Found two sets of twins yesterday

Yesterday I was potting up some early germinating polyanthas and am excited there were two sets of twins that shared the same testa (seed coat). Most of the time the smaller twin has half as many chromosomes as the main seedling and has the same genetic constitution as the originating unfertilized egg. On the female side after meiosis and there is a resulting cell that has half as many chromosomes, there are some cycles of cell division that lead to 8 copies of it. One becomes the egg, two are called synergids (with one on each side of the egg), and the others are positioned elsewhere in the ovule. Sometimes a synergid next to the fertilized egg develops into an embryo without fertilization. These smaller embryos are often not as competitive as their larger twin and die in nature. It is amazing how plants have natural mechanisms to both go up and down in ploidy, so if one ploidy is more competitive, it can predominate in a region. It is interesting, for instance, how there are higher ploidy R. acicularis it seems the further North and more difficult the climate. People have hypothesized that the more forms of genes/alleles present the more resources a plant has to handle difficult climates, but the “easier” the climate, the tendency is for lower ploidy and that tends to be associated with faster growth from having less DNA to replicate. Additionally, the lower the ploidy within a genetic background I’ve noticed more branching and more flowering too.

Since I germinate seeds in baggies and transplant them up as they germinate, it is easier to find a rare set of twins. I don’t have enough room to plant the seeds in flats and this method saves space. With some tender care, the smaller twin can live and grows. Some are weak and die out, but some of them grow and develop. Typically I’ve found these smaller twins with tetraploid moms and then the smaller twin is diploid. It’ll be interesting to see if with these diploid polyanthas if the smaller twin is monoploid with just one set of chromosomes. That may make them even more difficult to have survive. If there are some monoploids maybe some of the researchers doing genomics work would find them useful. Using monoploids is how the teams in France were able to sequence and assemble the rose genome efficiently. They took pollen from diploid roses ‘Old Blush’, R. wichurana, etc. and generated monoploid callus and sequenced it. With twins like this, they typically are noticeably different in growth/blooms, etc, so are not identical twins.

I’ll attach some pictures. The first set of twins had two radicles (roots) coming from the same seed coat. There is a picture showing that and then the picture of separated twins is the same set. The other set had the smaller twin stick to the hypocotyl of the larger twin. After I opened the testa I could see the depression in the cotyledons of the larger twin where the smaller twin was nestled during development. It would be fun to learn of the twins others have found. I’ll try to follow these and share some pictures as they develop. If twins each have their own testa, they are basically in separate ovules and, although visually different as they mature, are both products of different eggs being fertilized.
file-21 two.jpeg

Very interesting. Thanks, David!

This reminds me of something my late friend Les Hannibal wrote to me back in 2001.

I remember once I stepped on a > Crinum Moorei > seed and squashed it near flat. Much to my surprise the embryo blasted into near 20 seedlings. It probably damaged its immature basal plate. That might be developed into a means of multiplying seedling stock.

I have produced a research document on this subject, see attached. (Sorry the photographs failed to send) I could send the complete document by email if required, sorry I am not very tech savvy.

(a growing history)

Nepean Blue Mountains Hawkesbury Regional Rose Society, Australia
The purpose of this research was to follow the growth history, from an embryonic seed state to a fully developed flowering state. To aim was to watch each twin survive and proceed to maturity and eventually to flowering. The author has no formal horticultural qualifications and the language used throughout this document is targeted to a rose hybridizer.
The definitions that were created and used throughout this document are as follows:
Term Meaning
SENIOR The first seedling to germinate.
JUVENILE The seedling that germinated subsequently to the senior seedling.
POLYEMBRYONY Polyembryony is the phenomenon of two or more embryos developing from a single fertilized egg. Polyembryony occurs regularly in many species of plants.
SIBLINGS Twins share their DNA. The term is used to describe both seedlings growing together.
TWINS Twins are two offspring produced by the same hybridization.

