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N. Tabacum x N. Rustica

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Tutu

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Whereas the How to Intentionally Cross Tobacco Varieties is a great thread going into the technical aspects of how tobacco flowers can/should be crossed, and whereas Kasturi x Amersfoort was a good test case for a crossing of N. tabacum x N. tabacum, I think it's time to go a little bit deeper into inter-species crossing. In this case, the two species that are to most of our interest since they can be smoked. So should be the case for their hybrid version. This thread will be a discussion on what we know about crossing N. tabacum and N. rustica. I know it has been discussed in other threads, such as Sasha's Hopi, Red African and White Mammoth Grow, but I think there's a whole lot more for us to discuss. So let me summarize a few things that have been said already, then add a few new things I've recently dug up, so that we have a bit of a starting point to continue with

In the thread How to Intentionally Cross Tobacco Varieties it was Guzzy who asked whether one could cross N. rustica with a variety of N. tabacum. Deluxestoge replied that unless you tried a large amount of crossings, you are not likely to get any results. ChinaVoodoo posted a piece of text on Delgold, which tells us that after obtaining a hybrid, it can be crossed back to N. tabacum to overcome fertility issues. Upon a bit of discussion I posted a quote where someone held that crossing N. rustica with N. tabacum would only work if N. rustica was used as a female. I posted a link on "Hybrid lethality in the genus Nicotiana" and Deluxestogie inserted an image showing haploid chromosome numbers per species of Nicotiana. I'll post it here because it's a good start-off for this thread.

Chromosome Numbers.jpg

For the sake of this thread, let's continue a bit on the crossing of N. tabacum and N. rustica. Other interspecies crosses may be worthwile too, but if anything, the goal for me would be to create something that we can cure and smoke. From the previous "Hybrid lethality in the genus nicotiana" that I posted I got some interesting sources. In the lethality article you may find a table under the heading "5.1 Hybrid lethality in crosses using N. tabacum". This table shows results of crosses in the Nicotiana species made in previous studies. It summarizes the results. At the very end of that table, one page later, it shows crosses of "N. tabacum (24) x N. rustica (24)" as well as "N. rustica x N. tabacum". The table gives "Viable" returns for the F[SUB]1[/SUB] phenotype of the first cross and both "Lethality" and "Viable" for the second.

This points me back to when I asked why the cross of these two would only work in one direction, with N. Rustica as a female. By now I've read this statement in a few other articles as well. The current article lists them as possibly viable in both ways. I'm talking about viable to germinate, not fertile to reproduce. This hybrid lethality article does mention to take caution interpreting their findings. They themselves are looking for lethality occurrences in other published research, but they mention that for most researchers the objective is to find viable seed, thus omitting results where crosses actually did die.

In their table they refer to an article by E. M. East in 1935 when it comes to viable offspring with the crosses of "N. tabacum (24) x N. rustica (24)" and "N. rustica x N. tabacum". It leads to a series of articles called "Genetic reactions in nicotiana". There are three of them, and they will be the vocal point of this new thread. However, feel free to find and add other articles that may be interesting to share.


Genetic Rections in Nicotiana I. Compatibility

It is important to note that the aim of this paper is to get an overview of which species of Nicotiana are eligible for interspecies crossing. Personally my focus is on his trying attempting to find hybrids between N. tabacum and N. rustica. In case anyone is interested in attempting any other crosses with other Nicotianas, this is the place to look for what you can, and cannot do. The following is a table of compatabilities of interspecies crossing.
Compatabilities.jpg
On the left are the female parents of each cross, and on top are the male counterparts. The paper tells us that each cross was tried at least 25 times and in some cases over 1000 attempts. As you may find above, each cross in the table gives two results. The result on the left is the result that was most frequently found. The result on the right is the less frequent outcome of the cross.

