Flasking 101 - A how-to guide

[Royal]

Originally Posted by Zozzl

...and after we have a nice collection of those posts we can go back and break it down into steps so that it is very easily understood.

Hopefully, we can break flasking down and demystify the steps so anyone who reads this thread will be have no problem getting started.

Pat,

Shall we begin to break down the individual steps? We seem to have a good start. Others might chime in with their experience with certain aspects of flasking once this thread gets rolling again.

I think we'd all agree that "flasking" involves these key steps or processes (thinking very broadly):

- Obtaining or making good seed
- Choosing media and preparing flasks
- Creating a clean workspace
- Sowing - seed disinfection, sterile technique, tools, etc.
- Replating and growing on
- Deflasking and growing out

So, what do you all think? Are there any crucial steps that I missed that don't roughly fall into one of these categories? Once the "steps" are defined we can better discuss each topic at length. Does that sound good, Pat?

[Gwenchanter]

Can someone please explain more about the medium? I bought two flasks and de-flasked one of them so I know what it looks like once it's made but I don't understand how to make it or what it looks like when you buy it and how it acts when you are working with it. One of the posters mentioned that you mix it with water but I don't understand what exactly you are mixing with water? Is it a paste/ a jelly-like liquid... a powder? Secondly, how can you "sterlize it" in the pressure cooker. I can only imagine what would happen to jello in a pressure cooker and it's not pretty.

[Royal]

Originally Posted by Gwenchanter

Can someone please explain more about the medium?

Sure, we'll come back to seed.

In order to provide the right nutrients for a germinating seed, a nutrient mixture is made up. Every time we make fertilizer we are making a nutrient mix of sorts. Nutrient media is basically the same thing, just with a few other things added at different ratios depending on its purpose. The other additives can include plant growth regulators, activated charcoal, fruit powderss, etc. - but the base formula will consist mostly of fertilizer and sugar. Agar is only used to solidify the nutrient media, and contributes little other than a bit of calcium.

Mixed media are usually sold dry. If it has charcoal, it will be a black powder. If it has banana, it will be tan. If it has niether, it might be white. Some are sold with agar included, some do not have any agar added. Be sure that check before you order.

Mix with boiling water (agar needs to dissolve) and it turns into just a sticky, fertilizer scented broth. Pour into your sealable flasks, jars, petri dishes, etc. and process in the p-cooker for 15 minutes. As they cool, the agar will solidify. Let them sit for about a week to make sure they're sterile.

[Zozzl]

Hey Gwen,

I don't think it is too different than making jello Put it in a pot, heat it up and put it in the containers. Add water to bottom of pressure cooker, put in containers and see above message

Usually the media will come with instructions for preparing and it is not complicated.

[Zozzl]

That sounds great Royal Would love to hear where everyone gets their seeds and what suggestions for flasking containers.

Pat

[Royal]

The best way to get seed is to make your own. You are in control of its maturity, harvesting, and storage conditions. Another option available to us these days is the Orchid Seedbank Project. The OSP sells orchid seed packets of many species, some from the wild and some from cultivation. They are only about $4 so it's not super expensive to order a few. (OSP also sells germination and replate media, a one-stop shop.) Seedbanking is a valuable conservation tool, and proceeds go toward this noble effort.

One more option is getting some from a friend or fellow grower. Trading is fun, but some of us have seed to spare for those just starting out. Don't be afraid to ask - I mooch from OB members all the time!

An important point to note is that moisture is a seed capsule's enemy. Some harvest immature green pods, but I normally let them go until they start to split on the plant. I remove the capsule, put it in a coffee filter and set it on a bookshelf in my house. I usually snip the flower end off so that it continues to open as it dries. In a week or so, the AC has dried the capsule material completely and the comes out with a few taps. I store the seed in a cool, dry, dark, place. I've used the fridge, but only if I'm storing seed long-term. Never store fresh seed, or fresh plant material in plastic - it will grow all sorts of contaminants that may ruin the seed.

[Zozzl]

Here is a link that I mooched from another post. It has a lot of info on seed capsule maturation times,et. Worth checking out!

Lotte & Thomas Orchids

[Royal]

Pat, That's a great link. I'm going to refer to it again. OK, I'm going to move on to creating a sterile work environment. I hope others chime in with their pics and tips. Come on people!

From reading thus far, we know that we'll need to put good seed into sealable containers (flasks) that contain a nutrient media. Since this media will grow all sorts of fungi and bacteria, we go through the extra step of sterilizing them. Now, how do we get the seed into a sterile flask without introducing anything else, not even one little speck of dust? Answer - a sterile workspace. Like anything involving orchids you can get creative, but there are a few standard set-ups that people use reliably.

A Laminar Flow Hood is the most desirable option. Flow hoods work by forcing a constant flow of highly efficiently purified air (HEPA) across a workspace. The blanket of cleaned air eliminates contaminants and small eddies or air currents that can carry contaminants. These are very costly if purchased but the handy orchidist can piece one together. The Lotte and Thomas link above has a good explanation. So does OSP.

Glove boxes are the next best option (IMHO). They don't provide the freedom of movement of LFH, but they get the job done. The biggest positive is that they can be cheaply made. With a little practice you'll find what works for you. Mine is just a big clear rubbermaid tote with with a slot cut across the front and plastic kitchen wrap taped to the top. I used gloves taped into holes at first, but found that I do OK without a completely sealed box. You can still create air currents inside a sealed box, and I found that without gloves on, I can move more slowly and deliberately. For me, the benefits of control outweigh potential negatives of having an "open" glovebox. I guess it's more of a hybrid version of a glovebox and the next method(s).

Finally, many growers have had success making due without the equipment above. The Lotte and Thomas link shows a steam-bath method. It didn't work for me, but it does for some. An old AOS article discusses sowing seed under a bleach-soaked towel. I read somewhere that it has been done between two panes of glass as well. There are also quite a few growers who eliminate the need for a clean space by using a syringe. The options are endless and these stories everywhere. This is what inspired me to go with a make-shift glovebox - if people where getting good results over a steam-bath, then surely I could make a primitive glovebox work. I just move slowly and deliberately, minimize exposure of the medium, and spray a fine mist of isopropyl alcohol between flasks. All it cost me was about 8 bucks. Do a search for a flow hood and an 8 dollar glovebox will start looking pretty good.

[King_of_orchid_growing:)]

I've flasked some seeds by sterilizing the bottle using a microwave oven.

The problem was the bottles were too tall for my mini-oven. So I just put water up to 1/3 of the bottle and microwave for about 10 to 15 minutes. The water is dumped out and is left in the microwave to cool a bit.

I sterilize the caps by soaking in isopropyl alcohol. The cotton ball for covering up the vent on the lid is also soaked in alcohol.

After the medium is prepared, I pour it into the bottle. Be careful! If the temperature of the bottle is very hot and the the temperature of the just prepared medium is hot but cooler than the bottle, the bottle will break. Then I microwave the bottle with the media for about 2 to 3 minutes.

I cover up the bottle immediately with the cotton ball pressed tightly against the vent on the lid. Then into the freezer it goes for at least 24 hrs.

No contamination has occurred with this method of sterilization yet.

The culture media will not look pretty though if it matters to you all any.

[King_of_orchid_growing:)]

A thing about the bleach...

I've been told that certain orchid seeds are very sensitive to bleach.