laughing gas to make tetraploids?

Rose Genetics Ploidy Level
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41

Okay, (sorry, but I woke up thinking about this thing…) so now I’m wondering if germinating seeds within Jim’s apparatus, or micropropagules might work?

Jim, have you any experience with micro tissue propagation?

And with seeds, it is afterall the cotyledon which supplies nutrients so light would not be required, obviously… But would you be able to get a pure tatraploid, or would one more likely expect ‘success’ to yield only a chimera where one layer of cells mutated?? My biology is rusty, but I’m thinking at the germinating seed level, the layers are already pretty distinctly differentiated, no?

I’m still thinking the best would be to use fertilized hips to try to nail the earliest divisions of the ovule, but I can’t think of a species I’d be interested in which would fit in there as a mature blooming plant…

42

Hi!

Has anyone of you tried out this until now - or you Jim, again?

I unfortunately had absolutely not the time - and also the money, this spring.

I hope very much to try it in 2012 (would be a good year for this fascinating technique, at the end of the world :wink: )

One problem with wild roses is also, that one has to do it with very small potted specimen.

Its not always possible to get them blooming, I guess.

Perhaps as Bonsai plants (and after 10 years practising) :slight_smile:) !

Any ideas?

Grx!

Arno

43

Here is an old paper from 1973 about the N2O Microtubuli Effects, which I found today, the pdf is free.

Grx!

Arno

Link: jcb.rupress.org/content/58/1/96.abstract

44

I haven’t treated any roses with N2O recently, but I’m thinking about trying it with some of my R. roxburghii normalis X Mons. Tillier seedlings.

45

Hi Jim,

good to read you, a nice combination, your R. roxburghii normalis X Monsieur Tillier, I still also have R. roxburghii x diff. Species Roses seeds, but this year I hadn’t the time to stratify them in winter/spring. So I will do it this February, 2012.

I thought a lot now about the different options that are offered by this N2O theme … . Isn’t N2O toxic to the cells, I mean its at least an oxygenizing substance with two O-atoms? That would mean that there are also damages after treatment, perhaps even genetical damages in the Embryos … not only polyploidization.

And that for I also thought about other possible gases, that are perhaps less dangerous and damaging, but with similar effects.

The noble gas Xenon (Xe) for example perhaps should also be an interesting element for that treatments under pressure, that perhaps (! only a suggestion, so far I found nothing that verified my idea !) also inhibits the Microtubuli - as its also used as a inhalational anaesthetic.

(Wanted to write “inhalalalala”, guess, thats exactly the drugs influence :wink: ).

Don’t know if there is a connection, but maybe there is.

Only problem that occurs before simply trying it out: Xe is not dangerous but very expensive. About 15 Euro / 20 Dollars per Liter.

(I don’t know not for sure if they mean per liter gas at room temperature or per liter of the liquid gas under pressure. I think per liter GAS - and this is really expensive then. …)

So, what remains to be said for me is: Perhaps a Xe-N2O mixture should be also a good try in the future.

Further I think treating germinating seeds is not as good as mature fertilized plants, as there will for sure be created genetically mosaic plants then, and in such a plant the amount of polyploid cells is very often not stable, as the diploid celss are able to grow and divide faster, so that such a plant may be, after a while, again nearly a diploid as further developing whole plant.

And what you mention is also the pressure: perhaps the 90 psi are not enough to get good results. …

Problem here: All pots which would fit which I have checked are up to 90 psi maximum.

I found only one with a 150 psi maximum but it has got only a very small front hole of 15 / 10 cm. Normal plants will be difficult to be packed inside under such circumstances … .

Anyway. There will be a way … . :wink:

Grx! & good luck!

Arno

46

Hi,

the above sentence “Isn’t N2O toxic to the cells, I mean its at least an oxygenizing substance with two O-atoms?” of course is not right, should be just one O-atom. :wink:

Meanwhile I found an upright standing pressure tank up to 16 bar / 232 psi with a front hole that should be big enough for putting in plants of interest.

