damaclese

well market demand i say if agrastarts wants to keep are business then they should cut down the minimum order to accommodate hobbyist but i really don't think they are all that interested in small orders or they would have adapted already? besides i think we are all capable of taking care of each others Banana needs. I'm sure if we took and inventory of all the plants and supply's that every one on the org had we would find that theres more then adequate stocks to keep every body happy for years!

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rohsen

hello there you all im from amsterdam i make aliving tissue culturing plants and would like to share the following files with you, for anyone trying to micropropagate banana it is essential information!!!

here goes...

Attached Images

	medium banana shoot tip.pdf (260.7 KB, 288 views)
	protocol musa spp.pdf (881.0 KB, 247 views)

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Robert Anton Wilson

Chironex

rohsen, thank you for sending these protocols and propagation media formulations. Do you have a recommendation list of which medium to use for best results on different types of banana plants? Do you have a method for preparing the medium?

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Scot

rohsen

as im just getting started on musa spp im not sure yet, i do know some cultivars respond well to different formulations, banana is known as an easy species! what i usually do and i advice you the same is check the references given with the media, you can google this reference and see what comes up. if i cant find info i just set up trials using different media, i make my choices depending on what ingredients i have available what growth regulators (hormones or plant growth regulators PGR's for short such as 6-BAP GA3 IBA IAA NAA) required and the price of growth regulators. as far as media preparation goes i usualy buy pre-formulated media such as MS , MS gamborg, Rugini olive medium etc. only rarely do i formulate the media myself. (its a hastle) I only use agar in the initiation of cultures to screen for contaminants, after initiation i use liquid media in a temporary immersion system. any advice on media for musa basjoo ? im looking into propagating this species im also interested in ensete ventricosum

regards
Dave

in my gallerie pictures of my modest lab

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Chironex

Dave, I am also just beginning to tissue culture bananas. I use pre-measured M&S medium and add BAP for initiation and shoot proliferation. For rooting, NAA is commonly used. I have not yet tried Basjoo or E. ventricosum. I have 3 varieties in culture now as well as trials of 3 different bananas in embryo rescue. For these I want to make a modified N&N medium which is reportedly more robust for embryo rescue.
Photos of my laminar flow hood are in my gallery, too.

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Gabe15

Some banana varieties are very slow in the normal media (such as Lakatan, Fe'i bananas, some Callimusa and Australimusa species), so to help them grow I use full strength MS (instead of the normal 1/2 strength), and its also been recommended to use 30g/L sucrose, currently we are using only 20g/L for normal micropropagation. There is also an issue with different varieties reacting adversely to BA, some varieties proliferate too much and go through some type of dedifferentiation to the point where I cannot regenerate usable explants from the tissue (perhaps it is possible with somatic embryogenesis, but that would be a whole different project...), so for these bananas that have reacted poorly to the BA (E. glaucum and Fe'i varieties), I will experiment with using little or no PGRs, especially upon initiation.

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Growing bananas in Colorado, Washington, Hawaii since 2004. Commercial banana farmer, 200+ varieties.

rohsen

First of all nice to meet you guys!! Gabe, for species proliferating too fast you may want to consider using a growth retardant such as Paclobutrazol. im attaching a publication on use of this and other substances with musa. you'll find it an interesting read! as for embryogenesis .. not so much a different project as you think, very do-able especially if you have a protocol and media formulations.....! which are , naturally , attached!

I dont see to much here on temporary immersion systems, and if there ever was one technique for making loads of musa ....
I have used it succesfully with papaya pineapple and other species.

regards

Dave

Attached Images

	musa spp TIS growth retardants.pdf (5.14 MB, 124 views)
	cell suspension culture musa spp.pdf (132.3 KB, 114 views)

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Gabe15

Thanks for the papers Dave. I would love to try all of these things out one day, but I've been so busy that I don't have time to be making different media for side project this semester, I just use what our lab produces for normal micropropagation for the time being. Hopefully I will have more free time next semester and can experiment more with different media for things like embryo rescue (I use the normal microprop. media which works sometimes, but not always great), somatic embryogenesis, and these hard to culture Musa.

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Growing bananas in Colorado, Washington, Hawaii since 2004. Commercial banana farmer, 200+ varieties.

Chironex

Dave, I have read some about liquid media, but from everything I have read, it seems more expensive in terms of equipment, for the hobbyist like me. Is this the same process as developing callus culture and so on? Is there an inexpensive way to set-up this process for a home lab?
Thanks for the research articles.

