Yearly Archives: 2015

Creating a new ’super fruity’ yeast strain – The best of Conan and WLP644?

In this blog post I’m briefly going to summarize how I created and improved a hybrid yeast strain between Conan (isolated from a can of Heady Topper in 2013) and WLP644. I’ve used both strains in several homebrews previously, and I’ve really liked the fruity esters they produce during fermentation. While Conan produces a really nice aroma profile, it doesn’t seem to attenuate as well as say WLP001 (most probably as a result of incomplete maltotriose use). This means that it often leaves a slight sweetness in the beer, and is a bit unpredictable. With WLP644 I’m not that sure. I’m fairly sure it uses maltotriose and ferments the beer quite dry, but this observation is only based on a single homebrew I’ve done with it. It seems to grow and ferment slightly slower than most ale strains though. So, I had the idea to generate a hybrid between the strains, in order to create the ultimate ‘super fruity’ yeast strain:

  • Lots and lots of fruity esters
  • Complete maltotriose use for high attenuation
  • Bonuses are fast fermentation and moderate flocculation

Spore-to-spore mating is the traditional method of generating yeast hybrids, but I myself am more of a fan of rare mating. Also, spore-to-spore mating couldn’t really be applied here in this case because of the poor sporulation ability of Conan. To begin, I needed to ‘tag’ the parent strains with selection markers. To do this, I selected spontaneous auxotrophs of Conan (ura-) and WLP644 (lys-) on 5-FOA and α-AA agar, respectively. This means that my Conan mutant strain isn’t able to synthesize its own uracil anymore, while the WLP644 mutant strain isn’t able to synthesize its own lysine anymore. Any hybrid between these two would inherit the functional genes from the other parent, meaning the hybrid is prototrophic again, and can again synthesize its own uracil and lysine. Hybrids can thus be selected on minimal media, which doesn’t contain uracil and lysine.


The actual hybridization process is easy, all I have to do is mix cultures of both parent strains in rich media (YP-Maltose in my case), incubate for 3 days, pellet, wash and starve the yeast, and finally spread it out on minimal media agar. Any colonies appearing on the minimal media agar are most likely hybrids. The other option is that the colonies are the parent strains, which have undergone spontaneous mutations to return to being prototrophic again. In order to ensure that we in fact have hybrids, the colonies are first purified by restreaking on fresh minimal media agar, and then transferring and streaking a single colony to YPM agar. DNA is then extracted from a single colony on the YPM plate, and interdelta-PCR is performed on the DNA.

Interdelta-PCR is a technique that is usable for differentiating Saccharomyces cerevisiae strains. Below you can see the gel and the profiles that our hybrids (H1-H4) and the parent strains produce. As we can see, the hybrids contain DNA (i.e. the bands) from both parent strains, so it is confirmed that we have successfully generated a hybrid. This is especially evident in the areas I’ve marked with red asterisks on the DNA Ladder. I won’t go into more details on the mating mechanisms, but the short answer is loss of heterozygosity at the MAT locus (for more details see here). Being a result of rare-mating, this hybrid will most likely be tetraploid (assuming that both parent strains are diploid), and contain approximately the whole genome of both parent strains (as can be seen from the gel below, hybrid H2 has already lost some DNA).


To stabilize the hybrids and also generate some potentially interesting phenotypes, I utilized meiotic recombination and segregation! During ascospore formation in yeast, meiosis and chromosomal crossover takes place, during which genes are shuffled between pairs of chromosomes. First we have to sporulate our hybrid (this can be done on 1% potassium acetate), then treat the forming ascospores with a lytic enzyme (e.g. Zymolyase), and finally dissect the ascospores with a micromanipulator. I choose to do this to hybrids H1 and H2. Both strains sporulated quite poorly and the spores had low viabilities as well (34% for H1 and 8% for H2). I selected the fastest growing colonies for further characterization, and this was four segregants from hybrid H1: H1/A4, H1/C3, H1/C4, H1/H1. The characterization was done in some small-scale (35 ml) wort fermentations. This was done in order to confirm that they still are able to grow and ferment well in wort and that they produce a lot of fruity esters.


tetrad_dissectionThe four segregants that were chosen for further characterization