Only thing is: this tank is over 2 m tall and its weight for that reason is 300 kg . :frowning:

With abt. 1600 Euros its not really expensive but at least not cheap … . We’ll see.

Grx!

Arno

47

Arno,

You should consider using colchicine instead. Crocus bulbs are a lot less expensive.

Don

48

Hi Don!

Yes. But it might be worth it. Razor Blades are less expensive too, if life isn’t beautiful anymore. But I like my naturally given health so far. :slight_smile:)

Colchicine is really dangerous. And I think the gases are much easier to handle, with a very good polyploidization output rate.

Grx!

Arno

49

Colchicine is really dangerous.

I don’t understand why you think so. It’s not something that I would let the children play with but handling it with care is easy enough. 100% water soluble, too, so cleanup is easy.

50

I agree there with you Don, if you are aware of the hazzard then your handling practices should be spot on. I use Colchicine and Trifluralin. Trifluralin has given me some cracking results so far, better than Colchicine.

Warren

51

Trifluralin has given me some cracking results so far, better than Colchicine.

Do you have a protocol that you can share? Where do you get yours?

52

What actaully is the point of trying to increase the ploidy??

53

actaully=actually

54

…I mean what is the point in using chems to do it… nature increases the ploidy from time to time, and that is a pretty safe way, no?

55

Don’t get me wrong, I am not challenging those of you who want to experiemnt this way… it just seems, well…a a whole lot of messing around whne nature can do it sometimes. Each to their own!

56

Nature wants chromosomes to match up during fertilization. This can only happen when both donors have the same number of chromosomes and it happens best when that chromosome number is even. For roses this is diploid or tetraploid, though rarely also hexaploid, octoploid and even decapolid.

It turns out to be easier to increase the number of chromosomes artificially than it is to decrease them artificially. It is also easier to tell when you’ve accomplish an increase because the plants often have more leaves, thicker stems, larger flowers and fruits, a greater number of prickles and so forth.

57

…not wanting to hijack the flow of conversation here believe me, however I once enquired about trifluralin availability here and found it was fairly easily obtainable. I didn’t go ahead with buying any and left it up to nature instead to do the job, particularly because research into this chemical showed up a whole lot of local regulations governing its permitted useage and its correct disposal.

To my mind, it read like the sort of thing a university lab might apply for permission to use and it like totally scared me off the whole idea.

58

Don;

The Colchine was obtained through a chemist friend of mine and the Trifluralin from a farmer who had 20 lt or 5 gallons of it. The Trifluralin solution was 0.013% and it was applied to the seedling terminal bud at the cotyledon leaf stage, put into a humidity chamber for 24hrs and then rinsed off with tap water.

George;Thats why I acquired a Chemical Handlers Certificate for my work, it enables you to acquire some things which the public do not have access too.

Warren

59

Hi you!

I am not sure if one always is aware of potential long term risks of some of the substances; colchicine e.g. is extremely durable where it is applied once.

And since I have a girl friend with a 3 years old child I am thinking more often critically about risks of substances in the near environment.

If its not necessary I would avoid it. I haven’t testet Trifluralin but a few years colchicine; in at least one case it seems I have got a tetraploid influenced chimera of a Rosa persica seedling. In species roses this method seems to produce infertile “offspring” One has to take hybrids, as I must say now.

My main reason for why I will do it this way:

1.)

The method of doubling hole chromosome sets by using N2O seems to lead to better results (in Percentage-ranges) than the using of colchicine, e.g…

2.)

Further the situation shortly after fertilization gives you much more often the chance to get a highly percentage of cell layers that are polyploidized, if at all there is a statistical chance to have a polyploidization.

Here colchicine is only usable in calli to get a similar quality of output - and before I start to breed with calli, I prefer to test the N2O way, - while it seems to be much easier in the end.

Grx!

Arno

Link: en.wikipedia.org/wiki/Callus_%28cell_biology%29

60

Off Topic and by the way: Trifluralin is a very nice molecule … look at the symmetry.

Link: en.wikipedia.org/wiki/Trifluralin