// Twitter Cards // Prexisting Head The Biologist Is In: genetics
Showing posts with label genetics. Show all posts
Showing posts with label genetics. Show all posts

Thursday, December 5, 2019

Tomatillo Breeding (1/n)

Tomatillos are a wonderful vegetable plant to grow. There are several distinct varieties available, but nowhere near the numbers we see for tomatoes, peppers, or other crops. What's the difference?

Tomatillos are almost exclusively out-breeders. You need two or more plants growing in an area to get good production of fruit. As a result, every plant is a new hybrid and a population will maintain a high degree of genetic diversity. This also makes it difficult for different varieties to be grown in the same area, as they will generally cross and meld into one diverse population.


A few years back I started an experiment with breeding tomatillos. I grew one plant of a variety with small purple fruit next to one plant of a variety with large green fruit. I had saved seeds from a CSA and the local grocer, so I don't have any specific variety names to give you. (If you want to replicate the experiment, the purple variety was similar to: https://www.edenbrothers.com/store/purple-tomatillo-seeds.html; the green to: https://www.edenbrothers.com/store/rio-grande-verde-tomatillo-seeds.html.)

Four tomatillo fruit, from left to right. 1) Medium purple. 2) Small purple. 3) Large green. 4) Medium purple.
#1. Medium purple fruit.
#2. Small purple fruit.
#3. Large green fruit, with purple dots.
#4. Medium purple fruit.
Because the plants are such extreme out-crossers, every seed that year was expected to be a hybrid between the two different varieties. The next year I grew four plants, from seeds I saved from the purple plant. Each plant grew distinct fruit. (1-4, left to right in photo at right.) This diversity tells us that both parental varieties were highly heterogeneous, so the specifics of each hybrid plant depended on exactly which allele they inherited from each parent. As none of my neighbors were growing tomatillos, we can be pretty sure each one was pollinated by the other three.


Two large green tomatillo fruit at right. Six small pale purple tomatillo fruit at left.
F2s from F1#3.

The next year I planted seeds I had saved from plant #3. I grew 11 plants, but only 5 produced any fruit. The plants looked like they'd been exposed to an herbicide from the commercial garden soil I had added to the garden at the start of the season (Herbicide carryover). All the fruit were green, with some later developing some purple pigment as they ripened off the plant.

Ten bowls filled with tomatillo fruit. Contents of each bowl are different sizes and/or shades of green and purple.
F2s from F1#4.
This year I planted seeds I had saved from plant #4. I grew 12 plants and all produced fruit. These showed a much wider range of pigment levels, including a pair of plants with visible purple pigment and large fruit.

One plant had a trait I didn't like at all. The fruit from spoiled very rapidly after picking. (Previous year's fruit stored for months.) That plant was one of two in an isolated garden, so I immediately culled all of the fruit from both plants. I didn't want to risk the genetics associated with spoilage turning up in the garden again next year.

Overhead view of orange plastic bowl filled with large tomatillos. The fruit are combinations of green and dark purple. One fruit at center is mostly green with three purple stripes starting at the bottom.
One plant had fruit I really liked. The fruit were large and developed purple pigment, the traits I have been trying to combine in one plant. I wasn't expecting the fruit to develop stripes as they were maturing, however. These fruit are not lasting as long as I'd like, but the other good traits means I'll be saving seeds from them anyhow.
Overhead view of green plastic bowl filled with medium tomatillos. The fruit are dark purple, with the most ripe looking black..
A couple other plants produced intensely dark purple fruit, appearing ink-black. This is the color I've been looking for, but the fruit aren't as large as I want. I'll save seeds from these as well.

Because the plants are out-crossers, I know they will have been pollinated by the others in the garden. Even though these two have trait combinations I really like, it will be unlikely to find offspring with the same traits because of all the other traits in the garden.

I've tried to diagram the overall history of the project so far. (I didn't have any photos of the original varieties, so they get cartoon representations.)

At top are a small dark purple and large green circle, representing the original varieties I crossed. From the dark circle, a black line goes down to a second row consisting of four tomatillo fruit pictures. (From left to right: medium purple, small purple, large green, & medium purple.) Black lines are drawn from beneath the right two fruit downwards to photos. Left line goes to a photo of 5 bowls of green fruit, with a photo of pale purple fruit to the left. The right line goes to a photo of 10 bowls of fruit with varying colors of purple and green.
Tomatillo project history so far.

About this point I started thinking about how I might get around the issues caused by the potential for genes from every plant in a garden to turn up in the next generation. I don't want to have to cull everything from a garden when something strongly negative turns up in the population. Right now I only have two isolated garden spaces, so that strategy can only go so far.

For my solution, come back in a week for part 2!


References:
Posted by at 1 comment:
Labels: , ,

Thursday, April 5, 2018

The Naming of Things

If you've been following me here for a bit, you've probably noticed I'm interested in plant breeding (especially garden veggies). My main goals are to have healthy plants that grow and produce well for me with minimal inputs in my short-season climate. The measure of, "tasty" I go by is what tastes good to me and my family, with what other people consider tasty (when I occasionally do taste-tests) held to a lesser significance.

