Increased gelling agent concentration promotes somatic embryo maturation in hybrid larch (Larix×eurolepsis): a 2‐DE proteomic analysis

Abstract

An integrated physiological and proteomic approach was used to investigate the effects of high gellan gum concentration in the medium during maturation of somatic embryos (SE) of hybrid larch, by comparing embryos incubated in media with a high gellan gum concentration (8 g l(-1) ) and the standard concentration (4 g l(-1) ) after 1, 3, 6 and 8 weeks of maturation. Because of the reduced availability of water in the 8 g l(-1) medium, the cultured embryos had a lower osmotic water potential (Ψπ) and water contents, but higher dry weights (DWs), at 8 weeks compared with embryos cultured on the standard medium. The high gellan gum concentration induced a desiccation that is characteristic in zygotic embryo maturation. Total soluble proteins were extracted from SE with trichloroacetic acid (TCA)-acetone after 1 and 8 weeks of maturation on media with 4 and 8 g l(-1) of gellan gum, and separated by two-dimensional gel electrophoresis (2-DE) at pH 4-7. More than 1100 proteins were reproducibly detected on each gel. At 1 and 8 weeks respectively, the abundances of 62 and 49 spots detected in analyses of embryos matured at the two gellan gum concentrations, significantly differed. Among 62 significantly differing spots at 1 week of maturation, the corresponding proteins of 56 were reliably identified by liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS), and were found to be mainly involved in 'carbohydrate metabolism', 'genetic information processing' or 'environmental information processing' according to kegg taxonomy. Both physiological parameters and the proteins identified suggested that the embryos were stressed when they were cultured on 4 g l(-1) of gellan gum.

Different environmental conditions at initiation of radiata pine somatic embryogenesis determine the protein profile of somatic embryos

“…Our results are not in accordance with Teyssier et al (2011) when they reported a higher level of this type of proteins only in somatic embryos produced under high water availability conditions.…”

“…close relatives, according to SwissProt database (Table 1). The fact that for one spot (enolase) more than one protein was identified may be caused by the degradation products of some proteins (Teyssier et al 2011) or by the presence of multiple isoforms of a protein (Lippert et al 2005). The presence of protein isoform with different expressionpatterns during seed maturation has already been reported in barley (Finnie et al 2006).…”

“…One approach to generate such markers and to better understand the regulation of in vitro embryogenesis is to use global transcript expression or protein profiling (Lippert et al 2005;Zhao et al 2015). Indeed, proteome profiling has been successfully applied to the systematic analysis of protein expression in several broad-leaf (Correia et al 2016) and conifer species (Teyssier et al 2011) during the different stages of the development of the somatic embryos. Comparative studies using proteomics to find markers have been carried out by several authors for different aspects of the SE process (Correia et al 2012;Morel et al 2014;Teyssier et al 2011;Vale et al 2014).…”

“…Indeed, proteome profiling has been successfully applied to the systematic analysis of protein expression in several broad-leaf (Correia et al 2016) and conifer species (Teyssier et al 2011) during the different stages of the development of the somatic embryos. Comparative studies using proteomics to find markers have been carried out by several authors for different aspects of the SE process (Correia et al 2012;Morel et al 2014;Teyssier et al 2011;Vale et al 2014). Proteomic approaches have a great potential to study non-model species, because protein sequences have the advantage of being more conserved, making the high-throughput identification of non-model gene products quite effective by comparison to orthologous proteins (Liska and Shevchenko 2003).…”

Short communication: The effect of changing temperature and agar concentration at proliferation stage in the final success of Aleppo pine somatic embryogenesis

“…However, few studies have focused on the impact of temperature (Kvaalen & Johnsen, 2008), and the effect of different concentrations of agar in the culture medium has been studied mostly at maturation stage (Teyssier et al, 2011;Morel et al, 2014). In this sense, in our laboratory García-Mendiguren et al (2016) described in Pinus radiata (D. Don) that different temperatures and water availability conditions at initiation affected subsequent phases of SE; however, this effect on subsequent phases of the process disappeared when different environmental conditions were applied at proliferation.…”