During a hybridization of Rosa ‘Joyce Abounding’ and Rosa ‘The Golden Child’, four sets of twins were produced. The first of the twins appeared on or about the 30 April 2018, the second of the twins appeared on or about the 28 May 2018, which equates to approximately twenty-eight days or four weeks, the third of the twins appeared on or about the 15 Jun 2018, and the fourth of the twins appeared on or about the 05 Jul 2018.
The above phenomenon is referred to as Polyembryony and pronounced as (poll·e·em·bree·on·nee) and is also defined, in part, as a phenomenon of two or more embryos developing from a single fertilized ovule. Polyembryony occurs regularly in many plants and animals. The Nine Banded Armadillo, for instance, usually gives birth to four identical young. The term is used in botany to describe the phenomenon of two seedlings emerging from one seed.
Polyembryony was first described by Antonie van Leeuwenhoek in 1719 when the seed in Citrus was observed to have two germinating embryos. As with other species, due to the many embryos developing in close proximity, competition occurs, which can cause variation in seed success.
I found it interesting that in several species, polyembryony has been linked to genetic causes and appears to result from hybridization (Maheshwari and Sachar, 1963).
All twins were treated the same way as all other seedlings, the twins generally tended to lag behind all other seedlings which germinated at about the same time. The twins fell behind in growth, which would be expected as two siblings are surviving on the same available nutrients in the seed.

There are some roses that seem to give twins more often than others and some of those that produce this phenomenon more than others are Rosa ‘Rise 'N’ Shine’, Rosa ‘Dorcas’, Rosa ‘High Voltage’. It should be noted that Rosa ‘Rise ‘N’ Shine’ has Rosa ‘Little Darling’ in its Parentage Tree, as both the seed and the pollen provider at different stages, and this could just be a coincidence, Rosa ‘Rise 'N’ Shine’ is part of the pedigree of Rosa ‘Joyce Abounding’.

In total nineteen seedlings from this cross germinated, four of which produced twins, this equates to approximately, 21%, of all seeds to germinate, which is reasonably high percentage one would think.

The twins growth cycle has been chronicled by photographs each mid-month (on or about the 16th of each month), on a monthly basis.

The researched documentation on this subject indicated that both stems are likely to develop at the same rate, but this was not the case with my rose twins. One of the siblings would advance more than the other as they progressed, although this was more prominent in the more senior twin. When we have twins coming from a common testa or seed coat, one twin is normally much smaller than the other, and may need to be nursed along. Both the junior and senior siblings progressed normally, alongside other rose seedlings, whilst the third twin failed and it has appeared to have strangled itself, with the two stems twisting in on them. At that stage, approximately six weeks past germination, it had only advanced slightly past the leaf opening stage. The fourth twin simply failed to advance past the germination stage and had two distinct sets of cotyledons or seed leaves. It had only advanced to the leaf opening stage into its fifth week past germination.
The two remain siblings were potted up approx. eighty days after germination. The senior sibling slightly more advanced and the junior sibling slightly less so. Both sets of siblings had a good root system, and each set of siblings also had five, or better, true compound rose leaves.
The senior set of siblings started to show signs of yellowing and minimal dieback on some leaves and in the main looked in a feeble condition after a period of seven days after being potted up.
The junior set of siblings, also started to show signs of some yellowing of some leaves but in the main looked to be healthy and was showing signs of new growth on both siblings over the same period of fourteen days, after being potted up.
The senior set of siblings has started to show signs of the second stem failing and is not showing any signs of growth after another period of seven days after being potted up. The juvenile set of siblings has started to show signs of the second stem failing and is not showing any signs of growth after another period of twenty-one days after being potted up.
Both sets of the remaining twins are now growing strongly and it would be expected that both will move on to their next growing stage of flowering. The juvenile twin was more advanced than the senior twin at the flowering stage, I believe this to be due to the fact that at the time each stem failed, one stem of the juvenile twin was growing more dominantly then its sibling. The stems of the senior siblings were consistent in their growth, throughout all stages of their growing cycle, therefore sharing all nutrients and nourishments, as depicted in the photographs below:
The twins have failed on both seedlings, the sibling to fail was the second, and the underperforming stem on both. It is interesting to note that both twins failed within 28 days of each other, after germination. As an example Twin A, which germinated on 30 April 2018, failed on the 13 August 2018, whilst Twin B, which germinated on 28 May 2018, failed on the 10 September 2018. They germinated 28 days apart and subsequently, failed 28 days apart. It is as if they have a life expectancy of 105 days. (The dates are an approximation, as I could not say for sure when each stem had failed, they are based on a comparison of the appearance of each stem). Both the remaining seedling are growing well and proceeding to the flowering stage of their growth cycle. As shown below in the Siblings Timeline:
Siblings Timeline
Relevant Dates
Siblings Hybridized Planted Germination Potted Up Failed
Senior Sibling 25 Oct 17 11 Apr 18 30 April 18 18 Aug 18 13 Aug 18
Junior Sibling 25 Oct 17 11 Apr 18 28 May 18 18 Aug 18 10 Sep 18
Third Sibling 25 Oct 17 11 Apr 18 26 Jun 18 08 Aug 18
Forth Sibling 25 Oct 17 11 Apr 18 04 Jul 18 18 Jul 18

