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Universidade de Coimbra (2011)

Strategies for the improvement of Arbutus unedo L. (strawberry tree) : in vitro propagation, mycorrhization and diversity analysis

Gomes, Maria Filomena Figueiredo Nazaré

Titre : Strategies for the improvement of Arbutus unedo L. (strawberry tree) : in vitro propagation, mycorrhization and diversity analysis

Auteur : Gomes, Maria Filomena Figueiredo Nazaré

Université de soutenance : Universidade de Coimbra

Grade : Doutoramento em Biologia (Fisiologia) 2011

Résumé
Arbutus unedo grows spontaneously around the Mediterranean basin. The species is drought tolerant and able to regenerate following forestry fires making it quite interesting for forestation programs in Mediterranean regions. Fruits are used to make jellies and a spirit which represents the main income for owners. Considering the sparse information about the potential of this fruit tree to be propagated in vitro, a project to clone selected trees based on their fruit production was initiated. The role of several factors on propagation was evaluated and studies concerning the mycorrhization and genetic diversity analysis were also performed. Shoot apices from epicormic shoots showed higher rates of in vitro establishment. Of the three basal media tested, the Fossard medium with the micronutrients of the Murashige and Skoog medium gave the highest rates of multiplication. Kinetin (8.9 μM) gave the best results although not different (P>5%) from those obtained with other cytokinins such as benzyladenine or zeatin. Thidiazuron or 1-naphthaleneacetic acid promoted callus growth and had a deleterious effect on the multiplication rate. The genotype was also a factor affecting in vitro multiplication and the conditions used for shoot multiplication greatly influenced further behaviour of shoots during the rooting phase. In this phase, the inclusion of an auxin significantly increased rooting yields. Anatomical studies data indicated that adventitious roots had a deep origin in the shoot stem, probably from the cambial zone. From the five substrates tested during acclimatization, perlite without fertilizer gave the best survival rate (98.0±1.23%) after 4 months. Mycorrhization was tested in vitro and under nursery conditions. Arbutoid mycorrhizae were observed in vitro 1 month after inoculation indicating compatibility between A. unedo and Pisolithus tinctorius. Both mycorrhizae inocula treatments tested in nursery (vegetative inocula and dry sporocarps) improved plant growth compared to control plants and seedlings after 20 months in a field trial (P>5%). The genetic diversity in 27 Arbutus unedo genotypes was assessed by molecular markers (RAPD and SSR). The RAPD primers generated 124 bands from which 57.3% showed to be polymorphic. Eleven SSR primers first tested in Vaccinium (an Ericaceae) were previously selected. Five SSR loci were polymorphic displaying a 75% mean expected heterozygosity which is a higher value than that observed with RAPDs (27%). Thus, the Lynch (1990) similarity coefficient revealed a similarity among trees higher for RAPD than SSR, respectively of 83% and 21% (Mantel test : r= 0.64 ; P<0.001 ; r=0.75 ; P<0.001, respectively). No genotypes could be grouped according to their geographical origin for both markers. Some of the produced plants are now in the field for further studies. The data presented in this thesis have provided protocols concerning the in vitro propagation of adult plants, mycorrhizal synthesis and allowed the identification of five polymorphic molecular markers (SSR), used on genetic diversity analysis of 27 genotypes. The results are discussed and further research envisaged.

Présentation -> http://hdl.handle.net/10316/17964

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