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rohsen

Scot, indeed the initial cost will be higher than when using babyjars plastic jars or other disposables, however the results are amazing, and after the initial layout the returns wil justify the expense, in some species the multiplication rates will improve by 500% !!! for commercial labs these numbers are .... well you can imagine. added to this are less handling, no use of agar, and especialy in temporary immersion systems you get partially aclimatized and sturdier plants, every time you immerse the plants in liquid media you also renew the air in the culture vessel wich has considerable benefits. I distribute a TIS system in europ and south america wich includes an airpump culture vessels, the works. for 40 1 liter vessels including pump airfilters and all you need. would set you back at least 2500 euros, you would than be able to produce 10s of thousands of plants. so this is for pros, however it is possible to build a set yourself using lowcost options.
attached a paper listing lowcostoptions for developing countrys. and since a home TC lab, and indeed many commercial labs, needs to watch spending it will have some interesting options.

basicly if you can micropropagate conventionally...you can use bioreactors. you would first propagate the old way, and then seed your bioreactor with the product after you established your cultures in agar. see it as the next step towards lots and lots of plants

hope you enjoy the papers,

regards

Dave (who has no university education whatsoever started out as a hobyist)

ps. in this publication you will find in the chapter BIOREACTORS AS A LOW COST OPTION FOR TISSUE CULTURE a discription on how to build a very cheap aparatus that wil absolutly do the trick. ofcourse if anyone cares to try i will happily assist!!

Attached Images

lowcostmicroprop.pdf (1.04 MB, 228 views)

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rohsen

I love to share, i have many interesting books on micropropagation tissue culture etc.
if there is a special section of this site to do so please let me know.
for now a very good book for all people interested. Its a real knowledge booster!!!
regards

dave

please if im breaking any rules by sharing this let me know asap!!

Attached Images

Plant Propagation by Tissue Culture, vol.1 - The Background (Springer, 2007).pdf (6.85 MB, 218 views)

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rohsen

For those interested,

here's another paper i found in one of my books, it has some info on optimal media formulations for specific cultivars. interesting.

regards

Attached Images

banana - ec pua.pdf (2.67 MB, 187 views)

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griphuz

If I'm correct all TC from bananas is done from meristem tissue right? So you would have to sacrifice a sucker to go TC right?

Did anyone ever try to TC starting with just a piece of leaf?
I've heared it might be possible to re-differenciate the cells to differenciete them later on into leaf, root, or whatever...
Kind regards,
Remko.

Chironex

I have heard of being able to use male bud tissue, but have not tried it. It seems to me that I have read about someone being able to develop callous tissue from other parts of the plant, yet I have not read any research using thismeans. The article that rohsen attached "bananas" above, included some charts showing propagation results from other means - as best I can recall. If there are other places on the plant where adventitious tissue is formed, I would think that it could also be used.

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Gabe15

You do have to "sacrifice" a sucker, but if successful, you won't actually kill it and will end up with exponentially more identical plants.

Creating meristematic tissue from leaf tissue is not done, and has not been done with bananas as far as I know (one method of somatic embryogenesis). However, pieces of corm and the very bottom portions of sheath have been used during somatic embryogensis, but this is not done to micropropagate due to unacceptably high mutation rates, it is often done when trying to transform a plant through gene transfer however. Also, immature flowers (usually male, sometimes female) are used in the same fashion. Only shoot tip cultures (meristem) is normally used for mass producing banana plants.

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Growing bananas in Colorado, Washington, Hawaii since 2004. Commercial banana farmer, 200+ varieties.

Jack Daw

Once the cell has been assigned a role in the organism, it cannot be "reprogrammed, booted and reinitialized", if it were possible in such a manner, we would use that on people, to kill cancer cells for instance and force them to reinitialize properly and get a new role, which wouldn't harm the organism.

BTW - we would be talking here also about the possibility of reinitializing old cells - literally immortality key.

It would not only be the key to bioprogramming, but also to mutation processes beyond our technological levels and the only way we must go to truly understand the ways of the G .

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griphuz

Ehm, Jack,
From what I read, differenciated cells (in animals and humans) can indeed not go back to stemmcells. But in plants tissue-cells (differenciated cells) can sometimes indeed go back to meristematic cells to later form other tissues...or I must have read it wrong meaning the differenciated cells go directly to OTHER differenciated cells.
This trait is called 'totipotenci' and occurs naturally in plants when propagated vegetatively (e.g. through cuttings or layering).

This is invoked in TC, end in SOME plant species leaves, roots, or other differenciated tissue can indeed be used for TC and the mass production of plants.
Experts, please correct me if I'm wrong...
Kind regards,
Remko.

Chironex

These links should help to explain it. It will also spare you my lengthy repy.

Biology: Totipotency in plants v. little totipotency in humans, mammal cells, totipotency in plants

Home > Publications >

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griphuz

Jack & I discussed this a little over PM and the links from Scot really helped;
It seems indeed that plant cells (most tissue) is not as far differencieted as animal/human cells are, and do not have the 'on-off' switched genes. Therefore they can return so to speack to other differentiations;

"A large portion of a plant consists of cells that are not highly differentiated. These are parenchyma cells which make up most of the soft tissues and are involved mostly in food manufacture and storage. They are capable of dedifferentiation so that they can form other highly specialized tissues, even meristemetic cells which are capable of forming shoots and roots under the right circumstances. "

(From the links Scot posted)

It seems we were both a bit right;
They can form meristematic cells again, but also other differencieted tissue, without meristem in between, right?

So indeed it should be possible to mass produce a banana into TC going from just a piece of leaf, if you have the right formula to reduce the unacceptably high mutation rates e.g.
For now, If I try, I'll just usa a pup,

Kind regards,
Remko.

griphuz

Yeah, thanks dude, I think you need to be removed...