The fertilizing regime for all seedlings was half strength Neutrog “Sudden Impact for Roses” liquid fertilizer, applied twice a week. It was thought unwise to either increase the strength or application timing for the twins.

The seedlings shown below are shown in their order of appearance and not in the order of seniority.
The juvenile seedling is growing well and has advanced on to the next growing stage of flowering. The juvenile seedling flowered forty four days after its sibling had failed.
The juvenile seedling has been the more advanced of the twins, as detailed earlier, than the senior seedling throughout the whole growing cycle, therefore the junior seedling was the first to produce a flower. It would not be expected, nor would it be logical to expect, that both seedlings would produce the same flower as at this stage they are both different roses, on their own root system. The senior seedling is growing at a consistent rate and has advanced on to the next growing stage of flowering. The senior seedling flowered seventy four days after its sibling had failed. The senior seedling has been the more underdeveloped of the seedlings, as detailed earlier, and the junior seedling was the better performer throughout the whole growing cycle. This seedling appears to be lacking the qualities to advance past the benchmark required to avoid being rejected. The senior seedling was the second to produce a flower.

Within the genus Rosa there are over three hundred species and thousands of cultivars, yet there is only a selected few species or cultivars that are known to produce this phenomenon. Some Rosa species or cultivars hybridize and produce the twins or Polyembryony more easily than others. The production of twins is due to the characteristics of one or both of the parent’s recessive genes and will not only give us what we are looking for, but can also give us what we are not looking for. A degree of providence is also involved, (good or bad, whichever way you look at it). At the end of the day once one of the siblings fail, the remaining seedling is just another seedling to advance or fail on its own merits.

Happy to find this topic. I found twins too, mother plant is a tetraploid tea hybrid rose, so I suppose, a smaller one must be a diploid. The twins shared one testa.
I will keep an eye on those two. The small one also has a tiny bud now. The differences in growth are quite obvious, I think.

Not sure if these are the type of twins y’all are talking about. See pics.

Very interesting! Now I am wishing I had carefully labeled the twins I found this spring to track their progress and compare them.

found a picture I had taken. I wish I had thought to take a picture of them after I carefully dug them up, and then again after separating them.

They seem to have their own testa for each one. Would be nice to see how they develop.

Nice! So it is not such a rare event. Do you see the differences in growth of those two?

Hi Joe!!

Are the two pictures of the same seedlings at just different stages? It looks like each has their own testa inside of the achene. I wonder in this case if two of the four products of meiosis survived on the female side in this case and each developed into its own ovule and were uniquely fertilized.

Thank you Giessen for sharing the photos of your twins!! I hope you share updates with us as they open.

