Plant Cell, 1issue and Organ Culture 44: 155-159, 1996. 155 © 1996 KluwerAcademic Publishers. Printed in the Netherlands. Growth and regenerability of long-term suspension cultures of the U.S. rice cultivar Mercury Herry S. Utomo, Timothy P. Croughan & Suzan S. Croughan* Rice Research Station, Louisiana State University Agricultural Center, P.O Box 1429, Crowley, LA 70527-1429, U.S.A. (* requests for offprints) Received29 December1994;accepted in revisedform9 November1995 Key words: Oryza sativa L., regeneration, tissue culture, rejuvenation Abstract Suspension cultures of the U.S. rice cultivar Mercury have been maintained in modified General Medium for more than 3 years. These suspensions have continued to have high and relatively stable regeneration rates. Two different explants, immature panicles and seeds, were compared during the development of these embryogenic suspensions. Initial formation of secondary embryogenic callus from immature panicles on induction medium was greater than that from seeds. Suspensions of these two cell lines, however, did not differ morphologically and maintained similar regeneration rates. After 5 months in culture the rates of regeneration began to decline. The suspensions were plated onto regeneration medium without growth regulators for 2 weeks and then embryogenic cells were manually selected and used to develop secondary suspensions. Through this simple rejuvenation procedure, the suspensions retained high and stable regeneration rates. Variability in suspension growth, however, was observed during the culture period. Slower growth occurred at weeks 13, 15, 27, and 29 and was associated with a decrease in regeneration rates. Reproductive fertility of regenerated plants remained high for 3.5 years but then declined. Abbreviations: CH- casein (acid hydrolysate); 2,4-D- 2,4- dichlorophenoxyacetic acid; MS - Murashige & Skoog basal medium; SE- standard error Introduction The use of suspension cells of rice (Oryza sativa L.) can facilitate the accomplishment of several research objectives. They have been utilized to generate trans- formed rice plants, through both particle bombardment and polyethylene glycol treatment (Cao et al., 1992; Lee et al., 1991), and for in vitro screening (Yamada et al., 1986). Rice suspension cells also have been wide- ly utilized as a source for protoplasts that are used for somatic hybridization (Kyozuka et al., 1989), the pro- duction of protoclones (Kanda et al., 1988), transfor- mation (Zhang & Wu 1988; Li etal., 1992; Shimamoto et al., 1989; Toriyama et al., 1988; Zhang et al., 1988; Davey et al., 1991), and in vitro screening (Xie, 1991; Utomo, 1994). In other species, suspension cells have been used for rapid and continuous production of uni- form biomass for studies of basic plant cell physiolo- gy (Fry, 1990), enzymology (Herzbeck & Hiisemann, 1985), and secondary metabolites (Hall et al., 1988). Establishment, selection, and preferential culture methodology of embryogenic callus have been recog- nized as critical factors in maintaining long-term mor- phogenetic potential in cultures (Vasil & Vasil, 1984). In recent years, the establishment of embryogenic cul- tures has been achieved with several rice genotypes previously regarded as recalcitrant (Lee et al., 1989; Datta et al., 1990; Datta et al., 1992). This resulted from better recognition of the conditions required to produce embryogenic calluses (Vasil & Vasil, 1991; Datta et al., 1992). However, characterization and maintenance of long-term embryogenic rice suspen- sions has not been reported. This paper describes the characteristics of embryogenic suspension cells of the U.S. rice cultivar Mercury that have been in culture for over 3 years.