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Flasking -101

This is a discussion on Flasking -101 within the Flasking Equipment & Technique forums, part of the Orchid Propagation category; So far so good: These are tiny TINY baby orchid embryos (20x-30x magnification) C. cristata ...

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  1. #21
    mycologist's Avatar
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    So far so good:

    These are tiny TINY baby orchid embryos (20x-30x magnification)

    C. cristata


    D. antennatum (you can kind of see the lines of cells)


    I lost three of the flasks (of 9) to seed borne contamination the first round, but all of the species still have at least one clean flask and they have all germinated. For the rest I used a sugar pre-soak to encourage spore germination prior to sterilization. For those, only 1/12 was contaminated. I wish I had used clearer tubes, but hopefully they will get enough light until I replate.


    Quote Originally Posted by kfir from israel View Post
    ho boy
    i just saw this thrid now!!!!!
    cool lab man!!!!!
    i use the same lab-welcome to the war against fungus!!!!-most with seeds that dosent come from green pods!!!!!
    i cut the area with hot sterelized knife the fungus and put some oxigen\bleech\or any solusion that starilized for 12-20 hours and suck it back and hope that i did good
    i dont have 100% luck but somtimes it works
    u must cut abit from the good agar that didnt got infected too - to make shure for more luck-and i say luck,cause fungus spread the spores and somtimes the spores infect the rest of the petrry
    good luck
    kfir
    Kfir,
    Thanks for the info! I just discarded the contaminated tubes since I had made enough replicates. For plants that are further along, I have read that you can sterilize them (dip in 3% peroxide IIRC) and replate them onto clean agar. Pre-soaking in sugar solution made a huge difference! BTW, the fungi in the first set of photos are symbiotic and were inoculated on purpose.

    Quote Originally Posted by Ania View Post
    This looks like a lot of scientific fun. You put together a nice tutorial.



    I have a Rhyncholaelia digbyana seed pot to give away to ANYBODY who would like to try growing orchids from seed. It just crocked open and looks like its full of seeds. PM me if you want it!

    Ania

    Ps. You can search for my post if you want to see it in flower.
    Ania,

    It looks very nice! I would be interested but I haven't worked with any pods yet. If no one else takes them let me know.

    Quote Originally Posted by Feed_me_Seymour View Post
    What exactly is your agar mix? Is it simply activated charcoal and agar? Also if you add a little H2OH to the mix it will keep comtams down a bit. 14.9mL solution to 1-1.5mL of household hydrogen peroxide works. (For fungi at any rate) the idea is to keep spores from being able to germinate. I wonder if this trick could work for the task at hand... Did you experiment with agar mixes at all?
    The media contains everything including the kitchen sink. I haven't experimented with media - the formula worked for all six of the species I tried. There is a lot of literature and work already done so I will rely on that until I meet something stubborn. They also have good Cyp. media I will try next. If you go to the website for the company you can get the info sheet from the product number and it will tell you everything that is in it.

    I haven't ever used peroxide in media (but use it for seed sterilization regularly). I don't see how it would work because it breaks down to water when exposed to oxygen in the media. With the opposite approach using a sugar pre-soak to germinate the spores and make them vulnerable prior to sterilization contamination was minimal.

  2. #22
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    Update - the majority of the seeds didn't germinate. After referring to a book that shows the development of the embryos I now realize the pic I posted above is an ungerminated seed. I have some green protocorms in only one flask.

    I picked up a bunch more fresher seeds and will try again soon with the tropical ones.

    I ended up with a several green pods of local Cyp. acaule and started some mother flasks last week - no contamination so far so wish me luck. Of course it had to be the most difficult one

  3. #23
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    Good luck with the Cyp. acaule! That will be nice.

    Cheers,
    BD

  4. #24
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    This is a really cool tutorial. I just harvested a phal amabilis and sent it off. How cool to be able to do this yourself.

    Although I must admit it is a bit too techie for my little brain.

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    Hey just bumping this thread cuz I'm curious to know what happened!!!

  6. #26
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    Hi Myco,

    This is my first peek at your thread - I haven't been around much the last couple of years. But Aaron Tester [aka, ATester] was a guru at flasking media for differerent orchid genera.

    He and I did a lab flasking of a Cattleya cross I'd made. He was the flasking guy, I had the seeds and access to a lab at Dartmouth College.

    The seedlings did fine, all flasks, until they were nearly ready to replate. Then he had a power outage for a couple of days, and then the flasks seemed to succomb to a brown-rot and all croaked over the next month or two.

    He's not on the forum anymore, and I can't reach him. I know he was undergoing a lifestyle change, so neglect might have been the cause. I simply don't know. But after successful germination and growth, it was a bummer to lose the new cross (crossed in both directions).

    The thread is still probably in the archives, but I can't find it!

    McJulie

  7. #27
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    I'm glad I stopped back in today -

    I ended up with one tube worth of Cymb and was jsut thinking I should replate it and take some pics. I have a break coming up so I should get to it soon. The Cyp. acaule must have already had inhibitors (should have harvested greener) but they are clean and sitting in the fridge for the last month and a half (I ended up buying a bareroot C. calceolus for the garden for now).

    McJulie - I remeber the thread well. I was so excited when reading it and was sure it was going to come out well, then the bad news hit. They must have gotten cold because I know I have been guilty of ignoring them for some time with no apparent ill effects. You should try again! I got some more stuff from the OSB but I would love to try some fresh seeds.

  8. #28
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    Hi Myco,

    Cold could have been the problem. I know there was enough other strife in Aaron's life at the time that the "children" weren't getting all the hoped for attention. He's AWOL now, so it's as if he caught the brown-rot too.

    It was fun to flask them, but the strain on the pod parent did do her in, and I was sorry to lose that plant!

    NOTE TO FORUM: Don't try setting a pod on anything but a vigorous, mature plant, and never try and pollinate more than one flower at a time. It sometimes works, but more often the plant won't survive the strain of bringing a pod to full term.

    McBummed

  9. #29
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    I got the Cyp. acaule to germinate.

    I've also had some better luck with the newer seeds for the epiphytes and have been working on it a bit more with a couple of students. I think there are around 6 species that have germinated now. Here are some that just germinated (maybe the added a few too many seeds in that pile, it looks like a green blob but is a bunch of tiny plants) and some that were being replated. Brian is going to do a bunch more this semester.




  10. #30
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    WOW That is awesome!!! Congratulations!

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