· X: No stimulation
· P: Parthenocarpy
· WE: Non germinating seeds
· WS: Non-flowering seedlings
· HL: Large hybrids
· HM: Average sized hybrids
· HS: Small hybrids
· HD: Dwarf hybrids

Going back to our table we can see that with N. rustica as the female plant and N. tabacum as the male, there were often non germinating seeds, which he also calls weak embryos. Less frequent, though occurring, were large hybrids showing great heterosis. The other way around there were mostly no seeds. However, we can also see that on a less frequent basis there were large hybrids. The rest of the article discusses the results of each cross briefly. There is one more mention of this cross, which has the author stating again that it is more easier to cross in the direction of using N. Rustica as a female. The author also makes an attempt at explaining the reason for this. For me this was an important find in confirming that the direction of the cross is easier made in one way, than in the other, though not impossible, it seems:

"There are 11 examples where the cross is easier to make one way than it is the other. N. bigelovii x N. tabacum and N. rustica x N. tabacum -- both combinations where n=24 -- are easier to obtain than the reciprocal combinations"


Genetic Reactions in Nicotiana II. Phenotypic Reaction Patterns

The next step both for East and for us is to look at phenotypic aspects of the crosses. First at flowers, then at leafs. He explains in the beginning of his paper that his goal is not to simply give descriptions of what he observes, but to find specific rules for phenotypic reactions when making crosses. This would be interesting for us as well, not only for crossing N. rustica with N. tabacum, but also when making crosses of N. tabacum x N. tabacum. One interesting thing in this respect is that apparantly the red/pink colour of the flower of N. tabacum is always present, no matter what other Nicotiana species it is crossed with. Too bad if you want to cross N. tabacum with N. rustica only to get good smokable leafs with yellow flowers on top of your plant. Apparently, the only exception to this rule is when N. tabacum is crossed with a species having 9 or 12 chromosomes. Interestingly enough, the shape of the flowers do change accordingly. There are a number of example photos in the paper and I'll be posting two of these.

Flowers A.jpg

This image gives a good example of what hybridizing does to the flowers of tobacco plants. In the very middle you find the N. tabacum flower all by itself. To the outer left and right you find the flowers of the parents with which it was crossed. In between are the hybrid flowers. Now it should be clear to anyone that these hybrids do indeed resemble both parents. In the cross with N. bigelovii the flower becomes slimmer, whereas the cross with N. glutinosa the top of the flower becomes more bell shaped.

Flowers B.jpg

Even more interesting is to look at what we might expect as outcomes for our intended cross N. tabacum x N. rustica. In the above image we need to look at the middle row. On the far left an example of N. rustica, in the middle an example of N. tabacum, and inbetween we find its hybrid version. I wonder whether the Delgold flower resembles it in any way. East observes the effects of different crosses on the flowers of N. tabacum. He notes that N. rustica affects it by shortening and broadening the corolla tube.

So we move on to leafs. Now I'm afraid that the author looks at leafs from a different perspective than we do. That doesn't make his research any less interesting though. Before showing some of the results he finds, I would like to quote an important statement of his concerning the differences of genetic influence on the phenotype between flowers and leafs:

"The size of the leaves is influenced markedly by external conditions. Leaf-size is also influenced decidedly by internal conditions. If the hybrids are vigorous, the leaves may be large; if the hybrids are weak, the leaves may be small. It seems as if the machine does not run properly in certain combinations. It is also to be noted that the determination of sessile or of petioled leaves appears to be controlled by relatively few genes, as in varietal crosses between sessile and petioled N. tabacum varieties. The reaction pattern that seems to be comparable with the flower patterns is that which controls the shape or proportions of the leaves.

Most of the varieties of N. tabacum which I have grown myself had sessile leafs, except for Ainaro from Timor-Leste, which shows tall petioled leafs. The paper then gives a number of figures, of which I will show you the most complete, which includes the cross that I'm trying to investigate here. If you're looking for more examples, have a look at the paper itself.

Leafs.jpg

The variety of N. tabacum used in this experiment to cross with the N. rustica is Cuban. The N. rustica here is Texana. Most interesting in this version of the cross is the hybrid version of the sessile N. tabacum and the petioled N. rustica. The examples of hybrids with other species of Nicotiana are also interesting, especially if anybody here ones to give crossing a different shot.


Genetic Reactions in Nicotiana III. Dominance

The author of the previous paper wrote a third to approach the topic of dominant and recessive genes in the species Nicotiana. He attempts to do this for the whole genus of Nicotiana, finding general rules of thumb. For us, this might be interesting, but quite detailed, whereas right now I am particularly interested only in the genetic and phynotypic effects of the cross N. tabacum x N. rustica. The whole paper is still a very interesting read. But instead of unraveling the whole text here, I'd rather quote a piece of text which focuses on our specific topic of interest: N. tabacum x N. rustica:

