Abstract
The ability to grow xylogenic Pinus radiata D.Don in a liquid medium rather than on a solid one would produce a more homogeneous culture, and this in turn would improve cell and gene studies. We report the development of a liquid culture system for two xylogenic P. radiata cell lines and compare the subsequent formation of tracheary elements induced on the conventional solid media. The cell viability (fluorescein diacetate staining) in liquid cultures and subsequent tracheary element (TE) differentiation was as high as, or higher than, that observed with conventional callus cultures on solid media. The growth of cells in liquid culture was confirmed by comparing organic carbon consumption and dry weight increase. Conditions for optimal growth were determined by measuring substrate consumption and cell dry weight with two different cell lines, flask volumes, and starting inoculum densities. Changes to flask volume and cell line were observed to modify substrate carbon consumption within the cell culture, whilst having no significant impact on overall cellular yield. Inoculum density and cell line were the most significant factors affecting the percentage of TE produced. Overall, these preliminary findings confirm that P. radiata xylogenic cells were able to be grown in liquid cultures and did produce TE when induced on solid medium. Therefore, liquid culture has the potential to replace the current standard solid medium system for xylogenic culture of P. radiata.
Highlights
A unique in vitro technique developed at Scion (Wagner et al 2013) induces Pinus radiata D.Don xylogenic callus to differentiate, forming cells with lignified secondary cell walls similar to tracheary elements (TE) (Möller et al 2003)
Overall, these preliminary findings confirm that P. radiata xylogenic cells were able to be grown in liquid cultures and did produce TE when induced on solid medium
The aims of this study were (i) to determine whether or not primary P. radiata xylogenic cells could be cultured in liquid medium and, if so, (ii) test the efficacy of cultured cells in inducing secondary cell walls thereby demonstrating the potential for enhanced biotechnological development of these cell lines
Summary
A unique in vitro technique developed at Scion (Wagner et al 2013) induces Pinus radiata D.Don xylogenic callus to differentiate, forming cells with lignified secondary cell walls similar to tracheary elements (TE) (Möller et al 2003). This system enables candidate genes to be tested in vitro on solid media in order to achieve further genetic gains in P. radiata production. We report the development of a liquid culture system for two xylogenic P. radiata cell lines and compare the subsequent formation of tracheary elements induced on the conventional solid media
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