La coltura in vitro per la conservazione della biodiversità orticola

Abstract

Today, in vitro culture is a strategic tool to support medium and long-term conservation of plant genetic resources by using the slow growth storage of shoot cultures and the cryopreservation of organs and tissues. Over the last 30 years, considerable progresses were made in the development of both techniques that are nowadays considered as ex situ conservation strategies complementary to traditional seed banks and in-field clonal collections. Efficient protocols were developed for the conservation of a large number of crops, including important vegetables of the temperate environment (garlic, artichoke, asparagus, mint, potato, sweet potato, tomato, red chicory, thyme). Conservation in slow growth storage consists in modifying the medium and/or culture conditions to reduce the growth of plant material without affecting the viability and regrowth potential of shoots when moved back to standard culture conditions. The technique allows medium-term crops conservation, with a storage time of vegetables ranging from a few months to two years and more, without recurring subculturing typical of micropropagation. Cryopreservation preserves plant organs and tissues at ultra-low temperature, as liquid nitrogen temperature (-196°C). Currently, various techniques are available, based on cold tolerance induction by cell cytoplasm vitrification during the fast ultra-freezing through in liquid nitrogen immersion of explants. The term “vitrification” refers to the solidification of a liquid without crystallization. If induced in plant cells, it avoids the formation of lethal intra-cellular ice crystals during the ultra-freezing process, keeping tissues at stopped metabolism condition but vital. Working with vegetables, the techniques based on direct immersion of specimens in liquid nitrogen (“one-step freezing”, such as the “droplet-method”, the “PVS2 vitrification”, the “encapsulation-based” and the “cryo plate-based” procedures) are the most used. Further, the “two-step freezing”, i.e. the slow explants cooling before immersion in liquid nitrogen, still finds some applications. The experimental activity mainly focused on three economically important species, i.e., Allium sp., potato and sweet potato. Currently, almost 38,000 accessions of garlic, cassava, mint, potato, sweet potato, taro, yam are maintained in 17 genetic resources conservation centers, located in 12 Countries and 5 Continents (Europe, Asia, Africa, North and South America). Approximately 4/5 of these accessions are maintained in vitro by means of slow growth storage of shoot cultures, but more recent cryopreservation is constantly growing, and over 7,500 accessions from vegetables are stored at -196 °C. The germplasm of potato (Solanum sp.) is by far the most collected in the world, with almost 17,700 accessions presently maintained in in vitro banks and cryobanks.