Abstract
Many essential genes are required to participate in and regulate the progress of cell cycle which is an important process for plant growth and development. These key regulators not only participate in DNA synthesis and cell division but also get involved in the transmission of signals during growth and development. In order to understand the molecular mechanisms in controlling plant cell cycle accompanied with plant growth and development, Agrobacterium - mediated gene transfer method was used to introduce three cell-cycle related genes (AtCDC25, AtR2, and OsR2) which are driven under CaMV35S promoter into tobacco (Nicotiana tobaccum). The effects of these genes on tobacco growth and development were then investigated. The results showed that totally 45 transgenic tobaccos were obtained, including 27 lines with single gene insertion, 19 lines with two genes insertion, and 2 lines with three genes inserted in tobacco genome. Analysis of the transgene expression in T0 transgenic tobaccos by RT-PCR indicated that there were more than 50% transgenic plants with foreign gene expressed. The phenotypes of T0 transgenic plants showed no obvious variation. However, a few of them displayed aberrant leaves and delayed flowering. Microscopic anatomy analysis of leaf morphology showed that there were 1.5 folds increase in the number of spongy cells in transgenic tobacco; also the cell number of the lower epidermis was increased. In addition, we noticed that transgenic tobacco with high levels of gene expression, especially lines with three genes inserted significantly correlated with much more cell numbers and serious mis-arranged cells in leaf. Our results demonstrated that overexpression of AtCDC25, AtR2, and OsR2 can enhance the cell division and reduce the size of cells. Finally, analysis of the germination ability of T1 transgenic plants suggested that both the germination tendency and germination rate were lower than those of wild type. Taken together, overexpression of AtCDC25, AtR2, and OsR2 leads to delay of flowering time, increase of cell division rate, reduction of the cell size, and decrease of the germination ability of transgenic tobaccos. The phenotypes of transgenic tobacco were similar to non-transgenic plants. The interpretation for this may be due to the compensatory mechanism in cell cycle after disruption of this process by overexpress of foreign genes.
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