Very Easy Sterilisation Method - My Seedlings
 

:hello

This is my seeding in homemade media, with gaseous sterilization system (sodium hypochlorite fumes)


Epidendrum radicans
http://i51.tinypic.com/v45gmw.jpg


Epidendrum radicans replating with fresh medium
http://i54.tinypic.com/30m691t.jpg


Sophronitis coccinea
http://i53.tinypic.com/2edpjpx.jpg


Chysis aurea replating with fresh medium
http://i56.tinypic.com/eb7gqw.jpg


Bletilla striata alba
http://i54.tinypic.com/oiwqwh.jpg

In replating fresh medium I'm trying a supplement based on amino acids and short chain peptides (Siapton L10)

See how it works
:blushing:

Looks amazing, can you tell us about the gaseous sterilization?

I do not know if I can explain in English, but I try

is a system that prevents the sterilization of seeds and can work in non-sterile environment

I got the idea from this Protocol Brazilian http://www.scielo.br/pdf/hb/v24n2/19.pdf
Protocol explain how to disinfect the seeds with sodium hypochlorite fumes

I have adjusted the system to my needs

Prepare a sheet of plastic wrap with a hole
http://i712.photobucket.com/albums/w...orchidee/2.jpg

create an absorbent pad soaked in sodium hypochlorite
http://i712.photobucket.com/albums/w...orchidee/4.jpg

Sow without a sterile box
http://i712.photobucket.com/albums/w...orchidee/5.jpg



place the sheet with the hole on the jar and position the absorbent pad soaked in sodium hypochlorite
http://i712.photobucket.com/albums/w...orchidee/6.jpg

Cover with a sheet of plastic wrap without hole
http://i712.photobucket.com/albums/w...orchidee/7.jpg

just close
http://i712.photobucket.com/albums/w...orchidee/8.jpg


after 3 - 4 hours remove the absorbent pad by slightly raising the sheet.
http://i712.photobucket.com/albums/w...orchidee/9.jpg


close properly
http://i712.photobucket.com/albums/w...rchidee/11.jpg

cut the excess plastic
http://i712.photobucket.com/albums/w...rchidee/12.jpg

contamination 10% of jar (40 jar, 4 contamination)

I apologize for my English :Tdown:

Hello!

If I'm correct, you sow the seeds without any sterilization, then you place a plastic ( with a hole) on top, cover with a cotton pad saturated with bleach for a few hours, then remove the pad and cover right?
And you only got 10% contamination that way???

Yes is correct
no more than 5 hour, high time to fumes exposition kills seeds, or turn brown leaf of the young plant

I have never heard of that method of sterilization before, but it sounds worth a try! I'm very very surprised that you only get 10% contamination with this, that's amazing. I have some seed to flask, and I think I may try to flask some with this method, it sounds much easier than the 'normal' way of doing it.

Do you sterilize the the medium beforehand (pressure cooker for example?) Do you disinfect the seeds before or is the bleach enough to do it?

And would you mind sharing your homemade medium recipe?

Don't worry about your English, it's quite good and I had no trouble understanding!

This is very interesting. It would make flasking easy for all of us! Your english is just fine.

That's excellent. Thank you for sharing this info!

I am interested in knowing your recipe for medium as well. AND, I agree that your English is great!!! I totally understood everything, and I really like the idea of trying your methods! Thanks for sharing, and thanks for sharing the photo tutorial, it totally makes sense. I like having pictures so I know how to replicate ;)

I've never flasked, etc. before, but would like to try!

oven (150 ° C 25 minutes)

not disinfect the seeds, in fact if you go over 5 hours die fungi, bacteria and seeds

my medium is simple:

1 L RO water
1.5 g 20-20-20 NPK (Peters formula)
20 g sugar
1 g activated charcoal
5 drops NAA
5 g Agar

PH 5.8

This medium does not work with Catleya, aerangis, spathoglottis

has been tested with
Bletilla
Phalaenopsis
Doritis
Dendrobium
Sophronitis
Restrepia
Masdevallia


I'm happy if you try

There are few rules
clean all tools
do not put head and hands over the jar
prepare the absorbent pad on a blotting paper soaked in alcohol to prevent the electrostatic action of plastic. (picture 1 and 2)

I'd like to know the results :D

Thanks! I just need to find the NAA, otherwise I have everything else. I have Cattleya, Brassavola and Dendrobium seed, so I guess I'll try to make a flask of Den seeds. But I'm in the netherlands and my seeds are in france, so it will have to wait a few months.

I tried this method today. Hope the best.

well, update me about the results, positive or negative. :D

How many hours did you leave the pad



thanks

Hi Roby,

Thanks for the information, i wish i had some seed to sow id like to attempt this ;)

A question, why do you not also sterilize the seeds? Is it more hassle or will this much sterilization kill the seeds?

Thanks,
Jeff

Hi Jeff
This method is just to disinfect the seeds (see pdf)
Then I adjusted the system for seeding without boxes sterile.
For me, the classic method of disinfection of seeds and sowing is too complicated :D

Hi Roby...

I'm about to sow some Phals very soon and I will try your method! I'll keep you posted!
Thanks for sharing

Excellent! :D
The most delicate part is when you take away the pad, just lift the top sheet of plastic

Once again thanks alot Roby, this technique could potentially completely change the way we cultivate orchids in vitro - if you are the first person to use it for this purpose.

