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

Giardini Naxos, Italy - 18/21 September, 2002

ISBN 88-900622-3-1

Oral Communication Abstract - S1b

MORphological modifications of citrus by genetic transformation with rolABC genes

Deng Z.N., Gentile a., La Malfa S., Domina F. Tribulato E.

Dipartimento di OrtoFloroArboricoltura e Tecnologie Agroalimentari, Via Valdisavoia 5, 95123 Catania

citrange, rootstock, genetic improvement, tree size control, gene expression

Citrus improvement by conventional breeding is hampered by their long reproductive cycles, self-incompatibility, nucellar embriony and high degree of heterozygosity. Rootstock breeding, as a long term research, is of essential importance in citrus industry. One of the main breeding purpose is to select rootstock of dwarfing effect to scion cultivar, so that tree size is able to be controlled and consequently the production efficiency could be increased. For deciduous fruit trees dramatically improvement of production has been achieved through the use of dwarfing rootstocks. However , there is still not useful dwarfing rootstock available in citrus and the few existing dwarfing materials have serious limitations due to the low adaptability to different environmental conditions or the incompatibility with scions.

With the aim at obtaining dwarfing selection, a research program was set up to introduce rolABC genes from Agrobacterium rhizogenes into the genome of citrange Troyer [Citrus sinensis (L.) Osbeck x Poncirus trifoliata (L.) Raf.], an important rootstock as valid alternative to sour orange.

Stem segments were infected with A. tumefaciens strain C58C1 containing pDN3514 with rolABC genes and selectable marker (nptII). Shoots growing on selective medium were micrografted or autorooted. Putative transgenic plantlets, analysed by PCR, were acclimatised in greenhouse. Five different grafted clones and 25 rooted plants were chosen for observation of growth parameters (i.e. plant height, internode length, number of shoots, leaf area). Gene expression analysis was performed with total RNA extracted from leaves of transformed plants using RT-PCR beads (Pharmacia, Uppsala).

Transgenic citrange plants were verified by PCR analyses using specific primer for rolA, rolB and rolC. RT-PCR analysis with the RNA from the clones “B”, “D”, and “E” indicated that the inserted genes rolA, B and C were correctly expressed.

The observation of morphological characters indicated a strong dwarfing effects in the transgenic plants. However, the level of expression was different among the clones and also the growth habit observed was not uniform for all the transgenic plants. Plant height of different clones was about 50-66% less than the wild type. This was tightly linked with short internode length (60-66% less). In the transgenic plants it was observed that the apical dominance was inhibited. Every plant produced 2-4 branches while in the control plants had only the main shoot growing upwards.