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Ex vitro rooting and ex vitro acclimation by using the 'float hydroponics' technique - Alexfiraontheweb

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Ex vitro rooting and ex vitro acclimation by using the 'float hydroponics' technique
Introduction. Ex vitro acclimation is an essential phase in the process of micropropagation, as it allows in vitro - cultured plants to survive in normal, natural conditions. Ex vitro acclimation is the gradual adaptation of in vitro-cultured plants to the normal, natural environment. The usual, classical methods used for ex vitro acclimation involve the use of various types of trays or pots filled with a solid substrate (peat, perlite, vermiculite, sterilized soil or a mixture of some of these components). Acclimation is done in a more or less confined environment (greenhouse, tunnel etc.) in which humidity is maintained artificially either by artificial fog generated using a special installation or by covering the cultures with a transparent lid that retains humid air around the explants, such preventing them from drying out. Rooting of in vitro-cultured plantlets can be done either in vitro on media containing either auxins, e.g indolebutyric acid (IBA) or no growth regulators at all, or it can be done, depending very much on the species and cultivar, directly ex-vitro in the potting mix used for acclimation. Such, rooting and acclimation are done in the same phase. For some species this can be done, for others it is still impossible. At the Fruit Research Station of Cluj (www.scdpcluj.ro) a novel ex-vitro acclimation technique was set up by Alexandru Fira (myself) (email: firaalexandru@yahoo.com) and eng. Doina Clapa, PhD (email: doinaclapa@yahoo.com), based on hydroculture. We started research on this topic in August 2008. This technique was mainly applied for the acclimation of plantlets already rooted in vitro. Our method consists of culturing plantlets rooted in vitro and transferred exvitro in vessels containing a thin layer of ordinary tap water without any fertilizer and plant growth regulator. Information about this acclimation technique is provided on this website: https://sites.google.com/site/alexfirasplaceontheweb/new-ex-vitroacclimation-method Here is another movie of ex vitro rooted and acclimated blackberry on a floating cell tray: http://www.youtube.com/watch?v=CGBwI_ok1cA The use of a modified method of 'float hydroponics' for ex-vitro rooting and acclimation was inspired by the description of the 'float hydroponics' method used for vegetables and tobacco. Some info about float hydroponics is presented on these websites: http://edis.ifas.ufl.edu/hs184 http://www.fshs.org/Proceedings/Password%20Protected/1999%20Vol.%20112/313315%20(TYSON).pdf

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Ex vitro rooting and ex vitro acclimation by using the 'float hydroponics' technique - Alexfiraontheweb

http://www.simplyhydro.com/free2.htm

http://aesop.rutgers.edu/~horteng/floating_hydroponics.htm http://www.uky.edu/Ag/NewCrops/brassicamq_tw.htm There is also an impressive video on Youtube on this topic: http://www.youtube.com/watch?v=FHBhyqowSEc Using float hydroponics for ex-vitro acclimation and rooting was also inspired by a Bulgarian colleague and by one disadvantage of the hydroculture acclimation method: the need to replace and resupply the water in the acclimation vessels. By using float hydroponics one batch of water can be used during the entire rooting and acclimation period (1-1.5 month). A description of our method of using float hydroponics for the ex vitro rooting and acclimation of in vitro-propagated plants For ex-vitro rooting, take out the plantlets grown on the multiplication media, in the multiplication phase. These are shoot clumps (miniature bushes) proliferated on the multiplcation medium under the effect of cytokinins. Wash the bases of the clumps in order to remove any remnants of the nutritive medium. Cut off the base of the shoot clumps with a pair of sharp scissors in order to obtain a bunch of separate, individual shoots similar to a fascia. If the bunch is too thick separate it into smaller bunches. Then, plant each small bunch into cells in an ordinary cell tray and set the cell tray to float on the surface of the ordinary tap water in a basin in such a way that the bunches of shoots are partially immersed in the water. Most of the shoots should be well rooted in about 1 month. There are no special requirements of air humidity during the whole acclimation period, the cultures can even stand in open air ! Some species like some rose cultivars and the blackberry can withstand bad conditions. Well-rooted shoots should then be planted into pots containing potting mix and grown further in usual conditions. Dead, moulded or rotten shoots (there are very few of them) should be removed. Living but non-rooted shoots should be placed back into the water for rooting. Here there is a new video showing my colleagues transferring blackberry ex vitro for rooting and acclimation: http://www.youtube.com/watch?v=7KNc_VQyzgM

Here there is a video showing some blackberry rooted and acclimated ex vitro in float hydroponics: http://www.youtube.com/watch?v=cXQDgWsTg7w Here there is another video of a tray full of blackberry shoots, rooted and acclimated: http://www.youtube.com/watch?v=CGBwI_ok1cA http://www.youtube.com/watch?v=zlvxVvAtRRo&feature=related For the ex-vitro acclimation of in-vitro-rooted plants transfer ex-vitro
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Ex vitro rooting and ex vitro acclimation by using the 'float hydroponics' technique - Alexfiraontheweb

the plants from the rooting phase, wash the roots with warm tap water in order to remove the remnants of medium and plant one or more plants into each cell in the cell tray and set the tray to float in a basin containing ordinary tap water. There is no need for special conditions of humidity. The plants should be acclimated in about 1 month of culture and they should be planted into pots containing potting mix. Below there is a video showing some acclimated Drosera capillaris. The tray also contains black currant greenwood cuttings: http://www.youtube.com/watch?v=6XRx66sod0U Acclimation in hydroculture was a great succcess for Nephrolepis, also: http://www.youtube.com/watch?v=kx915TmpQFc Below I am going to show some pictures of plantlets either rooted ex-vitro in the floating tray or rooted in vitro and acclimated in the floating tray. Blackberry in the floating cell tray:

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Ex vitro rooting and ex vitro acclimation by using the 'float hydroponics' technique - Alexfiraontheweb

Rose cv. The Fairy rooted in vitro and acclimated in the tray:

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Ex vitro rooting and ex vitro acclimation by using the 'float hydroponics' technique - Alexfiraontheweb

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Kalanchoe blossfeldiana acclimated in the floating cell tray:

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A thornless blackberry variety rooted ex-vitro in the floating cell tray:

Blackberry cv. Thornless Evergreen rooted ex-vitro in the floating cell tray:
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Ex vitro rooting and ex vitro acclimation by using the 'float hydroponics' technique - Alexfiraontheweb

A thorny blackberry variety rooted ex-vitro in floatation hydroculture:

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Ex vitro rooting and ex vitro acclimation by using the 'float hydroponics' technique - Alexfiraontheweb

Images of roots grown into the auxiliary polystyrene floats:

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Ex vitro rooting and ex vitro acclimation by using the 'float hydroponics' technique - Alexfiraontheweb

I wish you success with applying this ex-vitro rooting technique !

Alexandru Fira

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