Two large cherry sized, blocky, white tomatoes. They're sitting on a notebook with a sketched map of the garden, showing where all the plants are and which plants were grown from the same batches of seed. There is a blue pen pointing at the specific plant which produced the fruit.
From 2017, with garden notes.
I've been working with tomatoes for several years and have developed some more fine-tuned ideas about what I want the plants to become. One of my lines, seen at right, is approaching stability. That is to say, most plants from one year to the next produce very similar fruit. The fruit are blocky, large-cherry sized, white (well, paler than yellow) in color, and have a very thick outer fruit-wall (not the skin). They've tested well with people in and outside my immediate family, so I've been thinking about the possibility of distributing their seed in the future.

A few dozen of the large white cherry tomatoes sitting on a white plastic cutting board.
From 2016.
In my personal notes, I've been using the rather uncreative name of, "Abbey White" for these tomatoes. It is sufficiently descriptive to let me know what I'm talking about in my notes, but it isn't a name I expect to attach to the variety when/if I start distributing it. I could easily adjust it to, "Abbey's White", but I'm not sure I want to go with that either.

In the forground is a ceramic bowl filled with diced white tomatoes. In the background is a large wooden cutting board covered in white, yellow, and orange tomatoes (as well as a few green tomatilloes).
From 2016.
"Wait. Tomatoes are red, right!?" A white tomato might seem kinda unusual, but it's just one of a very wide spectrum of colors that tomatoes can be found in. (Check out these companies I have no affiliation with: Artisan Seeds, Baker Creek Heriloom Seeds, TomatoEden, and SeedSavers Exchange. There's so much more diversity in color and taste available if you're willing to grow tomatoes from seed.) My tomatoes tend to be any color but red. Red fruit that have turned up in my garden have tended to have a taste I didn't favor, so over a few years I stopped growing as many red tomatoes. I expect I'll need to bring in some new genetics before I can grow red tomatoes that will taste good to me.


While I was thinking about how to go about naming this variety (and others in the future), I came across twitter user @JanelleCShane. She's been playing with Recurrent Neural Networks (basically a type of AI (specifically a type of machine learning)) trained on diverse datasets, like fruit names (or knitting patterns (or Irish melodies)), so I tweeted:

(I only later noticed my garbled grammar.) I was somewhat surprised when she responded back, asking if I had a list of tomato variety names she could train her AI with. I didn't, but I was pretty sure I could pull one together pretty quickly from online resources. After some looking, I found several sources ([1], [2], [3], [4], [5], [6]) with large lists of tomato variety names. To avoid spending too much time gathering the names, I wrote web scrapers to process each source and output text files with lists of names. In total, across the six sources, I collected 11,719 distinct tomato variety name strings. Some may represent extinct varieties. Some are in other languages. Some are numerical codes. There's also capitalization and spelling variations. I threw them all into a file that Janelle could use to train her AI.

Have a look at her blog post on the tomato name trained AI at: http://aiweirdness.com/post/172622965862/tomatonames


So. What did the trained AI come up with? Well, at first the AI got overly fascinated with the numerical code names in the training dataset. It produced lots of new "names" that would be quite not useful for naming a new variety. Janelle stripped out most of the code names from the list and trained the AI again.

This time there were some really good results, some really wrong results, and all sorts of weirdness in between. I've highlighted some of my favorites from each category.

The Good,the Weird,and the Wrong.
  • Floranta
  • Sweet Lightning
  • Speckled Boy
  • Flavelle
  • Market Days
  • Fancy Bell
  • Pinkery Plum
  • Mountain Gem
  • Garden Sunrise
  • Honey Basket
  • Cold Brandy
  • Sun Heart
  • Flaminga
  • Sunberry
  • Special Baby
  • Golden Pow
  • Birdabee
  • Sandwoot
  • Bear Plum
  • The Bango
  • Grannywine
  • Sun Burger
  • Bungersine
  • First No.4
  • Smoll Pineapple
  • The Ball
  • Golden Cherry Striped Rock
  • Eggs
  • Old German Baby
  • Frankster Black
  • Bumbertime
  • Adoly Pepp Of The Wonder
  • Cherry, End Students
  • Small Of The Elf
  • Champ German Ponder
  • Pearly Pemper
  • Green Zebra Pleaser
  • Flute First
  • Speckled Garfech
  • Green Dork
  • Cluster Gall
  • Shirve’s Gigant Bullburk
  • Giant Ballsteak
  • Black Crape
  • Brandywine, True Grub
  • Caraball
  • Ranny Blue Ribber
  • Roma Wasting Star
  • Scar Giant
  • Bug Beauty
  • Banana Placente
  • Bananana
  • Stoner
  • Speckled Bake
  • Ruck
  • Green Boor
  • Wonder Bagg
  • Sun Bung
  • Bellende
  • Shart Delight
  • Solad Piss


There were also a collection that would fit perfectly among the real tomato names, though they'd be kinda strange in other contexts.
  • Matt's Sandwich
  • Indigo Tree
  • Striped Hollow Potato Leaf
  • Lelly's Yellow Stuffers
  • Terra Pink Strain
  • Greek Boar
  • Ton's Oxheart
  • Babla's German Paste
  • Mortgage Lifter, Honey Blues

I really like when the AI tried to name a tomato after a person. It didn't have enough examples for real human names, but it gave it a good solid try.
  • Matt's Sandwich
  • Lelly's Yellow Stuffers
  • Ton's Oxheart
  • Babla's German Paste
  • Shirve’s Gigant Bullburk

Amusingly, the AI came up with an existing name that wasn't in the training dataset. "Sunberry" is the name of another fruit. It's a close relative of the tomato, so I think I'll call that a positive score for the AI.