The small-scale fermentations were carried out in 15 °P wort (specific gravity: 1.060), because I thought this would be representative of a typical IPA wort. 35 ml of wort was added to pre-weighed and airlock-capped plastic tubes (50 ml), after which 10 million cells / ml of wort of each yeast strain was added to start the fermentations. Fermentations were carried out at 18 °C (in retrospect, this might have been a little low), and they were monitored daily through mass loss. The alcohol content of the final beer was measured with an Anton Paar DMA5000M + Alcolyzer. Our HS-GC is in heavy use at the moment in other projects, so I wasn’t able to measure the individual ester concentrations. I did sniff the beers though to get a general idea of whether the hybrids are actually fruitier than the parent strains. The fermentations were done in wort that had only been hopped in the beginning of the boil, to make sure that the majority of the beer aroma was yeast- and not hop-derived.

minifermsThe small-scale fermentation vessels

fermentationFermentation progress

abv_ph ABV% and pH of the beers

As you can see, there was considerable differences in fermentation rate between the strains. Of the parent strains, Conan started fast, but ended slow, while WLP644 was the other way around with a slow start and faster finish. In early stages of fermentation, the H1 hybrid was also doing well, but in the end it was two of the meiotic segregants, H1/A4 and H1/C4, that reached the lowest final gravities after two weeks of fermentation (1.013 and 1.009 respectively). The Conan and WLP644 parent strains reached final gravities of 1.015 and 1.017 (this would probably have dropped slightly with a couple of more days of fermentation). There was considerable variation in pH as well, as the lowest pH values were observed in the beers fermented with H1/A4 (4.39), while the highest pH values were observed in the beers fermented with H2, H1/C4 and H1/H1 (4.64, 4.61 and 4.69). The aromas of the beers were very similar, all featuring a similar fruity tone that reminded me of various stone fruits (peach and apricot). While there were some differences in intensity between the beers, I’m not sure how much I would trust just my nose. In my opinion the strongest fruity aroma was found in the beers fermented with Conan, F1/A4 and F1/H1. The weakest aroma was found in the fastest fermenting beer, i.e. F1/C4. However, as I mentioned the actual ester concentrations would need to be measured to actually draw any conclusions. All aromas were ‘clean’, with no signs of any phenolics (both parents are POF-). The WLP644 beer had a slight sulfuric note to it though. Another thing I observed during these fermentations, was that WLP644 flocculates poorly, while Conan and all the hybrids flocculated quite well.

The next step will be to actually brew and taste some beers brewed with these yeast strains, in order to see if there actually is any real world difference. For this I was planning on brewing up a 25-liter batch of APA wort (OG around 1.050, IBUs around 50, whirlpool hops, but no dry hopping), which I would then split into 5 fermentation vessels. To these I would then pitch: Conan, WLP644, H1, H1/A4 and H1/C4. I’ll return when it is time for the brewday and ultimately the tasting notes.

References and additional reading for those interested:

Krogerus K, Magalhães F, Vidgren V, Gibson B. 2015. New lager yeast strains generated by interspecific hybridization. Journal of Industrial Microbiology & Biotechnology 42: 769-778. DOI: 10.1007/s10295-015-1597-6

Steensels J, Snoek T, Meersman E, Nicolino M, Voordeckers K, Verstrepen K. 2014. Improving industrial yeast strains: exploiting natural and artificial diversity. FEMS Microbiology Reviews 38: 947-995. DOI: 10.1111/1574-6976.12073

Tasting Impressions: Schwarzbock from 2013

I found a couple of bottles of homebrewed Schwarzbock from 2013, and I decided to see how it had developed during two years in bottle. This one ended up at an ABV of 6.8% and with a final gravity of 1.020, so a bit lighter than what was originally planned. I remember this one being very tasty when fresh, so let’s see how it tastes now!schwarzbock

The beer pours black and with a fluffy tan-colored head (it looks a lot lighter in the picture below). If you hold the glass against the light, you see that the beer actually has a dark ruby color (and is crystal clear). As the head collapses, it leaves patches of lacing along the glass. A nice appearance! The aroma is really nice as well, and it features a good combination of chocolate-like roasted tones, together with dark fruits and caramel. There is a slight alcohol note in the aroma as well, even though this is only 6.8%. The flavor is mostly roasty as well, with tones of dark chocolate and ash dominating. Behind this roast, there are tones of malt bread, biscuits and caramel that balance out the flavor. There are not many hop-derived flavors, but that was expected based on the recipe and the beer age. The finish has a moderately light bitterness though. The finish is quite dry as well, which adds to the perception of a relatively light body. This is quite a difference to the sweet and strong imperial stouts. The moderate carbonation level gives the beer some more mouthfeel though. It is quite easy to drink, and it has aged very well! Overall, a very nice beer that I would definitely brew again!