I have a question, did you harvest and pollinate the pods yourself? I have attempted to pollinate orchids alot of times but it never succeeded, any tips?

Hello Jeff9

I do pollinate and harvest seeds from my phals...
After a few try, I have better chance when:
1. I cross an old flower on a young flower ( same thing as humain no? ;-) )
2. pollinate in the morning
3. Don't remove the pollen on the mother flower
4. If selfing, flowers should be on 2 different spike
5. Orchid can be very stubborn!

Since I do that, I have almost no fail...

Helo again...

First, what do you mean by that? Second, how big is the hole in the plastic wrap?

thanks! ;-)

Hi

Let me explain with a drawing
the pad is between two sheets of plastic. one with the hole, and one is without a hole.

http://i51.tinypic.com/qrhd6s.jpg


When you remove the pad, not completely lift the sheet without a hole, but move only a small part of sheet and remove the pad.

The hole diameter is 1.5 or 2 cm

the jar volume is 100 or 250 ml

I don't believe in completely sealed flasks. Plants need gasous exchange to photosynthesize.

I did the following:

Sown the seeds in a autoclaved jar without a box, then dipped a multilayer gauze pad in Na-hipochlorite and put it on the jar and put a sheet of aluminium foil on the top then hold them with a rubber ring on the jar.

Few hours later I poked some holes on the foil with a needle to promote gasous exchange.

I can not tell if it is really necessary gas exchange, personally I never did do, but I know that many people use it.
But it is interesting to know what are the results of your method, I feel in the next seeding :D
Update me thanks.

Hello
gaseous sterilization, is not my invention. I just adapted the method to not use the box sterile

I have no tricks about pollination, which species have tried to pollinate?

LOVE IT! Thank you for this :)))))

Amazing! Thanks.

There is a problem, because you will never be able to get out chlorine gas out of the flask even of a vented flask because chlorine is 3 times heavier than air. That is why chlorine sits down to the medium and the seeds and may kill them.

True if the chlorine would last forever, but after completing his action is no longer active.
is important that the time is not more than 5 hours

Good point. But if you look at the pics of flasks in the first post, those young seedlings look pretty healthy!

So I guess the question to ask is Roby, have you successfully kept flasks until deflasking?

Hello

I do this technique from May 2010 so I never deflasking, but I think that the transplant in the new medium is fine, so I do not see obstacle to deflasking

I repeated the technique several times, both for improvement is to ensure that it was just a fluke.

In the first replanted, I killed the plants several times to find the correct time of exposure

I can only indicate the dates of sowing

photo 1) sown in June 2010

photo 2) sown in June 2010 - December 2010 replanted in new medium

photo 3) sown in May 2010 - replanted in new mediums January 2011

photo 4) sown in May 2010 replanted in new mediums January 2011

photo 5) sown in January 2011

Thanks to the cooperation of the members of my association I could try a good variety of species.
I will try soon with cymbidium, sophrotes (leptotes x sophronitis ), Angraecum.

In fact it may be that some species can not tolerate even small amounts of chlorine and could die.

but we will see this in the future :D

Thanks for your questions and thank you for your interest, this is useful to improve

Wow they grow that slow? Photo 3 is Sophronitis Coccinea right? Its almost a year old?

How much light do seedlings take to grow optimal, and how do they get light for you? Your lids of your flasks are not transparent, where does the light come from?

Yes is the Sophronitis coccinea

slow growth is due to 3 reasons
1) medium home-made
2) I should replant more often
3) Naples is very hot in late spring and summer, the species that prefer the cool stop completely and then return to vegetate in October.
I have cool in summer, otherwise the temperature goes over 35 ° C
Epidendrum radicans is much more vigorous.


14 hours of light
1200 lm Lamp - 6500 ° K - placed at about 20 cm

The lamp is at the center of a box, and jars around the lamp, the light comes from the side

Anyway, I did this when I wrote about it. Until now there is no sign of both contamination and germination.

Hello Mr. Tech
What you sow?

Pretty cool Roby. Thanks. I haven't heard of this method. I'll have to get Arditti's book one of these days as I see he tested it and suggest that this is a viable method.

For your media, how much NAA is in 5 drops? Have you tried Cattleya without it or lowering the amount?

Hello!

just did a quick search on amazon for joseph Arditi book and it is listed at $$$$ 475.00 can !!

Micropropagation of Orchids, 2 Volume Set: Amazon.ca: Joseph Arditti: Books

Hehe that's why I haven't gotten yet. I can buy a lot of stuff or waste a lot of stuff for that price. I hear it's worth every penny though. It's huge and covers every aspect of propagating orchids. Aaron Hicks' book is a bit more reasonable.

I do not know how 5 drops, I can tell you that the product is a solution of 0.01% NAA.
I have always used this dose (recommended by producer)

Hi Roby, never mind. You are using very little NAA, about 0.0005mg per liter. That should not be a problem.

Thank you Roby for the nice explaination and picture tutorials to go along with it. I'm interested to see how others do with the method. I will try this on a few other things besides orchid seeds.

What would be a most appropriate dose
I try :D

Thank you