Do any of these names fit my tomato? I'm not sure. I do rather like, "Flavelle" and, "Mountain Gem". I'll probably have to let the ideas ferment a while before I come to a decision.

I have recently seen a tomato that the name, "Speckled Garfech" would be perfect for. It came out of someone else's breeding program, so I won't share a photo. Imagine a yellow/orange striped tomato covered in green spots.
Two photos combined. The top half is a photo of a large yellow ceramic bowl filled with small cherry tomatoes. The cherry tomatoes area a mix of white and pale orange with a pink blush on one end. The bottom half is a photo of a closeup of a single larger tomato that is white with pale dark stripes. There are smaller red tomatoes and other items in the background.
From 2017.

I've got a couple more tomato lines that I'd like to stabilize (photographs at right). The upper photo shows a mix of small, very sweet cherries in pale-yellow/white with a pink blush on the bottom end of some. I'll be growing seeds from the ones with the blush. I expect the same phenotype will turn up next year, but I'm also sure there are lots of recessive alleles still hiding in them (for larger fruit, other tastes, and not having the blush).

The lower photo is of a larger, meaty white with pale stripes. This one is a bit further along already thanks to some lucky genetics, even though this phenotype only appeared in the last year. The fruit color, size, and shape are all due to recessive alleles, so those traits should already be stable. The stripes, flavor, and plant growth details probably won't be stable yet. I'll be growing several seeds from this fruit this year to find out.


References:
Posted by at No comments:
Labels: , , ,

Tuesday, March 20, 2018

Genetics of Male-Sterile Plants

Male sterile plants are an incredibly important piece of classical biotechnology. (To be clear, they're not the result of "genetic engineering".) They allow the efficient production of hybrid varieties, which dominate the corn, rice, sunflowers, etc. markets because of their high productivity (due to heterosis, hybrid vigor) and consistency (due to genetic uniformity).

Without male-sterile genetics a seed producer has to prevent pollen from one parent from being transferred to the other parent, somehow. This is time-consuming and arduous work (like detasseling corn), or was simply impossible (like with wheat). With male-sterile genetics there is no pollen to worry about in one parent, so there is no need for intensive efforts to prevent pollen transfer. All a seed producer has to do is grow the male-sterile plant inter-cropped with the intended pollen-donor, then collect seeds only from the male-sterile plant. Every seed will be a hybrid. It's as simple as that!


Most male-sterile mutations can be found in cytoplasmic DNA. The mutations can be found sporadically or generated by various experimental methods. With cytoplasmic-male-sterile mutants, all progeny of the plant will also be male-sterile. Once they have been found, they can be introduced into any variety (with some effort) by traditional breeding methods.

At the top are two circles. Yellow at left, with a female symbol beside it; pink at right, with a male & female symbol beside it. Immediately below them, halfway betwen, is another circle representing a hybrid of the above circles. This one is half yellow and half pink, to illustrate the genetic contribution from the parents in the top row. There is only a female symbol beside this circle. There are eleven further circles below. Each is placed horizontally halfway between the previous hybrid and the original pink circle. In each subsequent hybrid circle, the proportion of yellow (as a pie diagram slice) is reduced by half. All the subsequent hybrid circles only have the female symbol beside them. The very bottom circle is filled entirely in pink, representing a male-sterile version of the original [pink] variety.
Fig 1.
We start with a target variety (pink in diagram at left) that produces normal pollen and a source variety with a cytoplasmic-male-sterile trait (yellow in diagram at right). We cross the two varieties, with the first variety as the pollen-donor. The resulting seeds all carry the male-sterile trait, but only 50% of their genetics are like the target variety.

We cross the resulting plants back to the target variety and the new seeds will share 75% of their genetics with the target variety. If we do this backcross again, the next generation will be 87.5% identical to the target variety. (Then 93.75%, then 96.875%, then 98.4375%.) Each generation brings our male-sterile plants closer and closer (by 50% of remaining difference) to our target variety. Eventually the only difference between our target variety and the male-sterile plants is the male-sterility trait itself.

At this point, we've made a male-sterile version of our initial target variety. It can then be used in making large numbers of F1 hybrid seed. As long as the original target variety is maintained, the male-sterile version of that variety can also be maintained by continuing to cross with it.
The figure was drawn from a vague memory of a similar figure illustrating conversion of a sunflower variety into a male-sterile version. I saw the figure years ago and I think it was associated with some USDA research. I wasn't able to find it while writing this, but I'll add a note here with the citation/link if I come across it later. The original author/artist deserves the credit for the method of visualizing the illustrated concept.

(The figure also illustrates the process of introducing any single dominant trait into a target variety via recurrent back-crossing, with dominant-carrying individuals chosen at each generation. With recessive traits, it is more complicated.)


Similar to previous figure, but after every two generations two rows (2 and 3, respecitvely) of circles are added in to represent the selfing and screening for double-recessives that must be done. In total, this figure is much longer and appears much more complicated.
Fig 2.
Less commonly, a male-sterile mutation can be found in nuclear DNA. These are also called genetic-male-sterility and are harder to work with because they're usually recessive. With recessive traits (male-sterile or otherwise), you have to do test selfings every two generations in order to be sure you can re-capture the double-recessive individuals for the next back-cross generation.