"During the early years of my Nicotiana investigations, over 100 varieties of N. tabacum and N. rustica were studied. Some were discarded after one population had been grown; others were observed in selfed lines over long periods (maximum 28 years). Numerous crosses were made between commercial types of N. tabacum and, to a less extent, between commercial types of N. rustica. Several studies, involving hereditary differences in height, number of leaves, and size, shape, and texture of leaves, were published by Hayes, Jones and East. It is not necessary to discuss these reports beyond saying that the selection experiments on these quantitative characteristics, such as those published by East and Hayes (1914), were carried on to the F[SUB]13[/SUB] generation and were evaluated statistically, though not published. They are mentioned for one purpose only. This very considerable experience with continuously self-pollinated lines of supposedly pure types and with successively selected self-fertilized lines from varietal crosses indicated very strongly that though there is a rapid approach to homozygosis of gross characters, there still remains a large amount of apparently irreducible variability in all parts of the plant, some of which can be proved, by appropriate tests, to be heritable. For example, the tobacco leaf varies in characteristic items of shape from the tip to the base, in color, in size, in veination, in fullness between the veins, in thickness, and in various other factors that go to make up commercial quality after what is ordinarily thought to be a pure type is obtained. In selected descending self-fertilized lines, one may obtain a still more uniform progeny temporarily, as regards almost any peculiarity, thus showing a residuum of heterozygous genes affecting these characteristics. But there must be a high mutation frequency since, after 10 or 12 generations of closest inbreeding, one can not reduce, in an appreciable degree, the variability of any indices for the leaf as a whole."

Personally I think this is not only interesting for crosses between N. tabacum and N. rustica, but also for crosses N. tabacum x N. tabacum, or for N. rustica x N. rustica, as it is for non-crossed varieties on their own. I think it's something we've all been observing on our own, growing varies varieties from all over the globe, having them adapt to our local environment, sometimes more quickly than expected.

In this third paper, there is an interesting anecdote on self fertilizing N. rustica and observing the variation. It'll be another long quote, so maybe it's better to look it up in the text if you're interested.

------

All in all I think this is a very interesting starting point for myself and for others, to make an attempt at something that may be very difficult, but very rewarding. Personally I am not looking for anything specific in the hybrids, I'm just going to try and make it work. I'll see what the outcome is, if anything.

Please share your thoughts.
 

Tutu

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Old shouldn't be a problem, the other articles are old as well. Definitely interesting when genetic dominant and recessive genes are concerned. Although it mentions both N. tabacum and N. rustica, it's only for comparing nicotine content. As nicotine is higher in N. rustica, it might be economically more interesting to grow if merely for the nicotine. However, the focus on the article is on crossing N. rustica x N. rustica, without any N. tabacum being involved. Still worth your reading time, that's for sure. I'm quite impressed by some of the data tracking these fellows were doing back in the day. Unfortunately I don't think I'll be measuring the nicotine content. Neither have I done so for Kasturi x Amersfoort, which would have been interesting as the maternal parent is supposed to have high nicotine levels.
 

Hasse SWE

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Tutu I prefer old studies before newer pretty much before the variants I mostly grow are just old variant and match of the big leaf N.Rustica variants really are made by cross N.Rustica x N.Tabacum, I was for example trying to get some GCT-3 and DCT-4 seed last season (was also promised some seed but they never turned up), I was asking if they had some information about studies of the Alkaloids in the variants and the level they showed me was lower then much of the studies of older types of Dark-air/fire cured variants.
 

deluxestogie

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This is an interesting and well documented thread. For the questions that are addressed in your discussion, I'm afraid that there is nearly no new research. Tobacco used to be the premier subject of plant genetics research, in universities across the world. The tobacco plant has been largely displaced in plant genetics today by Arabidopsis thaliana, which doesn't bear the taint of being smokable. (It's the politics of science.)

Thanks for posting this, and for Hasse's link as well. I'll add this thread to the Index of Key Forum Threads: http://fairtradetobacco.com/threads/3868-Key-threads-in-the-FTT-forum

Bob
 

deluxestogie

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I've taken the table Tutu presented, and highlighted only those crosses that generally produce a viable hybrid.
HL: large hybrid
HM: medium hybrid
HS: small hybrid
HD: dwarf hybrid

NicotianaCompatabilities.jpg


Focusing on interactions involving either N. tabacum or N. rustica, we can draw the following conclusions.

