Abstract
This protocol presents a method to test the effect of sorbitol in vitro on hybrid larch plants derived from somatic embryogenesis. We have tested four different media variants, one control variant without sorbitol and three variants of decreasing water potential corresponding to sorbitol concentrations in the culture medium of 4%, 10%, and 20%. We cultured two hybrid larch clones on these media during 35 days and assessed their vitality and weight growth after this time.
Keywords: In vitro stress test, Larch, Tree breeding, Sorbitol
Background
Forest tree breeding can benefit from methods that permit a high-throughput screening of plant lines in early stages of its development. Such methods help to delimitate the number of genotypes to be tested later in the field, an issue that in case of forest tree species requires considerable amounts of space. One possibility is to test young plantlets in vitro, a procedure that has the additional advantage of facilitating homogeneous and well-controlled experimental conditions. Aspects of tolerance against biotic and abiotic stress factors can be for instance screened in vitro. Among them, drought stress is receiving special attention in improvement programs. A common way to mimic water deficit is to add an osmoticum to the root supporting medium which reduces water availability. The osmoticum can be ionic like NaCl or non-ionic like mannitol, sorbitol or PEG (polyethylene glycol) (Singh and Singh, 2015). We have developed a protocol that allows testing the effect of sorbitol in vitro on hybrid larch emblings derived from somatic embryogenesis.
Materials and Reagents
Equipment
Software
Procedure
Data analysis
Notes
Recipes
Acknowledgments
We thank Kurt Zoglauer and his team at the Humboldt-Universität Berlin, Department ‘Biology, Botany & Arboretum’ for delivering all in vitro plant material. We also would like to thank Ute Tröber and Ursula Franke from the Division of Forest Genetics and Forest Tree Breeding at the Wood and Forestry Competence Centre of the Public Enterprise Sachsenforst for their help in the laboratory and Marianne Kadolsky for facilitating sterile work. We are very grateful to the head of the Division Heino Wolf for lending the required unit facilities and making this project possible. This work has been supported by the German Federal Ministry of Food and Agriculture by decision of the German Bundestag (Grant No. 22034914). There are no conflicts of interest or competing interests.
References
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