In the figure at right, I have each circle labeled with their genotype with respect to the recessive male-sterile trait. ("msms" is the genotype corresponding to the male-sterile phenotype.) Each circle is filled in yellow and pink to represent the contribution from the genomes of the initial strains, as in the previous figure.

As a result of the additional complexity of maintaining and using male-sterile traits caused by nuclear mutations, very few varieties have been developed using them. (A couple are mentioned in [link].) As soon as a cytoplasmic-male-sterile trait is found or made for a species, it would become the trait of choice by seed producers.


The utility of any male-sterile trait is limited to those who are trying to produce large numbers of consistent F1 hybrid seed. These traits would be neutral or positive for home gardeners who don't save seed from year to year. (Positive because they're cheaper to produce than regular F1 seed.) Anyone who saves seed from year to year, from home-gardeners to amateur plant breeders like myself, would probably find the traits annoying and want to avoid them.

With the small number of plants in each generation that I have space to grow for most of my projects, I really don't want a few (or most (or all)) of them to be partly sterile. Fortunately, like any other negative trait, you can select against it if it does turn up in your plant breeding projects and it will soon cease to be a significant issue for you.


References:
Posted by at No comments:
Labels:

Wednesday, March 7, 2018

Potato Onion and Gene Networks

I've been growing onions in my garden for a couple years. Like most of my garden veggies, they're not exactly the typical sort. A few years back I received a large sample of "potato onion" true seed derived from work by Kelly Winterton. Potato onions are an old-fashioned perennial form of the typical garden onion (Allium cepa). The "potato" in their name is because they're grown by planting some of the previous year's crop, like as is done with potatoes.

Kelly's introduction into potato onion breeding came from a lucky break, when he planted some of his onions in the fall to see if they could overwinter in the ground at his northerly location. The next season, all of those bulbs flowered prolifically. The important thing he did was to save all those seeds an then try growing them the next year, next to his pre-existing potato onion clones. The high diversity and robust growth of his seedlings caught his attention and started a bit of a movement. (For more details of his work, read through his site.)


Onion plants growing in garden bed.
Seedling onions, next to seedling Siberian irises.
I'd read about Kelly's work, so when I had the chance to get a batch of seeds from one of his lines, I jumped at the chance. My first year working with them, I just tossed a scattering of seeds into a 4"x4" pot and let the seedlings fight amongst themselves for the rest of that year. (I wanted to select for aggressive growers.) At the end of the year, I separated the survivors and planted them in the main garden to overwinter. (I wanted to select for those that were very cold-hardy too.) In the end, I had six plants from that first batch.

Several small and narrow onion bulbs laid out in three groups. At top are narrower and whiter.
Two had lots of luxuriant leafy growth, while the other four seemed to grow a little while and then stall out. I was kinda sad most of the plants didn't seem to do anything, but I left them alone until the first frosty night. As I was pulling up the plants, I got some surprises. All four of the poorly growing plants had grown bulbs, with three of them perfectly formed (though small). The two that grew dramatically produced lots of divisions, but no bulbs.

One of the rapid-growing plants flowered twice over the season, so I was able to collect a next generation of seed. Since the plant didn't bulb up the way I wanted, I found myself with a puzzle. I had no idea if those new seeds would all grow into plants with the same growth habit and no bulbs, or if the nice bulb shape could be produced by some hidden recessive alleles. I really liked the aggressive and early growth shown by this plant, so I didn't want to discard its seeds either.


It took a while of searching before I stumbled on to some useful search queries to get what I was looking for. The first useful paper I found (Lee et al., 2013) goes into detail examining a set of six genes in A. cepa that are related to those associated with the control of flowering in the model plant Arabidopsis thaliana. These "Flowering Locus" genes are transcription factors that regulate how plants develop. The paper has a great deal of interesting information about these genes, but the parts I found most interesting were the experiments showing the interactions between the genes. In this sort of case, I like constructing an interaction network to help me understand what is going on.

Figure illustrating a model of how environment regulates flowering and bulbing. Vernalization (cold hours) increases AcFT2 which then increases flowering. Sunlight hours suppresses AcFT4, which suppresses AcFT1, which encourages bulbing. A pair of dashed arrows indicating sunlight hours stimulates AcFT1 and AcFT4 suppresses bulbing.
Basic model from Lee et al.
The basic model they came up with encompasses three specific genes (AcFT1, AcFT2, & AcFT4). AcFT2 is induced by sufficient winter cold and then induces flowering. AcFT1 induces bulbing and AcFT4 inhibits AcFT1. Sufficiently long days inhibit AcFT4. All together, we get the network at right.

The two larger inferences we can make from this network are drawn dashed and in color. AcFT4 inhibits bulbing (by inhibiting AcFT1). Sufficient daylight hours stimulates AcFT1 (by inhibiting AcFT4) and thus promote bulbing.


There are a couple more interactions in the biology, so we'll add them. When the flowering pathway is activated in typical onions, the bulbing pathway is suppressed. We'll represent this as a negative influence from AcFT2 to AcFT1, though logically the inhibition could manifest further along the bulbing pathway. The Lee et al. paper doesn't mention this, but I feel this is justifiable from my experience growing onions through to flowering. An interesting point mentioned in the paper, but not discussed in detail was that AcFT4 over-expression plants showed no senescence of leaves in the fall (in addition to no bulbing), instead growing vegetatively until being stopped by winter.