N. tabacum FEMALE can be reliably pollinated with:
  • N. glauca
  • N. glutinosa
  • N. tomentosa
  • N. sylvestris
N. tabacum can reliably pollinate:
  • N. bigelovii
N. rustica FEMALE can be reliably pollinated with:
  • N. paniculata
N. rustica can reliably pollinate:
  • NONE

So we can say that N. rustica usually doesn't play nice with others, whereas N. tabacum has better success in forming hybrids. N. bigelovii is a wild species that was widely cultivated by Native Americans in large areas of western North America.

Bob
 

ChinaVoodoo

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I find all of this fascinating, but I can't see having the time or space to begin cross breeding for the next few years.

A project I had thought of-far beyond my scientific ability-would be to take N.otophora, N.tomentosiformis, and N.sylvestris, and attempt to recreate an alternative line of N.tabacum.

Addendum:

Just some heavy reading
https://www.frontiersin.org/articles/10.3389/fpls.2016.00843/full
 

ChinaVoodoo

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Is it possible to take a single cell, remove the chloroplast, then replace it with a chloroplast from another species, then clone the cell into a full grown plant?
 

OldDinosaurWesH

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And you still wouldn't know exactly what you had. Is the tobaca-zilla you just created viable, and if so, will it take over the world? Hey! maybe we could make a bad science-fiction movie about this. I heard Harvey Weinstein is looking for work.

Wes H.
 

Tutu

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The table is indeed an interesting one Bob. Whereas I'm aiming at the N. rustica x N. tabacum for now, the other possibilities are very interesting too, and some of them possibly much easier to establish. I'm not quite sure what other hybrids you think might produce leaf that can be smoked? I'm not very educated beyond the tabacums and the rusticas of this world.

Delgold: A New Flue-Cured Tobacco

This is an article on the tale of Delgold, prviously highlighted by ChinaVoodoo. Now besides that it is a very very interesting experiment with outcome by itself, it is especially interesting regarding the cross I'm attampting to make. Delgold was created by a series of crosses. The first cross involved N. rustica x N. tabacum. The N. tabacum used was Virginia 115, and the N. rustica they used was Babor. The hybrid they obtained from this cross was not producing seed by itself. To get seed from the F[SUB][SUP]1[/SUP][/SUB] they had to make backcrosses with N. tabacum, for which they used Virginia 115 again. They call these generation BC[SUB][SUP]1[/SUP][/SUB]. Most of the plants in this generation were bushy and did not produce seeds by itself. They made another backcross with N. tabacum, this time to a Hicks Broadleaf variety. This BC2 was then crossed back to Virginia 115 to create a BC3. A single plant from this generation led to Delgold. To make a stable variety, it had to be selfed multiple times.

The remainder of the article is mostly concerned with the phenotype of Delgold in comparison to that of Virginia 115. It also shows a picture of Delgold. It's an interesting read, but I won't go into detail about the comparison of the two plants. For now, my point of interest is the fact that they were able to make a hybrid, and make it fertile again. My intention is to create a plant which looks a little bit more like an actual hybrid, rather than only obtaining a higher alkaloid level through crossing with N. rustica. No, my plant should look like a real hybrid!
 

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But that pdf on Delgold doesn't express the true hardship involved in making Delgold. It was like back cross x a gazillion, hundreds of mortalities. Crazy crazy science. It's elsewhere in the forum. You've probably seen it.

I'm not saying it can't be done again. If there's anyone who can do it again, it's someone like you who works for a tobacco company, but seriously, man, I'm just saying, make that move with pride.
 
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Tutu

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Yes I know. As was said in the lethality article, most authors focus on the successes they've had, not on the failed attempts. I do realize it will not be something achieved overnight. Perhaps, I won't achieve anything at all. But just the thought of giving it a shot excites me tremendously!
 

ChinaVoodoo

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Yes I know. As was said in the lethality article, most authors focus on the successes they've had, not on the failed attempts. I do realize it will not be something achieved overnight. Perhaps, I won't achieve anything at all. But just the thought of giving it a shot excites me tremendously!

I think it's really cool. I would love to see you succeed.

I haven't done enough research to say I'm correct, but it seems to me that the best tobacco research that's publicly available is the Canadian research. Google half of the Canadian varietals, and you'll find some really interesting documents. Not saying the American research was not great, just not as easily obtained. The Canadian research seems to be intended to be public from day one because socialism, while U.S. Research is private because capitalism.
 

BigBonner

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I think it's really cool. I would love to see you succeed.

