Infection and Synthesis Rate of Southern Bean Mosaic Virus in Soybean Callus Cells under Selected Cultural Conditions

WU, FANG-SHENG, and H. H. MURAKISHI. 1978. Infection and synthesis rate of southern bean mosaic virus in soybean callus cells under selected cultural conditions. Phytopathology 68:1389-1392. Soybean (Glycine max 'Harosoy 63') callus cells grown in with that of Prince bean callus treated identically or with either Eriksson's or Linsmaier and Skoog's media were infected soybean plants grown in the greenhouse. Vortexing inoculated with southern bean mosaic virus (SBMV) and of the cell-suspension/virus-inoculum mixture as is done for then incubated in either liquid or agar media. The tobacco mosaic virus-tobacco callus infection was not composition and type (liquid or agar) of the medium necessary with the soybean callus-SBMV system. Infection significantly affected the growth curve of the virus. Assays in Was achieved by adding virus inoculum to a suspension Phaseolus vulgaris 'Pinto' leaves indicated that soybean culture of callus cells, washing the cells with fresh medium callus grown in liquid and incubated in liquid medium after and incubating the cells in liquid media on a rotary shaker at inoculation gave the most rapid virus growth rate compared 120 rpm. Additional key words: virus-tissue culture interactions. The prospect of using pipettable suspensions of virusdependable bioassay host is also available (2). Callus infected cells or protoplasts has elicited considerable from its host plants can be easily induced and maintained excitement among plant virologists (16). Callus cultures in chemically defined media. These favorable (1, 9, 10, 11, 12, 13), separated cells (16), and protoplasts experimental conditions encouraged us to study SBMV(15) have been used by different investigators to study callus interactions. This report describes several virus replication and virus-host interactions. Early parameters that affect the multiplication rate and better studies of virus infection of callus cultures resulted in low synchronization of SBMV synthesis so that it could be efficiency of infection and slow growth rate of virus (4,6). used for the study of SBMV replication in cells at The low efficiency of infection was attributed to the macromolecular levels. inability of virus to penetrate the cellulose wall (4). Infection efficiency was improved by vortexing friable, MATERIALS AND METHODS suspension cultures of tobacco callus in the presence of Tissue culture and media.-Soybean (Glycine max virus and incubating the vortexed callus on agar medium 'Harosoy 63') or bean (Phaseolus vulgaris L. 'Prince') (11). When callus was not vortexed or when inoculated seeds were surface-sterilized with 70% ethanol for 3 min cells were incubated in liquid medium the infection and 20% Clorox for 20 min, rinsed in 0.01 N HC1, and efficiency was reduced significantly (10). washed several times with sterilized distilled water. The Plant virus vs. plant tissue culture relationships have shed severa times th sterile disted ter Th been studied so far using tobacco tissue and tobacco seeds then were transferred to sterile 50-ml test tubes in mosaic virus (TMV) almost exclusively (1, 4, 5, 6, 9, 10, which filter paper was folded and placed at the bottom of 11, 13). Other viruses with different morphology, the tubes to support the seeds. The seeds were kept moist transmission characteristics, and host range have not by the addition of 9 ml of Linsmaier and Skoog (7) beensstuied bcharauseeaisuitablenexperimentalesystemwas medium diluted 1:4 with water. The seeds were allowed to been studied because a suitable experim ental system was g r i a e u d r t e s m i h n e p r t r not available. Strains of southern bean mosaic virus germinate under the same light and temperature (SBMV) are world-wide in distribution and affect conditions used for growing tissue cultures. The primarily bean (Phaseolus vulgaris L.), soybean (Glycine hypocotyl was cut into 1-cm segments and placed on max L.), and cowpea (Vigna sinensis L.) (14). The SBMV designated media solidified with 1% agar. Then the is an isometric virus with a diameter of 25 nm (8, 14). It induced calli were maintained either on liquid media may be seed-borne and is transmitted by the bean leaf (liquid-grown) or on media solidified with l%agar(agarbeetle (Ceratoma trifurcata) as well as by leaf rubbing grown). Gro-Lux fluorescent lamps [861 lux (80 ft-c)] and (14). The yield of SBMV from intact plant tissue is a temperature of 24 C ± 1.0 C were used under all growth rela.Tivelyhigh;thel purficationprocdr i smpla ue, ad aconditions. Unless otherwise specified, calli grown in relatively high; the purification procedure is simple, and a liquid media were incubated in Erlenmeyer flasks and 00032-949X/78/000 252$03.00/0 placed on a rotary shaker at 120 rpm. Copyright © 1978 The American Phytopathological Society, 3340 The R3 medium is composed of Linsmaier and Skoog Pilot Knob Road, St. Paul, MN 551 21. All rights reserved, minerals (7), and 30 g sucrose supplemented with 0.5 mg