A more complicated network linking cold and sunlight hours to flowering, bulbing, and leaf senescence. Negative arrows between four genes (AtFLC, AcFT2, AcFT1, and AcFT4).
Expanded model for onion.
The Lee et al. paper describes how vernalization regulates flowering in a few other species. They don't examine in detail how it is happens in onions, but their review of how it is regulated in Arabidopsis thaliana gives us a good model for how it might work. The variations in the system in different species does highlight how transcription factor networks can easily be rewired to impact development.

The model doesn't clearly indicate the default activities of the genes. AcFT1, AcFT2, AcFT4, and AtFLC are all active by default. Because AcFT1 and AcFT2 are negatively regulated by AcFT4 and AtFLC, they (and the flowering or bulbing downstream development) are initially inactive.


The authors in Lee et al. describe how flowering is regulated in a few other model plants for comparison. In short, the same genes are used, but they the comparable interaction network between them has different links. Though the genes are highly conserved, how they work together to drive development is not. The upshot of this is that studies of these genes in other plants is of limited utility to understanding the onions that led me down this path.
Previous figure with three large red "X"s indicating parts of the network that are nonfunctional.
Mutations to expanded model for onion.

Even in onions there is evidence for significant diversity in how this regulatory network is put together. In Lee et al., the authors hypothesize that leaks may have an overactive FT4 homolog (shown at left as "X1" on the interaction between daylight hours and AcFT4), resulting in the lack of bulbing and leaf senescence seen in the plant.

Some onions have different vernalization requirements to start flowering. An onion could be entirely resistant to cold as an influence in its blooming, as in "X2" of this figure. Blooming in these onions would be triggered by other influences not described in my figures, such as plant size or age.

The third mutation I have in this figure ("X3") breaks the interaction between AcFT2 and AcFT1. This would prevent flowering from interfering with bulb formation and is what seems to be going on in potato onions. When a potato onion blooms, it will form a bulb from a different growth point that is almost as large as if it hadn't bloomed. This is much different than for regular onions, where flowering results in a tiny inedible bulb.


Though there are as of yet no genetic studies illustrating this hypothetical mutation in potato onions, it would be relatively easy to undertake. Potato onions and regular onions are the same species and easily cross (if both are flowering at the same time). Sequencing a large number of F2 progeny would help make a connection between variations in genetic sequence and the phenotype of flowering inhibition of bulbing. It might even be faster to sequentially modify one onion type with mutations to match the FT genes (and surrounding regulatory regions) of the other onion type.

The second technique would at the very least be helpful in validating any findings from the first technique, since what I have indicated as a single interaction could actually involve several other genes. The consequence of this could be that the two FT genes might show now sequence differences at all between the two onion types.



What inspired me to start digging into the biology of bulbing/flowing was the hope that I could make some predictions about the genetics of the potato onions I'm growing.

My robustly growing, but non-bulbing, potato onion seedlings appear to mimic what would be expected if the "X2" mutation described above was involved. I collected seeds from the best of these plants in hopes that they might contain a recessive bulbing trait that would appear in segregations in the next generation. Unfortunately, I still don't know. The phenotype could be due to either a dominant or recessive mutation. I'll just have to grow out as many of these seeds as I have room for and find out. Answering this question will take another two years, so stay tuned.

I'll also be growing a large number of seeds from the batch I originally received. I wasn't expecting so much diversity to appear in them, so now I'm really interested in what other trait combinations will turn up.


References:
Posted by at No comments:
Labels: ,

Monday, February 5, 2018

Speculative Biology: 3-Way Reproduction

The other day I found myself thinking about what would be the fundamental biological characteristics of a species having a system which depended on three individuals, instead of the two or one we're used to, for each reproduction event. (This is a distinctly different concept from a species having multiple sexes or genders. See references at end for examples of these.)

To simplify the discussion, I'm going to start with a big assumption that the hypothetical organism only differs from what we see here in that it has a 3-way reproduction system. It is carbon based, it has DNA organized into chromosomes, etc. Breaching that huge assumption would introduce far more variables into the discussion, when I'm only interested in the basics of sexual reproduction for this discussion.


There are different ways for Earth biology to control the sex of individuals. Sex in some species is driven by genetic differences (mammals and birds). In some it is driven by the number of chromosomes (bees and wasps). In some it is driven by temperature differences (reptiles and amphibians). In others the sex changes with age or social situations (some fishes). In large groups, there are almost always exceptions to the general patterns.

Almost all of these cases involve some chromosomes being contributed to an offspring by both a female and a male. Haplodiploidy in bees/wasps is an interesting exception. (Males grow from unfertilized eggs, while females grow from fertilized eggs.)


For our 3-way reproduction discussion, lets start by assuming each of the three individuals contributes chromosomes equally to each offspring. (Later we'll examine a more complicated case.)

We can easily enough abstract the concept of a Punnett square into higher dimensions, though it does get difficult to simply convey the results in a 2D format. In the regular version, the possibilities for a single chromosome contributed from each parent are aligned along the top and side edge of the table. For a 3D version we'll do the same, but split the contributions from the third parent into three sub-tables (left to right) with the third parent contribution at the upper-left corner of each. (I've added some color highlights to help visualize the contributing parent for each chromosome as well as the sexes of the potential offspring.)

Punnett square (and "cube") for 2-way and 3-way crosses.


The first observation that stands out from this is the difference in predicted sex ratios of the offspring. In our 2-way system, the calculations implies a 1:1 ratio. In the 3-way system, the calculations implies a 1:2.5:1 ratio between the three sexes of offspring.


