Willy's TC Journey

Discussion in 'Tissue Culture' started by WillyCKH, Sep 28, 2017.

  1. WillyCKH

    WillyCKH CPSC and PotM Moderator Staff Member

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    Hello all,
    I'll be posting updates of my LFH and photos of my cultures here :)
    Hope you enjoy this thread!
    Willy

    Drosophyllum lusitanicum culture, seed grown, 10 days old:
    DSC02007.jpg
     
  2. WillyCKH

    WillyCKH CPSC and PotM Moderator Staff Member

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    I've forgotten to mention that this thread is a continuation of my thread in the old forum here. You can check out my LFH and upgrades, if you're interested :)

    Here's a photo of the same Byblis liniflora culture, 30 days vs 38 days, can you spot the difference?
    This is a selected clone that grows the fastest! (well, I have two to compare, haha)
    I love how I don't have to worry about watering it inside the container. This is great for a lazy person like me.

    DSC01957.jpg DSC02066.jpg
     
  3. Samskwatch

    Samskwatch Carnivorous Plant Addict

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    I definitely can tell the difference. Willy, you ROCK!!!
     
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  4. housepet

    housepet Carnivore

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    Nice work Willy, it seems that you won the battle over sterilization and produced a nice gel for growing. I dont have any photos yet but hope to get something posted on my Dionaea Muscipula eventally. So far, I managed several clean cultures in baby food jars.
    My recipes are:

    A. For One litre Seed media -no auxins or hormones:
    Gelzan powder 3.0 g
    1/3MS@4.43/3=
    1.48g
    Sugar = 30g
    PPM-preservative= 1.0g
    PH

    Note*
    Without adjustment, this mix measured exactly to 5.1 ph
    Adjustment is needed in order to rech the desired pre-autoclave ph of 5.5


    For One litre of Explant media add the to the above:

    A. 1g kinetin-In order to promote callus in leaf and stems.

    B. 1g IBA3-In order to promote growth of roots
     
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  5. Dogrem

    Dogrem Carnivorous Plant Addict

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    Superbe.

    I'm always impressed about your in vitro ;)
     
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  6. WillyCKH

    WillyCKH CPSC and PotM Moderator Staff Member

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    @Dogrem, thanks! Most people refer this to TC, but I guess the correct term is in vitro, haha. :)

    @housepet
    Thanks for the recipe!
    This is mine (haven't added anything else yet, but I bought some BAP):
    1/3 MS salts
    30g table sugar
    1/8 tablet of multi-vitamin tablet with thiamine
    6g food grade agar agar

    On a side note, I usually mix half a liter at a time.
     
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  7. WillyCKH

    WillyCKH CPSC and PotM Moderator Staff Member

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    It takes good patience and a number of tries before something cool happens.
    Right now, I think the success rate for starting a clean culture is about 70-80%.
    I love it when something cool happens though!
    Here's the Drosophyllum culture and two Drosera cultures.
    DSC02095.jpg DSC02091.jpg DSC02090.jpg

    I'm still working on Darlingtonia and Sarracenia seeds, they tend to be either burnt or not completely sterilized. Anyone has tips to offer? :)
     
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  8. Lloyd Gordon

    Lloyd Gordon Parasitic Plant Aficionado Staff Member

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    I found Sarr. seed fairly easy in vitro.
     
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  9. WillyCKH

    WillyCKH CPSC and PotM Moderator Staff Member

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    Do you mind sharing your tricks? Do you use bleach? :)
     
  10. Lloyd Gordon

    Lloyd Gordon Parasitic Plant Aficionado Staff Member

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    It's been so long since I did Sarr's. I may have used a sterilizing solution of PPM, followed by PPM in the media. Possibly H2O2, then isopropanol.
     
