Doctoral Dissertations

Date of Award

12-1986

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Major

Plant, Soil and Environmental Sciences

Major Professor

B. V. Conger

Committee Members

Karen W. Hughes, Robert A. McLean, John H. Reynolds, Otto J. Schwarz

Abstract

Direct embryogenesis was observed from Dactylis glomerata L. [orchardgrass (2N=4X=28)] anthers incubated at 25 C for 6 weeks on Schenk and Hildebrandt (SH) medium containing 3% sucrose and 30 pM dicamba [SH-30 (3,6 dichloro-o-anisic acid)]. Subsequent experi-ments showed that SH-30 supplemented with up to 5.0 g/1 casein hydro- lysate did not affect and 0.1 to 5.0 mg/1 benzylaminopurine or kinetin hindered the embryogenic response. Furthermore, anther orientation had no significant effect on the embryogenic response and no embryos were obtained from culture of isolated microspores. However, SH-30 containing 94 sucrose promoted embryogenesis and an anther density of 10/ml initiated more embryos per anther than the 100 anthers/ml treatment. Anthers initiated embryo-like structures from within micro- spores after culture on SH-30 for 6 weeks at 4 C. In another experi-ment, anthers were exposed to 0, 3, or 6 week cold pretreatments (4 C) and then cultured at 25 C on SH medium containing 0, 10, 20, or 30 pM dicamba and 9% sucrose. Embryo initiation was affected by an interaction of cold pretreatment and dicamba concentration. Stereomicroscopy showed that the 6 week cold pretreated anthers initi-ated indirect embryos from callus masses derived from microspores. Five of six plants regenerated from the 6 week cold pretreated anthers consisted of cells with 28 chromosomes. However, 1 plant had 14, 28, and up to 112 chromosomes and 1 to 8 nucleoli per nucleus. Direct embryogenesis was observed from ovary and style regions of unpollinated pistils cultured on SH-30 for 3 to 4 weeks at 25 C. After 5 and 6 weeks culture, however, embryos disorganized and produced calli which proliferated into numerous secondary embryos. An upright pistil orientation initiated a significantly greater embryo- genic response compared to those pistils cultured flat on the medium. All examined plants regenerated from pistil culture possessed the somatic chromosome number of 28 except for 1 mixoploid with 14, 28, and 56 chromosomes. Culture of excised unpollinated ovules failed to initiate any embryos when cultured on SH-30 with or without kinetin concentrations up to 5.0 mg/1. However, 1 of 60 ovules excised from pistils cultured for 3 weeks on SH-30 and then recultured on SH-30 initiated callus and embryos. Histology of pistils cultured for 3 weeks showed possible cell divisions within the embryo sac.

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