Next lets discuss something analogous to the haplodiploidy of bees/wasps, where males only contribute chromosomes to their daughters.

Punnett square (and "cube") for 2-way and 3-way crosses, with haplodiploidy.

This shows the same possibilities for sex ratios of offspring. Since we're using bees as a model here, it's a good time to introduce the idea that a creature doesn't have to produce offspring at the ratios suggested by such simple calculations. Bees produce very few males, and only when needed for fertilization of new queens. Similarly, a hypothetical 3-way reproducing species could easily adjust the sex ratio of its offspring to be different from what the above calculations suggest.

An abstraction from Fisher's Principle (http://the-biologist-is-in.blogspot.com/2015/12/evolutionary-battle-of-sexes.html) suggests most species would evolve towards a 1:1:1 ratio between the three sexes. Cases where this wasn't the case would be interesting.


I imagine one sex evolving into an approximation of female (with a large immobile gamete), while the other two sexes evolving into an approximation of male (with smaller, more mobile gametes). It gets much more difficult to make predictions beyond this point, though a couple examples inspired from fiction and biology come to mind.

Maybe the two male-equivalents would actively court each other and then seek out the a female-equivalent together as a pair. This seems to be the pattern described for the fictional Pierson's Puppeteers (though their sexual biology is rarely detailed in the author's stories about them).

Maybe the male-equivalents would independently seek out the female-equivalent. I imagine something similar to the Deep-sea Anglerfish, with smaller male-equivalent individuals fusing to a larger female-equivalent and waiting for the opportunity to contribute to offspring when all three sexes have joined the party.


In the grand scheme of things, I expect biological systems requiring three individuals for reproduction would be rare in the cosmos. At an early evolutionary stage, any organisms which only required two partners to reproduce would probably out-compete those requiring three partners simply because it would be easier to arrange appropriate matings. This isn't to say it wouldn't happen, since all sorts of strange things happen in biology.

The Red Queen hypothesis (http://the-biologist-is-in.blogspot.com/2014/04/oxalis-and-red-queen.html) suggests why larger species don't simply have one sex. Yet, we do see this from time to time.


References
Posted by at 1 comment:
Labels: ,

Tuesday, October 31, 2017

Sex Chromosomes of the Triturus Newt

Salamanders and newts are an interesting group of animals. They're amphibians, like the frogs and toads you're probably familiar with, but they have an elongated body form that looks more like a lizard. They're generally rather small and tend to live in places where you don't, so you probably won't see one unless you go a bit out of your way in search of them. Both salamanders and newts start out life as a submerged egg that hatches into a swimming tadpole. As they grow up, they both generally metamorphose into a form adapted for crawling around on land (though they do prefer moist places). Salamanders typically live out the rest of their lives in this stage, while newts return to an aquatic life once they reach full maturity. Adult newts develop a flattened tail that helps them swim and then they go about their quiet lives underwater.

Figure illustrating relationships between species in genus Triturus, with two representative salamanders at right.
Fig1. Adapted from Grossen et al. and photos by @Blackmudpuppy.
Hidden within these shy critters is some really interesting biology. One group, the Triturus newts (the marbled and crested newts), have some peculiar chromosome weirdness going on that results in the death of 50% of their eggs. Evolutionarily, this is a very strange situation. You'd expect any trait that resulted in such a high rate of offspring loss would quickly disappear from a population. You definitely wouldn't expect the trait to become a permanent fixture of most species in the genus, as is observed.


Two very young juvenile newts, with dark body stripes and long extended gills.
Fig 2. Triturus babies.
Photos by @Blackmudpuppy
My initial thought was that maybe the dead eggs were fed upon by their surviving siblings. Kin selection could then explain why such an apparently "wasteful" trait would stick around. If the additional nutrition gained from eating a sibling resulted in at least a 2x increase in genetic fitness (survival and offspring), then the trait could be maintained by this mechanism.

The problem with this idea is that the newts lay their eggs individually, folding a bit of underwater leaf over them for protection from predators. Any given hatchling wouldn't be expected to find a separately stashed egg, so the dead eggs would probably be consumed by other organisms. If the eggs were laid in small clusters, the story might be different, but for now we have to abandon this hypothesis.


Fig 3. Hypothetical lethal male.
The next thought I had is about the 50% loss. That specific number implies a limited set of possible genetics patterns. The first I thought of is the way our biology (generally) uses chromosomes to determine our sex. The male gametes come in X and Y versions, while the female gametes are all X. The result is that 50% of our kids are XX and 50% are XY. (There are lots of subtleties and complications to this story, but for now I'm just going to use this simple model.) If the Triturus newts were doing basically this (without the sex-determination thing the way we do it), but one version of the sperm always resulted in dead embryos... Well, that chromosome would very quickly disappear from a population. This is another hypothesis we have to abandon.

Fig 4. Balanced lethal chr1 in Triturus.
At this point I dug up a 2012 paper by Grossen et al., which described what was going on and described a model for how it might have come to be. It turns out every one of these newts has two distinct versions of their chromosome 1 (the largest chromosome). The two versions have a region of sequence with inversions and deletions relative to the other. These differences mean the two regions don't recombine during meiosis like the comparable regions of most chromosomes. This is significant because each version of this region has a recessive lethal allele that is paired to a functional allele on the other version. (Probably some of those deletions I mentioned earlier.) If any egg is fertilized with a sperm carrying the same version of this chromosome, one of the recessive lethal traits is expressed. This results in the death of 50% of embryos, and leaves the survivors all heterozygous for chromosome 1. This is a pattern that can continue through generations.


