Читать книгу Biotechnology of Fruit and Nut Crops - Группа авторов - Страница 143

Since the introduction of oil palm tissue culture in the 1970s, clonal propagation has proven to be useful, not only in producing uniform planting materials (Rival and Jaligot, 2010), but also in the development of research in genetic engineering (Parveez et al., 2015). The scaling up to efficient commercial production has been successfully achieved in several producing countries, e.g. Indonesia, Malaysia and Costa Rica, with clonal material slowly making its way into the catalogues of seeds suppliers. Even if it is still difficult to get reliable figures, as clonal material is more often kept for internal uses by plantation companies which produce it and not sold in the seed market, the proportion of clonal planting material might not exceed 3% of the present offer. Currently the major limiting factors that continue to hamper the commercial take-off of clonal planting material are the cost of production and the risk of mantled somaclonal variation (Rival and Jaligot, 2010).

A mathematical model for the growth and conversion of somatic embryos was developed by Konan et al. (2006) to monitor the large-scale production of oil palm microplants. The predicted biomass of somatic embryos obtained and subcultured (B_n), together with the number of harvested shoots (Sh_n) – two key parameters for production forecasts – have been modelled for seven different shoot harvesting procedures. For the four different clonal lines studied, observed differences between experimental B_n values at the end of each culture cycle and their theoretical counterpart generated by mathematical models were found to range from −30% to +14% at the end of the first 6-week culture cycle, then from −50% to +70% after the sixth subculture (36 weeks). With respect to the predicted number of shoots harvested after conversion of somatic embryos (Sh_n), average variations between experimental and theoretical values ranged between −45% and +41%. Predicted values for biomass (B_n) between two culture cycles were found to vary slightly (+6% to +10%), indicating that the production of somatic embryo biomass, as predicted by the model, was rather stable, for a given clonal line, from one 6-week cycle to another. The established model could thus be regarded as valid, and the variations observed for B_n and Sh_n were found to be acceptable when compared to those described by other models. Taken as a whole, predicted values for the two studied production parameters were in agreement with the corresponding experimental data (correlation = 0.98).

In order to synchronize somatic embryogenesis and automate the production process of secondary somatic embryos, simple bioreactors based on the temporary immersion systems (Steinmacher et al., 2011) should be assessed, as they proved to be successful in improving the in vitro regeneration of peach palm (Bactris gasipaes) through secondary somatic embryogenesis.

Despite considerable progress in improving in vitro regeneration, callus initiation and somatic embryogenesis rates from proliferating callus cultures remain very low and mostly unpredictable. Thus, understanding the gene diversity and expression profiles in oil palm cell cultures is critical for increasing the efficiency of these processes. Low et al. (2008) generated a total of 12 standard cDNA libraries, representing three main developmental stages of oil palm embryogenic cultures. Random sequencing of clones from these cDNA libraries generated 17,599 ESTs. The ESTs were analysed, annotated and assembled to generate 9584 putative unigenes distributed in 3268 consensi and 6316 singletons. These unigenes were assigned putative functions based on similarity and gene ontology annotations. Cluster analysis, which surveyed the relatedness of each library based on the abundance of ESTs in each consensus, revealed that lipid transfer proteins were highly expressed in embryogenic tissues. A glutathione S-transferase was highly expressed in non-embryogenic callus. Further analysis of the unigenes identified 648 non-redundant SSRs and 211 putative full-length open reading frames. However, in order to confirm whether these genes are suitable early markers for embryogenic cultures, the genes need to be tested on earlier stages of initiation and induction and with a wider range of genotypes. This collection of ESTs is an important resource for genetic and genome analyses of the oil palm.

Konan et al. (2010) conducted a 20-year study to assess the problems involved in subculturing over a very long period. Oil palm somatic embryos were maintained in vitro on a Murashige and Skoog (1962) basal medium without growth regulators. Analyses of the proliferation rate of the embryogenic cultures, along with the survival rate of the regenerated plantlets after their transfer into soil, and of the flowering of the derived adult palms were conducted for cultures maintained in vitro for 1–20 years. From the ninth year of maintenance, the tissue quality of the somatic embryos gradually declined. After >20 years, 30% of the 20 clones tested still continued to proliferate satisfactorily on the same maintenance medium. Even though age of the lines suppressed survival capacity of plants under natural conditions, it is noteworthy that among the clones originating from 20-year-old cultures only eight (40%) exhibited the ‘mantled’ floral abnormality.