I haven't done enough research to say I'm correct, but it seems to me that the best tobacco research that's publicly available is the Canadian research. Google half of the Canadian varietals, and you'll find some really interesting documents. Not saying the American research was not great, just not as easily obtained. The Canadian research seems to be intended to be public from day one because socialism, while U.S. Research is private because capitalism.

I would say that you don't find much on a lot of USA research is because the research belongs to the Big Tobacco companies and is under lock and keys like their trademark casings that they add to their products .
 

Hasse SWE

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I most say that I think that Bigbonner have a big point on that. I have a similar problem to get information about a old time Swedish tobacco variant. I believe that I can find information both about the variant and around that variant but the biggest problem is named COPYRIGHT. often a pretty good thing but in this case it's pretty bad and silly. So you guys might have the same problem. No one cares about some of the studies but they have a copyright and don't just drop information. In other cases they know about studies and they really need to keep the information..
 

Tutu

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It's partly that, but it's also that a lot of people seem to have lost interest in tobacco. Now research on the plant is a very different thing from smoking the leaf. Nevertheless, some of it goes hand in hand. It just isn't the most popular thing around anymore. Let me see if I can dig up a few more interesting articles before my plants have grown to adulthood...
 

Tutu

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Smokes, this is great. It's like Tobacco Google, if you will. Found a few interesting things. The first article I want to discuss here didn't live up to its expectations regarding new information on the N. tabacum x N. rustica cross. However, quite unlike other scientific articles that focus on hybridizing, it turned out to have a smoke panel rating the hybrids by level of satisfaction, smoking them as cigarettes.

Comparative comporisiton of certain Nicotiana species and their F[SUB]1[/SUB] hybrids with N. tabacum

The article's main focus is the distribution of nicotine vs. nornicotine within the species Nicotiana, and how these levels change when their being crossed. For example, they mention that relative to other Nicotiana species, levels of nornicotine in N. tabacum and N. rustica are very low. On the other end, the level of nicotine is very low in N. glauca. For me, as usual, I'm extracting information on hybridizing different Nicotiana species. The first interesting quote I read was not so much on "how to", but more so on "why to";
It seems hardly likely that, in its evolution, all of the desirable qualities of the ancestral species would have been retained and all of the objectionable properties discarded

The goal of the plant is to survive and reproduce. Whereas our intention with it is not exactly the same. We look for certain qualities in tobacco that are unlikely to be selected by nature for the same purpose. This makes it interesting to make an attempt to hybridize, because you might create offspring that combine two or more good qualities from both parents. Equally, and probably more often, you will not. Nevertheless;
Because of their greater economic importance, intraspecific hybrids have been studied much more extensively

Most hybrids are N. tabacum x N. tabacum hybrids. Not only are they more economically interesting, they're also much more easy to make than crosses across species. Not more interesting to make though, I would like to argue. And in theory, the latter could be very rewarding;
Judging from their striking morphological contrasts, one might predict still other inherent differences in chemical constitution among the Nicotiana species. Some of these as-yet-unidentified unique occurrences or absences might conceivably be beneficial to quality

Unfortunately where the article fails to yield much results is where they have results for a comparative composition on many different constituents for many hybrids, except for a few where they didn't get a satisfying hybrid. One of these was N. tabacum x N. rustica. So there's no results listed either. with N. tabacum x N. rustica they attempted to use varieties SC. 58 and Brasilia. If your interested in the found components in other hybrids, please find the tables in the article.

For our current discussion I want to turn to something else. The "Hybrid Smoking Panel", so to say;
Leaf appraisal and smoke taste. Prior to manufacture into cigarettes, the surplus cured leaves of these experimental "tobaccos" were critically examined by research personnel of the cooperating manufacturer. Their appraisals are summarized in table 2 which also records certain physical measurements made on granulated samples.


Cigarettes made from these experimental tobaccos were smoked by a panel of experts. These panelists were trained to distinguish and describe rather subtle differences in the taste of flue-cured cigarettes. Generally, the taste of these "foreign" cigarettes exceeded their vocabulary of descriptive terminology. All of the foreign cigarettes were severely criticized; some were extremely irritating. The smoke of several was described as having a "mousy" taste which is usually associated with myosmine. In general, the hybrids were more offensive than the species. The panelists' appraisels of the smoke tastes are summarized by table 2

Leaf Appraisal.jpg


Perhaps I should be glad that there's no N. tabacum x N. rustica listed here. I probably would have discouraged me slightly.
 
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