Deflasking of Cypripedium seedlings - Cypripedium.de

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How to raise in vitro propagated Cypripedium seedlings to fully established hardy plants for the garden © Camiel F. de Jong, December 2004 climate in North-Western Europe. In these cases also the prevailing circumstances are mentioned so the reader is able to adjust to his or her own climate. Dormancy To survive the cold winter, the shoots of Cypripediums die down in autumn after having produced a new dormant bud. When it’s not freezing, roots continue to grow whereas the bud stays dormant. The dormancy of the bud is only broken after a three month cold period of temperatures below 5ºC ensuring the bud won’t develop prematurely into a shoot that will freeze to death during winter. The extent of the required cold period prevents the shoot from growing during warm spells during winter. In horticulture cold treatment is also applied and is called vernalization. Vernalization of Cypripedium seedlings is best carried out at 4ºC during a period of three months. Seedlings just out of flask have to deal with enough challenges already and should not be subjected to freezing. Although hardy plants are able to survive freezing, there’s always some damage done that may render the seedlings more vulnerable to attack by molds. If Cypripediums did not have proper vernalization, no shoot will develop. Plants may survive until next spring and then develop a shoot but it’s most likely they will perish during summer. Although some species only require a vernalization period of two months it’s best to subject seedlings to a vernalization period of at least three months to let the bud develop into a shoot in spring. Removal of Cypripedium Seedlings from the Flask When the seedlings have roots with a minimum length of around 3 cm and have visible dormant buds, they are ready to be removed from the flask. If not, let them grow on to this stage. Seedlings can be deflasked until January; after this date the cold period will become too short, because seedlings must be planted out before, or at latest in April (when only some light frost during night may occur). The first reason for this is that for strong shoot development the seedlings seem to require temperatures below 20ºC. The second reason is that for deflasked seedlings to develop their new bud, the growth season must be sufficiently long. The length of the required growth season depends on the species. The more Northern species such as C. guttatum, C. yatabeanum, and C. passerinum only require a short growing season but a relatively cold one as well. The species from warmer climates such as C. fasciolatum and C. kentuckiense need a longer growing season for proper new bud development. When having reached the proper size, the seedlings are carefully removed from the agar and transferred to a sieve. The plants are then washed with cold water under the shower until all agar is removed. The seedlings are then transferred to a clean ziplock plastic bag with a few drops of water added to it and labeled with name, date of removal from the agar (and the start of vernalization!), and number of seedlings. At this stage I always take an analogue picture from the bag with the seedlings and the clearly visible label. This will later demonstrate that the 2
How to raise in vitro propagated Cypripedium seedlings to fully established hardy plants for the garden © Camiel F. de Jong, December 2004 mature plants were raised from seeds. Of course this is not full proof but it forms a good indication. Vernalization Now the seedlings are ready to vernalize. The bags containing the seedlings are now transferred to a polystyrene-foam box. These boxes are available through laboratories or restaurants and are used to ship products that should be kept cool during transport. Make sure the box is clean. The bags with the seedlings are transferred to the box and the lid is closed. The box is then transferred to the refrigerator (4ºC). The box serves as a buffer for the fluctuations in temperature that occur in the refrigerator during normal operation. These temperature fluctuations cause condensation on the inner surface of the plastic bag and as a result some seedlings or parts of seedlings, dry out. The dried out parts die and form a good substrate for moulds when they re-hydrate again by contact with drops of condensation during normal storage or handling. The polystyrene box greatly reduces these problems. However, the seedlings should be checked regularly for development of mold. If this occurs, the affected seedlings must be removed. Alternatively, for larger numbers of seedlings, Bill Steele from Spangle Creek Labs, vernalizes his seedlings by transferring them to a polyethylene food storage box. Then a layer of water is added so that it’s just covering the bottom of the box and the box is placed in the refrigerator. Temperature changes during refrigeration cause condensation not only on the inner surface of the box, but also on the seedlings themselves, and so no desiccation occurs. The vernalization period should be at least three months and six months at most. Vernalization as described above requires the least cooled space. However, when one has sufficient space that can be kept frost free but below 4ºC, seedlings may directly be planted in the substrates and vernalized this way. This method is preferable as seedlings have time to settle themselves during vernalization, but is quite costly in terms of space. When using this method, care must be taken that there’s sufficient air movement to prevent molds from growing. However, drying out of the substrate must also be avoided. Substrate Mixes The mixes I use to plant the newly deflasked seedlings in are composed of Seramis, Perlite and Vulca. Seramis is a substrate that is developed for hydroponics. It consists of baked clay beads with a lot of pores. The material retains a lot of water and is also able to bind nutrients. It is not (yet?) available in the US but another product, Turface, seems to resemble it. This product is used to cover baseball fields. However, I never tested it. Perlite is an expanded volcanic material that retains some water but no nutrients. Vulca is basically pumice. Vulca itself is pumice from the Hekla volcano on Iceland. It makes the substrate mix somewhat drier and perhaps releases nutrients upon erosion. With the exception of Vulca these substrates cannot be 3
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Deflasking of Cypripedium seedlings - Cypripedium.de

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