Hi Hildegard, Sorry to hear you are still having problems. I am only able to guess - as you have not let us know which genera you are working with. I understand many terrestial South African genera come from acid nutrient poor habitats, which may have a bearing on your observations.
Aaron's suggestions to try vaious combinations is a good one and only your lab will be able to directly assess the results for your conditions and genera. Generally in-vitro seedlings modify the pH of the media they are growing in. Your terrestials may be doing this and still grow well. It does not always follow that a reduced pH will mean your plants die, especially if adapted to an acid habitat. There may well be exudates from your roots that are detrimentally modifying you media. Perhaps you could use larger containers with more media for more diffusion of the problem 'substances'. You plants may be simply reacting to transplant shock by producing exudates into the media on the second transplant. Some tc grade charcoal is acid washed. You should try and obtain neutralized chacoal or test the acidity from DI water washed prior to media addition to see if this is the problem. It is important to determine when the pH change occurs, is it after autoclaving, after replating or when the plants die? Almost all rooting hormones in excess can cause root inhibition, this is commonly seen in conventional cutting propagation. In-vitro, this is more critical. Low concentrations allowing root formation but high concentrations showing a charecteristic growth response of lack of roots and callus development. Also, IBA does not always produce the same results as NAA. Perhaps you could explore this. I know from my own in-vitro work with root modification of Gerbera and ornamental grasses that IBA produce branched short roots while NAA produce longer single roots! Terrestial orchids generally produce a good root system so perhaps you could culture your plants without any rooting hormone? Retuning to the subject of possible modification of the media making it toxic to plant growth.....this could be a result of any number of causes. High on the list would be to look at your in-vitro venting environment. It has been reported to me that other terrestials such as some species of Cypripedium react according to the rate of gas exchange by producing roots that either exit the media or dive deep into it. Your whole in-vitro environment is important. As for banana, as a complex additive, it does suffer the major problem that it varies from batch to batch or cultivar. (The same is true with Coconut water or milk, it is more a research tool rather than a commercial viable option). If you use banana you will almost certainly need to add charcoal - so you may be going around in circles! Banana is rich in cytokinins and these are generally inhibitory to roots, so if you remove your banana source you may be able to improve your roots. Both charcoal and banana attract a lot of myths and for many genera they are not essential. Millions of orchids are produced in temperate climates without a single banana being pulped, coconut drained or flasks being blackened with charcoal. Perhaps your genera can survive without charcoal and banana. Good luck. Alan L Winthrop http://www.tissuequickplantlabs.com > > Date: Tue, 13 Jul 2004 19:34:28 +0200 > From: "Hildegard Crous" <[EMAIL PROTECTED]> > To: <[EMAIL PROTECTED]> > Subject: [OGD] terrestrial orchids in vitro >Message: 4 > > Hi All, > perhaps someone out there could throw some light on this matter? > I get excellent root development on terrestrial orchids if the plants are > initially on a clear medium, (1/2 MS) and then replated onto the same medium > but containing charcoal. However, with the follow up replate, (charcoal added > to the medium), the plants lose colour, (pH drops and leaves turn white, ) >> regards Hildegard > ------------------------------
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