Date of Award
Doctor of Philosophy
Zoology and Genetics
Robert Michael Benbow
Apical meristem and axillary bud development in germinating soybean embryonic axes treated for 24 hr with the cytokinin, 6-benzylaminopurine (BAP) seemed normal morphologically, but was delayed by about 9 to 15 hr. After 5 days, numerous, rapidly proliferating meristematic regions (de novo vegetative buds) were observed in BAP-treated axes around the perimeter of the apical dome. These visible changes were preceded by transient inhibition of DNA synthesis in the shoot apices as early as 21-36 hr post-imbibition;Some of the new meristematic regions developed into multiple shoots, which were excised, subcultured, and used to generate mature plants. In the absence of BAP, multiple shoots were never formed. These results suggest that transient exposure to BAP interrupted normal chromosomal DNA replication and caused reprogramming of the developmental fate of a large number of cells in the shoot apex;Soybean embryonic axes were bombarded with microprojectiles to which expression vectors had been adsorbed. Gene expression was observed in 93% of bombarded axes within 24 hr. Maximal expression was observed in BAP-treated axes bombarded with vectors that contained a partial duplication of the cauliflower mosaic virus 35S promoter, an omega prime sequence, and a nopaline synthase polyadenylation sequence. Gene expression and persistence of exogenous DNA was observed in 12 out of 1,034 mature plants. Nine of the 12 plants showed expression and persistence of plasmid DNA at multiple nodes with distinctive patterns of expression;The effects of petunia transformation booster sequence (TBS) on the expression and persistence of supercoiled expression vectors in bombarded dicotyledonous (tobacco) and monocotyledonous (maize) plant cells were compared. Petunia TBS increased the transformation frequency 7.8- to 16-fold in tobacco, and 1.7- to 2.4-fold in maize. Although TBS contains a well defined transcription enhancer element, no significant differences were observed in levels of expression from transgenic tobacco or BMS cells. TBS did not appear to alter the complexity of integration patterns of exogenous DNA in either tobacco or maize transformants. In tobacco, transformants usually contained at least one copy of the introduced gene in a non-rearranged form, and often contained multiple copies of the gene in rearranged forms as well. In maize, however, most transformants contained only rearranged copies.
Digital Repository @ Iowa State University, http://lib.dr.iastate.edu/
Charisse Marie Buising
Buising, Charisse Marie, "Direct introduction of DNA into embryonic axes of Glycine max by microprojectile bombardment " (1992). Retrospective Theses and Dissertations. 9821.