Homebrew: Fresh Hop Ale

I collected this year’s hop harvest two weeks ago, and today we finally brewed a beer with them. This is the first time I’m using homegrown hops, so am really looking forward to tasting the final beer! Technically, this wasn’t actually a fresh hop ale, since I dried the hop cones before using them – but I’ll call this a fresh hop ale anyways. Since I’m not sure about the alpha acid content of these hops, we decided to use them only as flame-out hops, and instead use some Herkules at the beginning of the boil. This way we will hopefully extract the maximum aroma out of them as well. The homegrown hops weren’t very aromatic, so I’m expecting mostly grassy flavors and less of the typical citrus and pine resin. But hopefully I’ll be positively surprised! The post-boil wort had a slight perfume-like tone, which might have been hop-derived. For the malt bill, we went with a very simple 90% Maris Otter and 10% Carapils to an OG of around 1.050. This should let the hops shine (if they do). For the yeast, we used a really fruity ale strain that I’ve developed at work (more about that in a future post). I’m hoping it will compliment the hops, and make for a refreshing and crisp fresh hop ale! We will see in a couple of weeks!


This was the first time we brewed at our ‘new’ basement brewery (we moved there 1.5 years ago – yes, we’ve been extremely slow with the renovation), and I’m happy to say that everything went really smoothly. We were done in 5 hours and 30 minutes, reached almost 70% brewhouse efficiency, and the ventilation system (a really powerful inline exhaust fan) worked amazingly well. Looking forward to brewing a bit more regularly from now on!

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Homebrew: Wedding Dunkel

Time for another wedding beer. This time not for my own wedding, but for two of my friends’ wedding. The groom requested one dark and malty beer (the wedding is in January, so that would fit with the cold weather) and one pale and fresh beer. I’ve noticed that the most popular beers during events, where the majority of the people are not ‘beer nerds’, are balanced and easy-to-drink beers. American Pale Ales with a relatively mild bitterness seem to be especially popular, so I thought I’d go with a remake of my own Wedding APA for the pale and fresh beer. For the dark and malty beer, I decided to go with a dark lager.

I haven’t brewed many dark lagers before, but decided to go for a malt base dominated by Maris Otter and Munich malt. To this I added hints of Chocolate and Dark Crystal malt to give some color and flavor. I kept it simple with the hops, and decided to go with Tettnang at moderate amounts to a fairly low IBU. For the yeast I chose to use W-34/70, which is my favorite homebrew lager yeast because of the ease-of-use (just sprinkled two packs on top of the 21 litres of wort) and the clean and crisp flavor profile. The brewday went extremely smoothly, and 21 liters of wort are now fermenting strongly at 12C. The aroma coming from the airlock is really nice, which is always a positive sign! The wedding is in about three months, which will mean I will have some time to lager the beer once it has finished fermenting.

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Homebrew: Bavarian Hefeweizen

I’ve again been lazy and haven’t updated the blog for a while. There hasn’t been that much interesting to post though. About three weeks ago I brewed a Bavarian Hefeweizen for a friend, and today I decided to pop the first bottle as quality control. The recipe was really simple, with the malt base being 60% wheat malt and 40% pale ale malt. I hopped with Tettnanger to a modest 16 IBU, and fermented the wort with WLP380, supposedly the Schneider Weisse yeast.

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The beer pours with a hazy golden-yellow, almost grey, color. A fluffy white head is formed, but it collapses a bit too quickly for a Hefeweizen. A typical wheat beer appearance. The haze is both yeast- and protein-derived as it was quite clear before I put it in the fridge.

The aroma is typical Hefeweizen, with tones of cloves and banana. The spicy 4-vinylguaiacol dominates the aroma profile, but there are lots of fruity esters present as well. The aroma is quite one-dimensional, but I guess it fits the style quite well.

As with the aroma, the flavour profile is dominated by spicy phenols and fruity esters. The amounts of esters and higher alcohols almost go a bit too far as I get hints of solvent as well. On the other hand, it has only been three weeks since pitching, so these will probably subdue a bit with some time in the bottle. There are some malty and doughy tones hidden in the background, but I can’t detect much hop presence at all. As it should be in a Hefeweizen. The carbonation level is high and the body quite light. Refreshing and quite easy to drink.

Overall this is an okay Hefeweizen. I’m not that big fan of the style, so I have a hard time judging how successful this beer is. It is still very young, so it will probably change a bit with more time in the bottle. There are definitely a lot of esters and phenols present, as it should be, but there might even be a bit too much of them at the moment. Perhaps this could have been fermented at a slightly lower temperature?

