Plant tissue culture - (Mar/15/2006 )

Hi,
i am working in tissue culture lab i am facing a problem that the cultures for shooting turned back and death of the leaves occours,
plz tell me the soultion of that
there are some points in my mind and these are
-- old stock solution of MS
-- old stock of cytokinins
-- using hot forceps while culturing
plz help me if anyone knows about?

Muhammad Asif

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Hi Muhammad Asif!
Your description suggests that your plantlets (they are plantlets, aren't they? ) don't die immediatly after subculture, so this can be due to a lack or a mistake in your medium composition, so it is a good idea to try to prepare your substrate with new reagents.
Besides, I suggest you to compare your protocols at least with literature, to be sure that there's any mistake (sometimes it happens... ) and take lot of care while checking growth regulators (as you already understood, plants are really sensible to concentration and choice of growth regulators).
Sometimes also gelling agents could be the reason of similar problems.
It's definitely a bad habit to use hot forceps on plant tissue, but I don't think that it will cause anything more than a local necrosis in a sane plant, although if you "burn" your stalks with hot tweezers it will be hard for your plant to survive...
Hope this helps!
Good luck!
ILA

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Hi Ila!
thanks for the reply i really apriciate ur effort. The question i coted was that in the culture for shooting first the growth is good then the older leaved die of and turned black. the remaining are green, that sort of culture appears contaminted but in fact they are not. tell me reason of that thanks.
take care
bye

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Hi!
I'm afraid that it's quite difficult for me to understand the cause of your problem without any information about plant sp. and explant type. I'm sorry but I believe I can suggest you hundreds of causes that will be at least reasonable...

However, I believe that considering the descriptions you gave, we have already identified the main factors that effect shooting in general: media composition and way of preparation, growth regulators, undelicate manipulation of explants, gelling agents...
Have you any example taken from literature? Is your plant a recalcitrant one? In this case, maybe it's not your fault: it will always take patience and a long long time to set a satisfying culture of a recalcitrant plant. And you can't take it for granted anyway...

Another thing that came into my mind is that maybe your propagation procedure could be optimized: you are correctly transferring meristem (because your plantlets are in fact growing), but maybe you don't need to transfer all that structure. This is an important factor in particular if your plant need frequent subcultures...

Good luck!
Take care too!
ILA

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HI,
Which plant u are dealing with. Is it a tree species.Than many times problem arise.It depend on the physiological condition of explant also.So pls tell the species of plnt .
beena

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Hi,

i appriciate ur effort. i am not dealing with any tree species. i am dealing with stevia, a sweet herb of peraguy. The active content of which is 200-300 times more sweeter than sugarcane.
ok

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Did you see this site? It's completely dedicated to stevia...I admit it's not strictly scientific, but seems to be inspiring...

Cheers!
ILA

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i dont know many things about plant tissue culture,
but what about culturing physical conditions??
i mean temperature, light, and moisture?
are they adjusted?

i think moisture changes as the plantlets grow, what do you think?

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