Proceedings of the XLV Italian Society of Agricultural Genetics - SIGA Annual Congress

Salsomaggiore Terme, Italy - 26/29 September, 2001

ISBN 88-900622-1-5

Oral Communication Abstract

IDENTIFICATION AND CHARACTERISATION OF GENE SEQUENCES EXPRESSED DURING WHEAT HEADING

CIAFFI M., PAOLACCI A.R., MARABOTTINI R., TANZARELLA O.A.

Dipartimento di Agrobiologia e Agrochimica, Università della Tuscia, Via S. Camillo de Lellis, 01100 Viterbo, Italy

ciaffi@unitus.it

wheat, flowering control, flower development, differential expression, Northern analysis

Flowering is a crucial and complex event during the plant ontogenesis, which requires the switch from the vegetative to the reproductive phase. This transition is triggered through a series of endogenous and environmental stimuli (e.g. photoperiod and vernalization). The induced floral meristem is then canalised toward a second phase, consisting in the development of the different flower structures, which is under strict genetic control. Many genes involved in the control of the different phases of flowering (flowering time, floral meristem identity and organ identity genes) have been identified, cloned and characterised from Arabidopsis and Anthirrinum. The comparison of gene sequences controlling flowering induction and flower development isolated from several plant species has shown their high conservation among the dicotyledons. The present knowledge of the metabolic pathways involved in plant flowering and of their molecular control is very far from a whole and detailed comprehension. However, the available information seems adequate for starting these studies also in important crop species, such as cereals. The flower morphology of these species is quite different from that of Arabidopsis and Anthirrinum, therefore it would be very interesting to understand the relationships between the flower organs of monocots and dicots and the molecular mechanisms controlling their development. Up to the present the knowledge in the monocot is limited to some MADS box genes isolated from rice and maize.

The aim of the present research consists in the identification, cloning and characterization of genes specifically expressed in wheat spikelets at different stages of spike development. Spikes and leaves were collected, at interval of five days, starting when the spike was 1-1.5 cm long and until complete heading, from five bread wheat and six durum wheat varieties. The single organs forming the spikelet were separately collected from three varieties of bread wheat and two of durum wheat. The mRNAs differentially expressed in spike and leaf at heading time were analysed by the SDDM (simple differential display) method, which consists in the reverse transcription of mRNA using random hexanucleotides and then PCR amplification by decamers. Eight out of twenty decamers analysed during the first phase of the research detected 25 differential PCR products between spikes and leaves. The 25 differential bands were cloned and for each of them ten clones were analysed. A total of 45 inserts of different sizes were found and were analysed by Northern analysis with total RNA from spikes and leaves. Four groups were recognised on the basis of the Northern analysis: I) 15 clones hybridising only to the spike RNA; II) 20 clones showing a more intense signal with the spike than with the leaf RNA; III) 3 clones void of hybridisation with both tissues; IV) 7 clones with similar signals in both tissues. The 35 clones of the first two groups were wholly sequenced. Their sequences were searched in the nucleotide and EST databases and the deduced amino acid sequences were searched in the protein databases. For 16 of the clones it was not detected any significant homology, whereas seven of the remaining 19 sequences showed homologies only with EST sequences. Some of the sequences showed homologies with interesting proteins, for instance transcription factors, such as a zinc-finger of wheat and maize.

The short-term continuation of the research will consist in the following phases: I) Northern analysis of the most interesting clones with several tissues for a better clarification of their tissue-specific expression; II) further diffrential analysis of the spike at different development stages with more decamers and with the cDNA-AFLP technique; III) chromosome location of the isolated clones by aneuploid analysis.