Metla Project 3198

Application of biotechnology for genetic research and gene conservation of forest trees

Duration: 1997-2003 Keywords: DNA markers, biotechnology, cryopreservation, genetic transformation, molecular biology, tissue culture

Objectives

The main aim of the project is to develop and apply biotechnological methods for studies in forest genetics and tree breeding. Vegetative propagation methods can be utilised in forest tree breeding both in evaluating breeding values as well as in transferring genetic gain obtained into practical silviculture. Vegetative propagation is also used for gene conservation purposes. DNA-markers and genetic transformation techniques, on the other hand, provide a lot of information on the genetic structure and functioning of tree species.

The research going on in the biotechnology project is mainly basic research aimed at increasing current knowledge of the fundamental processes behind tree growth and other economically important characteristics, and partly applicable to breeding and research practices. Typical for the project is combining of different techniques and broad collaboration with other research groups both in Finland and abroad.

Research is currently focused on the following topics:

Vegetative propagation of Scots pine - development of somatic embryogenesis and studies on the effect of mycorrhizal fungi on the rooting of vegetative propagules (cuttings, somatic embryos)
Cryopreservation - conservation of tree individuals in liquid nitrogen and the efficient regeneration of plants from the cryostored material in Scots pine, silver birch and aspen
Transformation studies - development and application of gene transfer techniques for studying gene functioning in Finnish forest tree species

Collaboration with:

Other research projects within Finnish Forest Research Institute Universities of Oulu, Kuopio, Joensuu, Turku and Helsinki; Wood Wisdom Research Programme
EU / COST Action 843, Quality Enhancement of Plant Production through Tissue Culture, Working Group I, Developmental Biology of Regeneration
Institute of Biotechnology, ETH-Zürich, Sveitsi / Dr. P. Kallio
Michigan Technological University, Yhdysvallat / Prof. V. Chiang

Results

A vegetative propagation method based on the production of somatic embryos (SE) has been developed for Scots pine, and the first pine plants regenerated using this method. Also the technique for cryopreservation of the SE cultures has been developed. The explants used for the induction of the SE cultures are young, immature seed embryos with surrounding megagametophytes. So far, 28 procent of the randomly chosen and tested seed families have been compatible for SE production by the current method. The most problematic phase in the Scots pine somatic embryogenesis has proven to be germination of the embryos obtained, especially the development of the roots. For solving the rooting related problems, the effect of mycorrhizal fungi is being studied. The application of the mycorrhiza has already been observed to enhance the rooting of the Scots pine cuttings.

In the cryopreservation studies, functional conservation techniques have been developed both for the SE cultures of Scots pine and for silver birch material. In the case of birch, either buds cold-hardened in the nature or tissue cultured material cold-hardened in vitro and treated with cryoprotectants can be used in cryopreservation. The research for the application of these methods also in aspen is going on. In addition, the genetic fidelity of the cryostored material has been studied together with the effect of the different chemicals used as cryoprotectants. According to the results there are differences between the generally used cryoprotectants and their mixtures, both in recovery of the material after cryostorage as well as in the genetic fidelity of the samples, and these effects should be taken into account when applying the technique for practical or research purposes.

In the genetic transformation studies, several techniques applicable in silver birch have been developed and further on, used for obtaining specific information on gene functioning related to the following research topics:

the connections of the carbon and nitrogen metabolism together with the effects on tree growth and adaptation
the effects of plant growth regulators, especially auxins, on the wood anatomy and chemistry
the biosynthesis of lignin.

The chemical and molecular analyses of the transgenic birch lines produced are currently going on. In coniferous species, Scots pine and Norway spruce, the research has been focused on developing transformation systems for routine use. Using tissue cultured material as target, transgenic cell lines have been obtained in Scots pine, but the regeneration of plants from these lines has not been successful.

Transgenic Scots pine has, however, been produced through pollen transformation, and the method used is probably suitable also for Norway spruce. In this technique, transgenes are inserted in mature pollen that is then used for controlled pollinations. A high proportion (12-84%) of the pollen grains express the transformed reporter gene, but only a small proportion of the progenies have been found to be transgenic. Using the current method, genetic transformation is possible without any tissue culture methods or involvement of the selectable marker genes, such as antibiotic or herbicide resistance. The technique can be applied for numerous pollen lots and the crossings made with several mother trees.

Project leader: Aronen, Tuija
The Finnish Forest Research Institute, Punkaharju Office, Finlandiantie 18, FI-58450 PUNKAHARJU, FINLAND
Phone: +358 29 532 4233
E-mail: tuija.aronen@metla.fi

Other researchers: Hassinen, Minna (2002), Häggman, Hely (2001-02), Jokipii, Soile (2001-02), Kontunen-Soppela, Sari (2001-03), Luomala, Eeva-Maria (2002), Ryynänen, Leena, PU (2001-03), Tiimonen, Heidi, PU (2001-03), Timonen, Ulla (2002)

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Updated 12.06.2012
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