The difficulty arises when we consider how this arrangement could come to be. If either recessive lethal allele was present without the other, then it would be selected out of the population. The chromosome version without a lethal allele would quickly dominate in the population (and we wouldn't see a 50% death rate among the babies).

The Grossen et al. paper goes into some really cool details about how sex-determination works in these newts. They have an XY sex-chromosome system kind of like ours, but they're also strongly impacted by temperature. If baby newts are raised up in water that's too warm, they'll become functionally male. If they're raised up in water that's too cold, they'll become functionally female. If the temperatures are extreme enough, they'll all grow into one sex without respect to what their sex-chromosomes look like. (This happens with lots of cold-blooded creatures, though what sex is produced at what temperature varies by species.)

In this context, they propose that the 1a/1b chromosomes that are causing so much trouble started out as two versions of an ancestral Y-chromosome. Y-chromosomes tend to collect recessive lethal mutations (deletions and such) and different lineages of Y-chromosome will end up with different mutations. If a population of ancestral Triturus newts experienced a significant cold spell, some of the chromosomally male newts would have grown up female. They could then breed with more typical males to produce offspring with two Y-chromosomes. If the two Y-chromosomes have the same mutations, the offspring would die. But if they had sufficiently different versions, they could survive. (This has been shown experimentally in a few species, as described in Haskins et al. 1970.) Grossen et al. go into some detail simulating how this initial case could lead to the chromosome dynamics now seen.


Fig 5. Model for evolution of
balanced lethal Ys.
While I was reading the Grossen et al. paper, I was thinking of a slightly different version of the model. In this version, we see a female-promoting mutation develop instead of the male-promoting one that was modeled. In the associated figures to the right, I've included Punnett squares for all the possible chromosome combinations involved in matings at each stage of the model. The color of the progeny squares represents their sex, as determined by the interaction between genetics and temperature. (Red=female; blue=male; purple=either; black=dead.)
  1. The population beings with XY males and XX females. Multiple Y lineages coexist.
  2. As the temperature drops, the offspring of all XY*XX crosses develop as female. Sooner or later a XY female of the newer generation meets up with an XY male from the previous generation. If the Y-chromosome versions are the same, every baby again develops as female because the double-Y babies die. If the Y-chromosome versions are different, a fourth of the babies will develop as YaYb males.
  3. The older males eventually die off, leaving only YaYb males. There are still XX females around from the last generation, but all of their offspring will now be XYa or XYb.
  4. The XX females eventually die off, leaving only XYa and XYb females.
  5. The temperature drops a bit further. Now YaYb embryos can develop as either male or female.
  6. Some YaYb females meet up with some YaYb males and the first clutch of eggs is laid that experiences a 50% chromosomal-induced fatality rate.
  7. The newt population has been experiencing a major catastrophe and has dropped to very small numbers. The X chromosome carrying females die out, due to either random chance or some minor benefit the YaYb females have.
Fig 6. Model for evolution of
new sex chromosomes.
At this stage in the story, there are no further X chromosomes and half of every clutch dies due to incompatible Y chromosomes. We shouldn't really call them Y chromosomes anymore, so lets call them chromosome 1s for simplicity. The population is now in a stable evolutionary configuration, but only for as long as the temperatures remain consistent.
  1. The temperature starts to rise back up and new 1a1b babies start developing as males. Some very few of the females happen to have a mutation on another chromosome (2*) that encourages female development. The offspring carrying this mutation develop female at these warmer temperatures.
  2. All the older females die off, leaving only those carrying the mutation. The temperatures are still rising and some newts carrying the mutation start developing male.
  3. Some males with the mutation meet up with females also carrying the mutation. The first newts with two copies of the mutation are born and develop female.
  4. The temperatures are still rising. Newts born with only one copy of the mutation start developing male. The only newts that develop female have two copies of the mutation.
  5. The older females die off, leaving only those with two copies of the mutation. The only new males are those born with one copy of the mutation. The older males die off, leaving only those with one copy of the mutation.
At this stage in the story, a new pair of sex chromosomes has evolved. The copy with the mutation is now an X chromosome and the copy without the mutation is now a Y chromosome. The residual chromosomes from the original sex chromosome pair are still causing 50% of babies to die in early development.


I could extend my model using a simulation-based approach similar to Grossen et al. to make a better comparison, but I don't expect I will. With complex evolutionary history like this, it might never be possible to ascertain exactly how the process unfolded. There could be many equally-probably historical scenarios that would have led to the evolution of the situation we see today. It's educational to study how these systems could potentially have evolved, even if we can't sort out the exact path they took to get where they now are.

I don't often write about published papers, but when I do I prefer to carry the discussion past where the paper ended. Discussing an alternate model is in no way an attack against the one proposed in Grossen et al.. It's an honest reflection of what I was thinking of while reading and is part of an exploration of the ideas discussed in the paper. Frankly, if their writing didn't give me any new ideas to play with, I wouldn't find it anywhere near as interesting to read. Good science answers a question. Great science answers a question and then draws you in to ask your own new questions.