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  11. housepet

    housepet Carnivore

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    Nice work Willy, it seems that you won the battle over sterilization and produced a nice gel for growing. I dont have any photos yet but hope to get something posted on my Dionaea Muscipula eventally. So far, I managed several clean cultures in baby food jars.
    My recipes are:

    A. For One litre Seed media -no auxins or hormones:
    Gelzan powder 3.0 g
    1/3MS@4.43/3=
    1.48g
    Sugar = 30g
    PPM-preservative= 1.0g
    PH

    Note*
    Without adjustment, this mix measured exactly to 5.1 ph
    Adjustment is needed in order to rech the desired pre-autoclave ph of 5.5


    For One litre of Explant media add the to the above:

    A. 1g kinetin-In order to promote callus in leaf and stems.

    B. 1g IBA3-In order to promote growth of roots


    And finally the photos:
     

    Attached Files:

    Last edited: Oct 7, 2017
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  12. WillyCKH

    WillyCKH CPSC and PotM Moderator Staff Member

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    Great work, Kent!
    Where did you buy your PPM? It is so expensive..
    I adjust my ph to 5.8 before I put the jars into the pressure cooker.
     
  13. Dogrem

    Dogrem Carnivorous Plant Addict

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    I just want to tell you that I have a great respect for you guys...
    That's so complicated !
     
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  14. housepet

    housepet Carnivore

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    Thanks Willy, At first I had quite a time finding a source but I believe you can still get 30 grams for roughly 30 dollars plus shipping at 35 dollars from:


    Terri Wichtendahl

    Controller

    Plan Cell Technology

    1823 Jefferson Place, NW

    Washington, DC 20036

    Telephone: 202-621-5494

    http://www.plantcelltechnology.com/ppm-store/

    It sells for roughly a dollar per gram, so the small bottle should be enough for 30 litres.
    I also bought a 50ml bottle since then but cant recall who sold it.
     
  15. housepet

    housepet Carnivore

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    Your recipe looks like a few others Ive seen except for the multi-vitamin tablet.
    Whats that about and does it work?
    I read elsewhere, Dionaea seem to do better on Kinetin and Sarracenia do better on BAP.
    For seed cultures, I am staying with ph of 5.5.
    After a few months of trying, I still havent had any callus from leaves.
    I still dont know the reason for that.
    Perhaps I should have began them on kinetin and without any IBA.
     
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  16. WillyCKH

    WillyCKH CPSC and PotM Moderator Staff Member

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    I've only bought BAP to play with. I haven't added any to my cultures yet, I want to get them growing into decent sizes before inducing the multiplication.
    Here are some photos from my mini-lab:
    Some Drosera seedlings...
    DSC02179.jpg DSC02180.jpg
    Byblis
    DSC02182.jpg DSC02184.jpg
    A transferred culture, hopefully nothing goes wrong!
    DSC02186.jpg
    Sarracenia seed!
    DSC02188.jpg
     
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  17. housepet

    housepet Carnivore

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    I havent tried anything other than Dionaea so far because my big problem is always fungus on the tips of ebay seed even after 10 percent bleach for ten minutes. The only change lately is that I just started using tween 20 in the bleach bath before rinsing with tap water 5 minutes through a tea strainer. As an added precaution, I let them soak a few hours in sterile distilled water before sucking them out with a plastic pipette and then dropping them into jars. Even after all that, I will only get a few clean sprouts from each pack. My best average of 5 seeds without fungus came from an ebayer in Tisdale Sask.
     
  18. housepet

    housepet Carnivore

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    You had me fooled Willy. Judging by the amount of roots in this batch, I was certain you were using some magic formula.
    Have you had any success from cuttings?
     
  19. housepet

    housepet Carnivore

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    For the 1/8 tablet, which vitamin is this?
     
  20. WillyCKH

    WillyCKH CPSC and PotM Moderator Staff Member

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    I have a few ex-plant cultures just made few days ago. Let's hope that something will happen (beside fluffy fungus growing).
    I am just using the formula that I've written previously. I will be adding BAP later.
    The vitamin tablet is called Multi Vitamin, the one per day ones you see in drug stores. If you are buying some, make sure to check for thiamine (B1), I've read that it's important to have it in the tablet.
     

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