Agar mixture for plant tissue - (Jul/14/2007 )

Hi All,

I'm trying plant tissue cultivation and have, hopefully, a simple question.

I'm using powder Agar from a company called NOW. It just says that it is made from seaweed and is 100% pure.

My question is what mixture ratio of powder to water should I use?

I read a post and the person said they us a 15% Agar mixture which, makes me think I'm mixing mine way too strong.
I mixed 28 grams or two table spoons to 2 cups water.
The Agar in my dishes seems like it's to firm. Shouldn't it be more like Jello so the plant tissue can sink in some?

Thanks for any help in pointing me in the right direction.

Frank

>Edit<

Okay, I found that I should be using 7 to 8 grams per liter.
Is this what you would suggest?

http://www.carnegieinstitution.org/first_l.../preptcmed.html

Thank you,
Frank

we used 7g to prepare 1L of MS medium

Thank you Strawberry!

I'm putting saucer magnolia auxillary buds from this years growth into the agar mix. I'm pushing the bottom ends of the auxillary buds slighlty into the surface of the agar so the top of the bud is above the surface. The agar mix of 7 g per 1L looks right as the bud penetrates the surface with a little help. When placing tissue cultures into agar, do you suggest pushing the cutting all the way into the agar?

Thank you for your advice!
Frank

i'm working for the same purpose, axillary buds...
7g/L is suitable i think, so media is not hard or liquid...
i transferred my seedlings last week into magenta vessels, u need to dip them just to allow the tissue get the nutrients .....


hope this help u

Thank again you Strawberry!

Good luck with your a. buds.

Do you dip them all the way under the agar or leave part of the bud out?

Frank

i transferred the seedlings and now i'm waiting for the axillary buds to appear...they need 2 or 3 weeks to appear...but in general, i think we should dip them slightly not all the way

Hi Strawberry,

Can I ask you one more question? Sorry, I'm new at this. You said transferred the seedlings into Magenta vessels. Are you growing the plants first and then using the auxiliary buds? If so, is that so you can get "clean" tissue?

My first sad attempt:
Fungus grew on my auxiliary buds, taken from a 40 year old saucer magnolia, in only 5 days. I carefully trimmed them to remove any thing that might hold bacteria. I soaked them in a 10% chorine solution with a drop of soap for 10 minutes. I autoclaved the medium for 15 minutes under 15 lbs of pressure. The medium didn't contain any anti fungal or anti bacteria chemicals, just nutrients. I'm thinking the contamination was internal. Maybe it's too late in the year and the buds have bacteria in them.

I'll try again using different chemicals. If you can suggest any anti fungal or anti bacterial chemicals, I'd really appreciate it.

This is fun and I'm going to keep on learning and trying!
Thank you,
Frank

ampicillin and streptomycine should kill the bacteria. I am not sure about the fungi.

hi fbt,

yes, i'm starting from seeds, i am aiming to use the axillary buds of seedlings rised from seeds (not plants)....till now, there is no contamination since the seeds were already coated with fungicides and sterilized also with chlorox as usual

Thank you Strawberry and perneseblue!

Frank