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Degree

Doctor of Philosophy, Ohio State University, Horticulture and Crop Science, 2006.

Abstract

In plants, the cryptochrome gene family is responsible for mediating photomorphogenic processes in response to blue light. Here, RT-PCR and RACE techniques were used to clone CRY1 and CRY2 from Nicotiana sylvestris. In a reverse genetics approach, antisense and overexpression constructs for each gene were then used to transform N. sylvestris and N. tabacum cv. Maryland Mammoth plants by inoculating sterile leaf explants via Agrobacterium tumafaciens. RT-PCR results indicate that for N. sylvestris, one CRY1 overexpression line (S1-6) and one CRY2 overexpression line (NsS2-2) were produced. Two CRY1 overexpression lines (S1-3 & S1-4) and two CRY2 overexpression lines (NtS2-2 & S25) were generated in N. tabacum cv. Maryland Mammoth. Western analysis confirmed overexpression of CRY1 in S1-3, S1-4 and S1-6 plants. To characterize the transgenic plants at a functional level, hypocotyl length and leaf chlorophyll content was measured. Major reductions in hypocotyl elongation and increased in chlorophyll were observed in all of the CRY1 and CRY2 overexpression plants in both species. Also in both species, the effect of CRY1 and CRY2 overexpression on flowering time was examined. In marginally inductive and in naturally changing photoperiods, the appearance of visible flower buds occurred sooner than in wild type plants. Unexpectedly, flowering occurred in non-inductive conditions, indicating that the photoperiodic timing mechanism of these obligate species is influenced in part by CRY1 and CRY2. Additional effects on vegetative growth and leaf development were explored. Aspects specific to sun leaf morphology, including increased thickness of palisade parenchyma and higher stomatal density were seen in CRY1 overexpression plants that were grown in low light conditions. Finally, the N. sylvestris CRY1 and CRY2 overexpression constructs were used to generate transgenic petunia plants. Compared to wild type the growth rate of CRY1 overexpression plants was dramatically reduced and CRY2 overexpression resulted in more compact plants with plants that produced a greater number of branches. Taken together with the increased chlorophyll, dwarf phenotype, increased leaf thickness and higher stomatal density, it was concluded that CRY1 plays a role in mediating photomorphogenic processes involved with photoprotection and the development of sun leaf characteristics in response to light intensity.

Keywords

Cryptochrome; Photomorphogenesis; Photoperiodic Flowering; Vegetative Leaf Development

Advisor

James D Metzger

Pages

169p.

Document number: osu1155657216
Permalink: http://rave.ohiolink.edu/etdc/view?acc_num=osu1155657216

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