jarceleste

Hello.

I experienced to make my 1rst tissue culture of a canna plant.

I read many papers of tissue culture before starting the experiment.
On July 26 I made my first experience with a canna indica, but failed.

After 15 days there are no leaves and began to appear fungi.

I used the culture medium Murashige & Skoog with agar
Modified with Adenine,Inositol, and Thiamine
(With macro- and micronutrients, 80mg/L adenine sulfate, 100mg/L myo-inositol, and 0.40mg/L thiamine HCl)

I bought this medium in Caissonlabs at this link http://www.caissonlabs.com/formpdf.php?product=79

Please help me if I'm making some error or if is missing something.

Thank you!
Miguel

jmoore

I suspect it didn't work because you had contamination. Did you use the correct part of the meristem? Did you follow the method correctly? Did it say use that type of medium? You say you read many papers on this subject, try and repeat everything that you read, they should list reagents and method in scientific papers. If you follow what they did and it didn't work then it's your aseptic technique that's at fault.

jarceleste

hello,

Thank's for your reply.

I read some manuals of Tissue culture, but in general, not to a particular specie.

My question is if this ms medium is good to that tissue culture or it was missing something, like hormones or regulators.

About the correct part of meristem, is there some information about that?

Thank's for your help.

Miguel

jarceleste

any ideas???

adrift

It doesn't matter what hormones or media you use if you have contamination. If the fungus or whatever is at the site of your meristem, then that was likely what brought the contamination. Examine your technique for processing it. If the fungus began away from the explant, then your general technique is bad.


If you have general overall contamination, you can practice without any explant or hormones. Make up your media and put in dishes. Keep them under the conditions you will be keeping your TCs for several weeks, watching for contamination. Don't bother making up jars / dishes with hormones and explants until you have your sterile technique worked out. Maybe under 10% contamination.

Did you use any preservative in the media (such as PPM)? Can you afford to do so?

Did you do it in open air or in a box? Can you make a box, even if it doesn't have airflow?

Have you practiced simply opening your jar or dish with one hand, keeping the lid over the top to avoid anything falling in, and putting the explant in with the other hand?

__________________
I've got to stop asking people, "How stupid can you be." People are taking it as a challenge.

Frankco

Hi Miguel,

For multiplication you should use between 3mg/L and 5 m/L of BAP depending on what type of banana you are trying to tissue culture. Your other ingredients look fine. Just wondering how much sugar you are using. 30 grams to 40 grams per liter is what you should add.

If you do not have a flow hood or don't trust your clean area, do the cutting under a weak solution of bleach after you disinfect the cutting in a strong solution of bleach (strong solution = 1 part bleach to 4 parts water). For the weak solution you can use 1 part bleach to 20 parts water.

Trim the plant under the solution of bleach and then rinse with autoclaved distilled water. As long as the piece of banana is under the weak solution of bleach, you can trim it anywhere, kitchen table, desk, etc. just don't take too long to do it. The transfer into the autoclaved water should be done in a clean box. Since there will be a slight coating of bleach on the plant during transfer your risk of contamination will be small if you do it quickly.

Here is a video that shows you what to use for your explant.

__________________
Frankco
Zone 5
N.W. Indiana

alah

Hi Frankco. Please excuse me for deleting part ( OK, almost all ) of your reply.
I´m sure we´ll understand each other better this way.

At the end this guy takes out a shoot? for subculture?
He puts two pieces of explants into the same vessel.
Can you explain that?

"Place this material on it's side and carefully trim the associated corm
so as to reduce the amount present "without damaging" the meristem."

This is from KGBenson´s article.
Can´t I quarter this explant?

Thanks, Alberto.

novisyatria

dear all
I want share my experience about tissue culture banana. like Francko, I use MS + 5 mg BAP/l and add 40 gram sugar. sometime for certain variety using 10 mg BAP/l. tissue culture bananas is easy enough. for rooting samoetine auxib is not necessary because depend of my experience, without auxin such as IBA or NAA, bananas explan can growing root.

__________________
Novi Syatria
Lab of Tissue Culture , Dept of Forestry, Faculty of Agriculture, Univercity of Bengkulu, Indonesia www.kuljar.com email: novi_syatria@yahoo.com