Hop Harvest

Today I harvested the cones off the hop plants in my yard. They might not have been perfectly ripe, but we have some cold nights coming up and I didn’t want to risk them getting destroyed by frost. They were smelling really good though! I collected 400 grams of hops in total, which after drying will probably fall to around 80 grams. To dry the cones, I spread them out in three layers on a steel mesh and put a fan underneath to recirculate the air. I’m hoping to brew a beer with them in a couple of weeks!



Small Update

Just thought I’d write a short update on the recent beers I brewed for my upcoming wedding. They are all tasting really nice, which I’m happy about. Here are some short tasting notes:

Wedding Pils – Dry finish, grainy malts, and a floral hoppiness that ends with a nice bitterness.

Wedding Blond – Lots of spicy phenols and fruity esters from the yeast. Not my favorite style, but this is quite a nice Belgian-style ale.

Wedding Hybrid – Lots of hop aroma combined with fruity esters. A slight hint of spicy phenols in the flavor. APA meets Wit. I like!

Wedding APA – Citrus and floral hops mainly that end in a moderate bitterness. Some maltiness as well, and not a completely dry finish.

Wedding IPA – Lots of citrus and tropical fruits from the hops together with a huge bitterness. A really successful IPA!

Also, here is a progress picture of the ‘bar’ I’m building for the wedding. Five homebrews will be available on tap. I still need to paint it and build a drip tray, but it should be ready in a couple of days. Really looking forward to testing it!


Homebrew: Wedding IPA – American IPA

Yesterday I brewed the fifth and final batch of beer for my wedding in August (see previous posts for the Wedding Pils, Wedding Blond, Wedding Hybrid, and Wedding APA): an American IPA loaded with hops and fermented with Conan. I’ve brewed a slightly similar recipe before (see here), and while it was a nice beer, it ended up a bit too sweet with WLP007. This time I’m changing up the hop bill slightly based on what I have in the freezer, and I’ll be fermenting the wort with Conan. I’m hoping for a hoppy and bitter IPA, with loads of fruity aromas from both the hops and the yeast.

The malt bill is similar to the APA I brewed last week, and it consists of Maris Otter, Munich, CaraPils and CaraAmber.  I mashed quite low (63C) in order to get a very fermentable wort. I’ve used Conan a couple of times before and I’ve ended up with around 78% attenuation. I aimed for an original gravity just below 1.070, in order to get a beer with around 7.5% ABV. For the hops, I chose to bitter with Herkules (using up the last from a 100g bag), added some Cascade and Centennial during the boil, and at flameout I added even more Cascade and Centennial together with some Amarillo. The bitterness levels should be around 70-80 IBU, depending on how much the whirlpool hops contribute. I pitched a 1.5L starter of Conan after I had cooled the wort down to around 20C. I placed the fermenter in my fermentation fridge and set the fermentation temperature to 19C. I checked the fermenter 24 hours later, and it was fermenting violently with krausen coming out of the airlock. After a quick clean-up things were looking good again. Man it was smelling good inside the fermentation fridge!

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Homebrew: Wedding APA – American Pale Ale

Today I brewed the fourth and penultimate batch of beer for my wedding in August (see previous posts for the Wedding Pils, Wedding Blond, and Wedding Hybrid): an American Pale Ale loaded with Cascade and Centennial. I’ve brewed variations of this recipe several times before (one, two and three), so I know exactly what to expect. The resulting beer should have a hop-dominated aroma, with tones of grapefruit, pine resin and floral perfume, while the taste should feature a lightly bready maltiness together with a citrusy and floral hoppiness that ends in moderate bitterness and semi-dry finish. All in all, it should be a really easy to drink, yet still flavorful beer. The beers have been crowd favorites in the past, so am predicting that this keg will be the first to run out during the wedding.

The malt bill is simple, consisting of Maris Otter, Munich and CaraPils.  These should, together with the relatively high mash temperature (67C) and low-attenuating yeast (WLP002), yield a solid backbone to the beer despite the low ABV%. I aimed for an original gravity just below 1.050, in order to get a beer with around 5% ABV. For the hops, I chose to bitter with Herkules (since I still have some left from a 100g bag), and at flameout I added a large dose of Cascade and Centennial. The bitterness levels should be around 40 IBU, depending on how much the whirlpool hops contribute. I pitched a 1.5L starter of WLP002 after I had cooled the wort down to around 20C. I placed the fermenter in my fermentation fridge and set the fermentation temperature to 18C. I like using ‘English Ale’ yeast in my APAs and IPAs, since I think the esters they contribute go well with fruity hops. The high flocculation is also a bonus. Hopefully this one turns out as tasty as my previous brews of this recipe.

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