Stinky's Adventures in Micropropagation...

Hey all,
I just went out and finally got a digicam with flash so I can document my latest folly.

I've decided to let my plants be guinea pigs for a go at tissue culture, which I'm just going to call TC from now in because I type really not so fast! :D

Here's what I am trying to find out in these experiments:

1) What media/additive combination works best for shoot formation in Cannabis.

2) What media/additive combination works best for rooting.

3) What part(s) of a Cannabis plant are appropriate for TC:
-Nodes
-Internodal stem segments
-Growing tip tissue
-Leaves
-Roots (I might not use roots, depends how much space I have in the incubator)

4) What sort of growth pattern do you get- Can you get a single-stemmed plant out of a mass of TC'd organisms without damaging them too much?

5) How long does this take?

I'm working on a trial media and have come up with a list of components that several different sources that all agree on:
-A gelling agent
-A sugar
-A nutrient source
-One or more growth regulating substances:
-----inositol
-----B1 Thiamin
-----NAA
-----coconut milk (seriously)
-----Kinetin
-----IBA

So I'm putting this all together.

I found Agar locally at a Chinese market as an ingredient in a dessert mix... but it was almond-flavored and already had the sugar added, so no control over that.
GNC stores carry powdered inositol and Thiamin.
Coconut milk comes from coconuts.
NAA and IBA are found in rooting agents but I want them individuially and might have to go to a biological supply store for that.
I don't know where in the heck I'm going to easily find kinetin!

Keep ya posted.

This will definitely be a Favorite thread.

So far, 1st place in the Science Fair! :)

i agree you know your stuff.... good luck ill wait for futher posts definatly

Still trying to assemble ingredients for my media. Thank you for your patience!

Although I do have my tools lined up:

Containers will be half-pint jelly jars, because they're the right size and can take autoclaving.

My sterile cabinet will be a clear plastic rubbermaid-type tote with hand-holes cut in it.

I've got access to a nice analytical balance accurate to 0.0001 gram :D yay.

Sterilizing agents will be bleach spray and a spirit lamp for flaming off tools.

The autoclave is a beefy old pressure cooker that has a HUGE capacity, but it's at work and I'm still hunting around salvation army locations for a cheap canning processor.

I've narrowed my plant-parts choices to leaf segments and nodes.
We all know that nodes are the source of good totipotent cells when it comes to cloning.
But I want to try leaves as well because obviously the plant has plenty of leaves and not so many 'extra' nodes. Leaf sections will be 1cm square and placed cuticle-side down on the medium so the stomata can still breathe.

Are you making any progress, Stink?

Yeah, I FINALLY found a product that contains cytokinin.
It's Bonide Tomato Blossom Set Spray.
So add that to my supplies-on-hand list.

Sticky,

This shit is a little over my head but I'm watching you.

Loco

It'll be a while. I had to put this project on the back burner. The new autoclave we're ordering at work hasn't arrived yet.

Stinky does that make you a tatto artist?

Nope, a chemist.