MurCurY

What size plant can you start with for an explant for TC?

I have a few plants inside for the winter. My girlfriend noticed a sport popping up next to the main stalk...couple days later....another. So....I want to use one...or both...of these pups as explants for tc. The variety is Rajapuri.

http://www.bananas.org/gallery/uploa...plants_015.JPG

http://www.bananas.org/gallery/uploa...plants_013.JPG

Chironex

Generally, I ask for them to be at least 16" tall, I have done smaller, but dividing the tissue becomes more difficult.



For reference, this is a baby food jar cap.

__________________
Scot

MurCurY

wow that large!

At the lab i work at it's all about the smallest piece possible. would it be possible to take a pic of a "starting plant" and how much you actually use?

Gabe15

The bigger the plant, the easier it is to keep it sterile. With a large plant, you can do surface sterilization multiple times while removing layers from around the meristem, but with a plant as small as the ones you want to use, you would have to get it very sterile on the first try and would probably be able to only remove one layer.

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

Chironex

There is a photo in my gallery.

__________________
Scot

Bananaman88

MurCurY,

I'm just trying to make sure that I understand you. When you say "sport" do you mean an actual sport where what you see coming up is distinctively different from the parent plant or do you simply mean a "pup"? Thanks for the clarification.

MurCurY

i found the post with that pic chironex....but when i went to your gallery....couldn't find it.

I have a cavendish super dwarf that's at leaf 12/13 on 2 pups....the main plant died off a while ago. Maybe i'll sacrifice one of these plants and give it a try.

MurCurY

a pup....


and chironex...that tip in that baby food jar lid came from a 16" or so plant?

How hard is it to find? are there layers leading to the tip....like an onion?

Chironex

It is similar to an onion, yes. The closer you get to this tissue, the more compact it becomes. Don't peel away too much before you sterilize the surface though. As Gabe has said, you will lose some of the outer layers of tissue in the process.

__________________
Scot

MurCurY

and you use standard MS media......right?
any hormones?

Chironex

As Gabe mentioned before, we generally use 1/2 strength M&S medium, with BA or BAP during initiation. I have been experimenting with other media too, including kinetin and activated charcoal. Although, these are primarily to promote embryo rescue.

Then no hormones for rooting, or some use NAA, IBA or IAA (auxins) but I haven't tried them yet, as all of my cultures are still in stage 2 sub-culturing.

__________________
Scot

MurCurY

1/2 strength MS, with what concentrations of BAP and...do you not use NAA in the initial mix?

Chironex

Generally, 1 mg/ml is adequate although I have seen much lower concentrations of BAP. I believe that this will vary based upon which banana cultivar is being cultured.

I do not use NAA until the rooting phase as it is a rooting auxin. Some add no NAA and just leave out the plant growth regulators altogether in phase 3 - rooting. Try using some in a few cultures and none in others until you find which one works best for your banana plant selection.

__________________
Scot

MurCurY

so....1mg per liter of agar? I'm using BAP dissolved in dmso.

Chironex

No, 1 mg/ml in water. If you are dissolving it in DMSO use about 25 mg/ml. You will use about 5 mg of this BAP solution in your agar.
Note: It's quite alkaline, so be sure to adjust your pH.
This link might be of help to you: Micropropagation of Musa

__________________
Scot

MurCurY

excellent site!

One last question....for the moment....
He lists gelrite as his "stiffener"
What amount of agar would you use for musa?

Chironex

I use about 5-7 grams per vessel. I put about 3 tablespoons of medium per vessel and add 5-7 grams of agar. (This is about a third of a teaspoon of agar per vessel.) Gelrite is used instead of agar by some people. I like it more because it is colorless.

__________________
Scot

MurCurY

i've been really sick past few days. i'm feeling way better today so....i think i'm going to make up my agar and start disinfecting this afternoon/evening.

5-7g of agar in approx. 40ml of nutrient.......isn't that rock hard? when i make agar for my mushrooms i use 20g/l and it's firm enough i can't press into it but my finger makes a print when pressed.

I already mixed my 6-BA at 1mg/ml in dmso and do not have an accurate enough scale at home to add up the mix...my work lets me use their lab scale and i've been out sick the past few days.
Would using 5ml of dmso in a liter of agar be harmful to the explant? The lab manager says she "thinks a little helps the plant take up stuff better"....although i don't really know. and i've never asked her if there's an amount at which it might be detrimental. I do know if you touch it...your mouth tastes kind of like garlic in seconds.

I have some strawberry plants from work i might try to tc while i'm at it.

Chironex

You can use less agar if you wish, I was going from memory and I use a pre-sized measuring spoon when I use agar. I use 7 g of gelrite.
Not sure about DMSO as I have not used it. Good luck.

__________________
Scot

MurCurY

thank you very much! pictures to follow.