This post was inspired by an interesting conversation over on Twitter. (You can follow me there as @thebiologistisn.)
The photos of the Triturus newts were loaned to me for this blog post by photographer (and newt wrangler) @Blackmudpuppy.


References:
Posted by at No comments:
Labels: , ,

Tuesday, August 1, 2017

A Cross by Any Other Name

Figure illustrating how a recessive trait appears in F1, F2, and F3 generations after a cross. In F1, the trait is hidden. In F2, a quarter of individuals show the recessive trait. In F3, 3/16 of individuals show the recessive trait.
From [link].
I've been involved in a few discussions online lately about different types of crosses that can be used in plant breeding. There has been some mild confusion about basic terms, as well as about the implications of different types of crosses. A few years ago I wrote about backcrossing. Though that post is somewhat hard for me to read, as I imagine early writings are for most authors, it has some useful information. Here I'm going to try and do a more general overview. Lets see how this little ride goes.

Some of that basic terminology and common abbreviations:
  • P : Parental. An initial variety used in a cross. Multiple parents can be numbered, like in "p1 x p2".
  • F : Filial, relating to progeny generations after an initial cross. F1 is the initial hybrid. F2 is the result of crossing two F1s. F3 is the result of crossing two F2s, etc.
  • Self Cross : Crossing the male and female parts of the same plant.
  • BC : Back cross. Crossing a filial generation back to one of the parents.
  • CC : Complex cross. A cross involving more than two parents.

P : To simplify things, we usually use highly stable varieties as initial parents in a hybridization project. This means that several generations of each parent variety have been grown out without any visible variation appearing. At the basic genomic level, this means the varieties are highly homozygous. In theoretical cases we consider the parents to be absolutely homozygous, though reality is never quite so clear-cut.

F1 : Our initial hybrid between two parents can be written out in a bit longer form like "p1 x p2", or just referred to as an F1 between the two parents. In our idealized scenario, every F1 produced by crossing the same two parents will be identical. F1 stands for "first filial generation".

If a group of F1s aren't identical, this says one or both of the parents wasn't entirely homozygous. (Or new mutations were introduced, or epigenetic effects are at play, or etc. It can get complicated). Because they're (more or less) identical, selection usually isn't very important at this stage.

Figure showing Punnet square of a dihybrid cross in peas, with each potential offspring plant indicated by a cartoon of the resulting pea-pod. A quarter of potential plants have yellow pods. Independently, a quarter of potential plants show pinched pods, with seeds visible through the constricted pod surface.
From [link].
F2 : Our second filial generation is produced by crossing two F1s together. For those plants that can self cross (like peppers and tomatoes), the F2s would generally be produced by crossing one F1 to itself. For those that can't (like tomatillos), the F2s would be produced by crossing two separate F1 siblings.

The F2 generation is where the different alleles from each parent are recombined. Almost any combination of traits from each parent can turn up in an individual among the F2s. This is where the magic happens in a plant breeding project really happens. This generation is where selection is most important.

F3...Fn : Subsequent filial generations would be produced in a similar way to the F2s. If you produced F3s by selfing an F2, each F3 will have about 50% of the heterozygosity of the F2. Selfing another generation will result in another 50% loss of heterozygosity. Continue this process for enough generations and you will have a new stable variety, with an essentially homozygous genome.

If you produced F3s by crossing random F2s, you'll keep mixing up the genetics instead of automatically losing 50% of the heterozygosity each generation. If you do this with relatively few plants, you will still be losing heterozygosity each generation, though calculating exactly how much becomes a bit complicated.

If you produced F3s by crossing specific F2s that had a trait you liked, you'll keep mixing up all the other genetics while selecting for that specific trait. You would be losing heterozygosity near the genes responsible for the trait of interest, but the rest of the genome would still be maintaining heterozygosity through generations.

BC : In basic back crossing, each subsequent generation past F1 is crossed back to one of the parents. BC1 would be diagrammed something like, "[p1 x p2] x p1" (or "F1 x p1"). For one hypothetical mutation found in the first parent, a BC1 individual would have a 50% chance of having two copies (and a 0% chance of having no copies) since it is assured of inheriting one copy from the parental strain used in the backcross.

Through each generation of back-crossing the resulting plants will lose 50% of their heterozygosity, but it will be replaced with whatever mutations are found in the parental strain. The result will end up more and more like the recurrent parent strain over the generations. If you do this randomly, you will end up with essentially a genetic clone of the recurrent parent. To get anything different, you have to persistently select for a trait that was originally only in the second parental variety. Doing this will eventually produce something almost exactly like the recurrent parent, but with the one trait that was originally in the other parent variety. (That's all detailed in the link I mentioned in the intro.)

CC : A complex cross involves three or more parental varieties. A simple case would be taking an F1 and crossing it to an independent F1, "[p1 x p2] x [p3 x p4]". In these scenarios you would get a very diverse population, just like with F2s, but the mutations contributed to the population can come from all four parent varieties.

A mutation that was found in only one of the parental strains would only be found in one copy in 25% of this mixed up population. If one of these plants was selfed, the chance of a plant being homozygous in the next generation is 6.25%.
If the plants were allowed to cross randomly, the chance of a plant being homozygous in the next generation drops to only 1.5625%. You would need to be working with very large numbers of plants to routinely recover double-recessives using this strategy. I strongly advise you not use this strategy.


References:
Posted by at No comments:
Labels: ,
Subscribe to: Posts (Atom)