Micropropagation of walnuts (Juglans regia L.) in vitro

Najčešći problem u mikropropagaciji oraha predstavlja posmeđenje hranjivog medija. Značajan je utjecaj i genotipa. Najprikladniji medij za orah je DKW medij s koncentracijom saharoze od 3 %. BAP, IBA i GA3 imaju pozitivan utjecaj na klijanje embrija, razvoj kalusa i proliferaciju pupova. Kao zgušnjivač najčešće se upotrebljava agar. Okolišni uvjeti svjetlosti od 1000 do 3000 lux-a (cca. 40 – 92 μmol/m-2/s-1) i temperatura u rasponu od 22 do 28 °C najčešće se koriste u kulturi tkiva oraha. Rizogeneza mikroizdanaka može se postići u in vitro i ex vitro uvjetima. Predtretman s IBA-om uz tamnu fazu preporuča se za indukciju rizogeneze. Smanjenje koncentracije soli, povećanje sadržaja saharoze, dodavanje aktivnog ugljena i vermikulita u medij bez hormona predstavljaju odlučujuće čimbenike u poboljšanju uspješnosti rizogeneze. Postupak aklimatizacije oraha vrlo je težak zbog brzog propadanja presadnica i njihove osjetljivosti na bolesti uslijed visokog sadržaja vlage tijekom faze rizogeneze. Uspostavljanje tekuće kulture (TIB/TIS sustavi) daje nekoliko drugih prednosti te je važan korak u automatizaciji samog procesa mikropropagacije oraha.The most common problem in micropropagation of walnuts is browning of the nutrient medium. The influence of genotype is also significant. The most suitable medium for walnut is DKW medium with a sucrose concentration of 3%. BAP, IBA and GA3 have a positive effect on embryo germination, callus development and bud proliferation. Agar is most often used as a solidifying agent. Ambient light conditions of 1000 to 3000 lux (approx. 40 - 92 μmol/m-2/s-1) and temperatures ranging from 22 to 28 °C are most commonly used in walnut tissue culture. Rooting of microshoots can be achieved in in vitro and ex vitro conditions. Pre-treatment with IBA in dark phase is recommended for induction of rooting. Decreasing the salt concentration, increasing the sucrose content, adding activated charcoal and vermiculite to the hormonefree medium are importand factors in improving the performance of rhizogenesis. The process of walnuts acclimatization is very difficult due to the rapid decay of seedlings and their susceptibility to disease due to the high moisture content during the rhizogenesis phase. Establishing a liquid culture (TIS/TIB systems) provides several other advantages and is an important step in automating of walnut micropropagation process