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Post by Medina on Jul 30, 2009 15:08:56 GMT
I sown five seeds of R. gorgonia in vitro, the seeds were disinfected in 20% bleach 15 min, wash in sterile water 15 min and put in MS medium. ¡¡¡29 days later two seeds germinated!!! There are the photographs
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Post by Medina on Jul 26, 2009 14:12:59 GMT
Robet128 is the firts time that i see Nepenthes in vitro!!!. How many days the seeds germinated? Do you give it any treatment?
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Post by Medina on Jul 24, 2009 21:09:22 GMT
My plant growing in vitro has produced a flower!!! There are the photographs The plant in the flask is a cutting and it has not any root!!!.
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Post by Medina on Mar 14, 2009 19:10:48 GMT
I think that sphagnum resist the sterilization wiht bleach. I am interesting in in vitro culture of sphagnum moss, but i belive that is quiet difficult to see if the red coloration is due to nutrients or to abiotic factors.
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Post by Medina on Dec 6, 2008 23:58:58 GMT
I had used the next method for seed disinfecting with success: 1) With a piece of filter paper make an envelope( 3 cm X 3cm ), put the seeds (no more than 50) in the paper. fold it in two, fold the ends and staple it. 2) Prepare a 20% solution bleach (20 mL bleach + 80 mL water), add two drops of liquid detergent and fill a flask. 3) Put the envelope with seeds into the flask, mix and wait 10-15 minutes. 4) Now take the envelope out, with sterile forceps and put it into another flask that contain sterile water, mix and wait 15 minutes. in the photograph the flask with red lid is sterile water and the another is solution bleach 5) Take the envelope out with sterile forceps and put in a sterile petri dish. In this case I put the envelope over the lid, but it is better the petri dish. With sterile forceps and scalpel quit the staple and open the envelope, using forceps transfer each seed into the flask with medium. With this method I have germinated seeds like: D muscipula red form D. broomensis Byblis D paradoxa D ramellosa and D lusitanicum That is all, Enjoy it. --------------- ADMIN: I removed 2 images that had nothing to do with this article or CP. One of them greatly exceeded the size guidelines.
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Post by Medina on Nov 18, 2008 21:45:46 GMT
In the case of photographs no, but in the flower stalk yes, the medium was 1/3MS, 0,1 mg/L BA, 0,1mg/L NAA and 2g/L activated charcoal.
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Post by Medina on Nov 18, 2008 2:55:45 GMT
Matt I have sterilized leaves from alfalfa, violets cephallotus and juvenile flower stalk from VFT with success, followed the next procedure. 1. Choose a juvenile piece of plant(explant) which is growing and wash thoroughly in running water, next for flower stalk of VFT, dip it in bleach solution( bleach 30 mL + water 70 mL with + one or two drops of liquid detergent) for 15 min. Dip in sterile water for 10 min( I use a bottle with sterile water and then I will introduce the explant in the bottle). Now transfer the explant into flask with medium. The success depend of explant and the sterilization procedure. in general is more easy sterilize flower stalk than leaves, the concentration bleach solution is critical, for that you make try with different bleach solutions and times(usually 10 o 15 min) Bleach 10 mL + water 90 mL= 10 % Bleach solution Bleach 20 mL + water 80 mL= 20% Bleach solution Bleach 50 mL + water 50 mL= 50% Bleach solution For example, in cephallotus leaves I used a 50% bleach solution for 15 min, the explant resisted this treatment. Before I used 20% and 30% bleach solution without success. The method that you read should work, I suggest in point c) that you try with different bleach solutions beginning with 20% but no more that 50%. 2) What size cuttings do you put on the media? this is your second question. If you have a plant growing in vitro you do not need sterilize again, only to take the plant out the flask and cut it up into smaller pieces and each piece transfer into a flask, like this photographs. take the plants out Cut the plant in pieces transfer into the flasks Matt, finally I do not use BAP or IBA in this case, only 1/3 MS. Matt is quite difficult to me write in english, I will hope that you understand me.
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Post by Medina on Nov 12, 2008 2:07:50 GMT
The plants have four months in the flask, but I leave them until eight months in the flasks.
With VFT I never have used 1/2 MS. 1/3 MS I meand 1/3 of macronutrients plus full micronutrients and 30 g/L of sugar.
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Post by Medina on Nov 11, 2008 19:28:43 GMT
Very nice cultures nepaholic, congratulations is very hard introduce into culture in vitro, a leaf of plant which is growing in a pot . Some of your flask contain a blue medium, what kind of culture medium do you use? I use 1/3 MS and the VFT responds very fast, here is a photograph with VFT, I started it with a small plant and in 3 months you get many plants like this flask.
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Post by Medina on Nov 2, 2008 18:29:03 GMT
Brian the sepecie is B. liniflora and I have the plants removed out of the flask and they were laid in a mix of peat moss and sand(1:2) but in a 3 days they died. I think that is due to damping off.
How do you grow this specie? is the peat moss and sand used to grow this plant?
I have micropropagated this specie by shoot tips cultured in 1/3 MS, in a few days they produced roots and flowers in vitro.
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Post by Medina on Oct 31, 2008 21:15:03 GMT
I sown five seeds of Byblis, only two germinated. The plants grow and produced flowers in vitro, then were micro propagated and now I have many plants but I have not success in flask them Plants growing in vitro Plants flowering Many plants obtained in vitro Flasks with 3 shoot tips
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Post by Medina on Oct 31, 2008 14:08:26 GMT
Thanks a lot thwyman for your information
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Post by Medina on Oct 30, 2008 18:09:33 GMT
Thwyman, as can i see ¡¡you are an expert in culture in vitro!! for other side, does some of tuberous species produce tubers in vitro? in your opinion why are the tuberous species difficult to deflask? do you think that the tuber plays an important role in deflask?.
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Post by Medina on Oct 29, 2008 1:55:47 GMT
In this picture are the forceps and scalpel that I used And here is other seed germinating.
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Post by Medina on Oct 28, 2008 19:08:12 GMT
Thanks thwyman, are some tuberous species difficult at hardening?. I am a beginner in carnivorous plants culture and I know that deflasking in critical to plants survive. I had deflask D. muscipula, Drossera broomensis, Byblis and orchids with success.
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