Plant Cell, Tissue and Organ Culture 58: 155–157, 1999. © 2000 Kluwer Academic Publishers. Printed in the Netherlands. 155 Research note Establishment of callus and cell suspension cultures of Opuntia ficus-indica R´ omulo M. Llamoca-Z´ arate 1,2 , Cla´ udia Studart-Guimarães 2 , Joerg Landsmann 1 & Francisco A.P. Campos 2,∗ 1 Institute for Plant Virology, Microbiology and Biosafety, Federal Biological Research Centre for Agriculture and Forestry, Messeweg 11/12, D-3810 Braunschweig, Germany; 2 Department of Biochemistry and Molecular Biology, Federal University of Cear´ a, P.O. Box 1065, 60001-970 Fortaleza, CE, Brazil, ( ∗ request for offprints; E-mail: bioplant@ufc.br) Received 13 April 1999; accepted in revised form 9 November 1999 Key words: Cactaceae, forage crops, in vitro propagation Abstract Friable callus cultures were initiated from cotyledons and hypocotyls of Opuntia ficus-indica. Explants from cotyle- dons produced significantly more callus than those from hypocotyls. Optimum callus growth was observed on Mur- ashige & Skoog medium supplemented with 0.9 μM 6-furfurylaminopurine, 2.3 μM 2,4-dichlorophenoxyacetic acid, 1.0 μM 4-amino 3,5,6-trichloropicolinic acid, 400 mg l −1 casein hydrolysate and 3% sucrose. The same medium without agar was used for establishing cell suspensions. Abbreviations: kinetin – 6-furfurylaminopurine; 2,4-D – 2,4-dichlorophenoxyacetic acid; picloram – 4-amino 3,5,6-trichloropicolinic acid; MS – Murashige & Skoog basal salt medium The prickly-pear (Opuntia ficus-indica Mill.) is a cac- tus species widely utilized in semi-arid regions as a forage crop (Felker, 1995). In periods of drought, its fleshy phylloclads are the sole source of water and nutrients for farm animals. Tissue culture techniques are essential in order to facilitate the application of modern biotechnological methods to this crop. The techniques for in vitro culture of Cactaceae are still not well developed, and the effect of different growth reg- ulators on morphogenesis is not well understood (Fay and Gratton, 1992; Palomino et al., 1999). We present the conditions for the induction and establishment of friable callus and cell suspension cultures. Callus cultures were initiated from cotyledon and hypocotyl sections of 21-day old light-grown seed- lings. Seedlings were rinsed under running tap water, submerged in 70% ethanol for 1 min and transferred to a flask containing 100 ml of 1.5% sodium hypochlorite for 10 min, then rinsed three times (5 min each) in sterile distilled water. Hypocotyl and cotyledon sec- tions were then transferred to 12×90 mm Petri dishes containing 30 ml of semi-solid medium, consisting of full-strength MS salts and vitamins (Murashige and Skoog, 1962), 30 g l −1 sucrose, 8 g l −1 agar and supplemented with different combinations and con- centrations of growth regulators: 2.3, 11.3 or 18.1 μM 2,4-D, 0.9 μM kinetin, 0.2, 0.4, 1.0 or 2.0 μM picloram. The pH of all media was adjusted to 5.8 with 1.0 M KOH and autoclaved at 121 ◦ C and 103 kPa for 15 min. The cultures were incubated in the dark at 28 ◦ C. Callus was separated from explants and subcultured at 3 or 4-week intervals onto medium sup- plemented with 2.3 μM 2,4-D, 0.9 μM kinetin, 1.0 μM picloram and 400 mg l −1 casein hydrolysate. This medium was designated friable callus medium (FCM). Cell suspension cultures were initiated by shaking 35-40 g of friable callus at 150 rpm in the dark at 28 ◦ C in a 300 ml Erlenmeyer flask containing 150 ml of liquid FCM. After 48 h free cells and small cell aggregates were separated by sieving through a 1 mm stainless steel screen and allowed to settle in Falcon plastic tubes for 30 min. Inocula of 10 ml of packed cells were transferred to a 300 ml Erlenmeyer flask containing 50 ml of FCM. After 3 weeks of culture the