At frequencies that low and in stressful environments, genetic drift will eliminate those mutations super quick.

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Old 12-09-2011, 01:39 PM   #6
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Quote:
Originally Posted by orangehero View Post
The yeast and brewery practices differ so it is just a rule of thumb, probably a very conservative one to account for slack home-brewing practices. It not only has to do with genetic drift, but inevitable contamination.
I agree. I would guess the home brewer is prone to contamination without the luxury of specialized equipment to help maintain aseptic technique, but I've never tried to see how long I could go.

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I've read wheat beer yeast in particular have to be renewed fairly quickly. I've also read that many brewers consider a yeast to perform best at around the third repitch.
Based on what? That's my point, I have heard these things as well, but I haven't seen the data to back that up. Where's the science?

The commercial brewing company (Bridgeport Brewing) that participated in this study re-propagates "on an extremely infrequent basis". The data from this paper comes from the 98th generation of their house strain and is compared to the master colony.
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Old 12-09-2011, 01:59 PM   #7
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Quote:
Originally Posted by Reno_eNVy View Post
Well the sheer odds of genetic mutations reaching any measurable (read: non-negligible) frequency in a huge population like active-fermenting yeast are pretty dang low.

Let's say your beer requires you pitch 200 billion yeast cells. That means that within that population size, the frequency of a new mutation would be 1/200 billion (1/400 billion if yeast are diploids... not sure though.) At frequencies that low and in stressful environments, genetic drift will eliminate those mutations super quick.
That sounds right (400 for diploid) but the rate would be dependent on selective pressure, right?

I just looked up a paper from 2001 (Genetics 159 441-452) that reports a mutation rate of 0.0011 per cell division for Saccharomyces Cerevisiae. So if yeast will divide 2-3 times maybe during fermentation the number of cell divisions starting with 200 billion cells would be (200 + 400 + 800 = 1.4 trillion). With a rate of 0.11 % that would be 1.54 million mutations in that one fermentation. But those could lead to different flocculation charateristics, etc that could very well get filtered out.
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Old 12-09-2011, 03:58 PM   #8
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Quote:
Originally Posted by insubordinateK View Post
That sounds right (400 for diploid) but the rate would be dependent on selective pressure, right?

I just looked up a paper from 2001 (Genetics 159 441-452) that reports a mutation rate of 0.0011 per cell division for Saccharomyces Cerevisiae. So if yeast will divide 2-3 times maybe during fermentation the number of cell divisions starting with 200 billion cells would be (200 + 400 + 800 = 1.4 trillion). With a rate of 0.11 % that would be 1.54 million mutations in that one fermentation. But those could lead to different flocculation charateristics, etc that could very well get filtered out.
The initial mutation frequency I was providing of 1/2N (the 2 assuming diploid and N being population size) is assuming Neutral Theory. It's basically a null-hypothesis for an evolutionary system in which the only evolutionary force is genetic drift (i.e. random chance.) Therefore you don't really need to worry about selection.

However there is a selection-mutation equilibrium model for Neutral Theory which is (p-hat) = sqrt(μ/s) [where μ is mutation rate, s is selection coefficient, and p-hat is the equilibrium frequency of allele p]

But of course, the above information is useless unless I had a way of quantifying s for the yeasties.
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Old 12-09-2011, 04:05 PM   #9
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Also important information I forgot to add:

Even with a lot of mutations, if you look at the codons for amino acids you'll see that the genome is imbedded with a lot of redundancy. Many amino acids have multiple codons that will code for them, so most mutations are actually synonymous (i.e. base is swapped with same base or new base is a part of this redudancy.)

Let's also not forget nonsense point-mutations that code for STOP codons.... this usually triggers a cascade that results in programmed cell-death.


In both of these cases, the mutation is silent either on the individual level or the population level.

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Old 12-09-2011, 05:59 PM   #10
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Quote:
Originally Posted by Reno_eNVy View Post
Also important information I forgot to add:

Even with a lot of mutations, if you look at the codons for amino acids you'll see that the genome is imbedded with a lot of redundancy. Many amino acids have multiple codons that will code for them, so most mutations are actually synonymous (i.e. base is swapped with same base or new base is a part of this redudancy.)

Let's also not forget nonsense point-mutations that code for STOP codons.... this usually triggers a cascade that results in programmed cell-death.


In both of these cases, the mutation is silent either on the individual level or the population level.
I love it. Redundancy was incorporated into nature for home brewers!

You bring up very good points. I had a feeling you had a biological background. So I think we can put to bed the 3-generation rule or the 5-generation rule. That sh!t is baseless.
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