Thank you Gary for your very detailed description of your twins. It is sure hard sometimes keeping the weaker one alive… It would be challenging being a citrus breeder… In their case the majority of the embryos from my understanding (some seeds have 3 or 4 embryos) are clones of the mother and are from maternal cells within the inner layer of the ovule and then there is one sexually derived embryo typically. It is hard for the breeder to determine which is the new desired hybrid. In the case of having a 4x and a 2x seedling in roses sharing the same testa from a 4x mom, I suspect apomixis (without or away from mixing) reflects what is happening here more than polyembryony as the smaller lower ploidy twin is likely from a synergid and not a fertilized egg that splits to make identical twins later. Once years ago I had twins that shared the same testa but were about equal in size and they developed into plants with the same growth and flowering characteristics and were both tetraploid. I suspect they were from polyembryony-splitting into 2 embryos after fertilization. I didn’t run DNA markers on them to see if they had the same fingerprint. It is fun all the variations how embryos can be derived and the genetic consequences depending how it occurs (clones of mom, haploids, identical twins of each other, etc.).

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A little update to my twins (s. image above)…
Both are blooming now.
The left one is a ‚small‘ twin, and the one to the right – is a ‚big‘ twin (still not opened completely).

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Thank you for the update Giessen! It is fun to see how different the flowers are. It’ll be nice to see continued updates and learn if the smaller twin has fertility for you. Many of the smaller diploid twins in my experience that I’ve counted aren’t very fertile, but thankfully once in a while some are. I want to focus more this year on using a couple diploids that have fertility out of ‘Gaye Hammond’ and High VoltageTM more in crosses with polyanthas to try to bring in warmer colors to polys.

Thank you David for your response.
It is indeed fascinating.
Of course, I would love to check the fertility of the diploid twin. It actually has a very nice scent and lovely bloom color. The other one looks promising as well. They are still in tiny 4x6 cm pots, I will have to repot them.
This year I am also going to cross some diploids.
Last year I crossed Penny Lane (seed) with a groundcover rose Charmant from Tantau (pollen), and suprisingly, the seedlings have the appearence of Charmant (very similar foliage). I wonder, if Penny Lane is 4n and Charmant is obviously 2n, shouldn‘t the seedlings result in 3n. However they look like 2n. Of course, to be sure, one has to make chromosome counts (or at least to analyse pollen)… But could it be that Penny Lane was 3n?

I got to thinking about this, and a couple of cases I’ve encountered in reading.
Michurin (1934) p. 219
Fig. 39. The result of fertilizing almond with apricot pollen (the fruit obtained had an almond seed with four kernels from which four plants developed.)
H. H. B. Bradley (Hybridising at the Antipodes) 1906)
Calostemma luteum x Pancratium maritimum gave a quantity of fertile seed, and the seedlings are growing strongly, but none have flowered yet, so it is uncertain if the cross has taken. Curiously, several of the seeds developed two plants, and one seed gave three plants from the one seed.

Then I went searching for other info:

American Journal of Botany 30(6): 408-415. (June 1943)
Haploid-Diploid Twin Embryos in Lilium and Nicotiana
D. C. Cooper
TWIN SEEDLINGS have been noted in a number of species of angiosperms wherein the individual plants differ from one another in that one member of the pair has the normal diploid chromosome number characteristic of the species and the other may be either haploid, triploid or tetraploid (Muntzing, 1937). The haploid-diploid combination was first reported in 1933 independently by Kappert in Linum and Ramiah, Parthasari and Ramanerjam in Oryza. Since these reports this combination has been found in Dactylis, Triticum, Poa, Phleum, Solanum, Nicotiana and Gossypium (Webber, 1940).
A synergid is stimulated to divide in approximately 1 percent of the ovules of the seven species of lily examined. Twin embryos, one haploid and one diploid, are thus produced. Both embryos may continue to develop but usually the haploid embryo disintegrates at an early stage of development. …
It is suggested that embryos arising from synergids may account for many of the haploid plants which have been discovered.\

I read (somewhere) about a tetraploid American Rosa species (which one?) that was rarely found as a diploid. Maybe these were really haploids.

Does hybridization provoke more twinning? Here’s another suggestive example.
Hereditas 30(4): 567-582 (May 1944)
The original material was derived from three different stocks, two of which were pure M. sativa, while the third was the so-called Ultuna lucerne, which consists of a population of M. sativa X falcata crosses. Treatment of seedlings at the cotyledon stage with colchicine-agar produced some tetraploid plants (2n = 64), and these were propagated by means of cuttings. Crosses within these gave rise to three tetraploid seed plants. After crossing a tetraploid plant of pure M. sativa with a diploid plant of the Ultuna lucerne from one seed twins with a triploid complement of chromosomes (2n = 48) were obtained.

Well, this one does not support the synergid model, so maybe twining is not a single phenomenon. And it may be hereditary.

Polyembryony and Breeding Behavior of Polyhaploid Twins in Alfalfa (1958)
Dean Stewart Bishop
Masters Thesis: Kansas State College of Agriculture and Applied Science
Frequenoy: The percent of polyembryonic seeds occurring in various varieties is shown in Table 2. The results suggest a difference in frequency among the varieties. The highest monogerm to multiple embryo seed ratio was shown by Buffalo (1,330:1) while Rambler showed the lowest (514:1. The overall total indicated that polyembryos occurred about one in 300 germinated seeds in the population studied.

A striking difference was evident when frequency in the varieties was compared with that of the s1 seeds of twin plants. The results show an increase of nearly 10 times that of the studied varieties. An increase in the number of multiple embryos was evident in three sets of twins, T13, TlU and T104. T13 and T104 produced about one in 60 and one in 90 germinated seeds respectively. T14A and B exhibited a definite increase in the occurrence of plural embryos, A producing one in 6.4 seeds and B producing one in 7.5 seeds.

Thank you Karl for this interesting information.
Maybe I can contribute a little bit by mentioning that last year I perfomed about 140 different crosses (which produced seeds), so, out of these I had about 450 germinations, and this was a single event of producing twins among them. The crosses I made were mainly in modern roses with a few older remontants. No species roses were involved.

My suspicion about species crosses being involved was probably due to the first cases I encountered being somewhat difficult crosses. Still …

I remembered what I had forgotten: It was C. C. Hurst who wrote that Rosa carolina (L. 1753) is tetraploid, rarely diploid. It is not difficult to imagine special cases where a diploid seeding (haploid of tetraploid species) might gain an advantage. This would be another way higher polyploids can give lower ploidy offspring.

Going through my notes I found another example that I hadn’t added to my web page. This one involving iris twins. One was larger than the other. I have not yet found any follow up.

And another regarding tomato twins.

By the way, regarding the diploid-haploid or tetraploid-diploid twins. If the smaller one came from a synergid, it would be purely maternal. The other one would be “daddy’s little angel.”

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So, if I understand correctly, if the smaller one comes from a synergid, doesn’t it have to have the bloom color like mother plant? Or not necessarily? I imagine, it has to resemble its mother qualitatively (not quantitatively). So, what I mean is, it may have less petals and shorter habit, but the bloom color, also maybe the type of fragrance, the texture and the color of the foliage have to be be very similar.

Hi Giessen,

Not necessarily. It just has a subset of the genes of mom, so depending on what it inherited it may look like mom or a bit different. For instance, I had a diploid small twin out of the Buck rose ‘Dorcas’ and it was a soft yellow. ‘Dorcas’ must have some genes for yellow in it that are recessive and/or masked some by the pink anthocyanin pigment. So, without the anthocyanin being produced (likely didn’t get those genes that allow for its production) and maybe not getting a gene that suppresses yellow pigments from being produced, it was a soft yellow. Haploids like this are a fun way to sometimes unmask and directly see some of the recessives a parent may possess and be transmitting in some of its gametes.
Dorcas and haploid.JPG

That’s a great example to have on hand.

HelpMeFind gives the parentage of Dorcas as Minigold x Freckle Face.

Is the yellow haploid at all fertile? It would be a nice example of “breeding down” from tetraploid to diploid, rather than the more familiar upwards trek. Maybe give ‘Isabella Sprunt’ a more dramatic daughter.