Establishiment of molecular markers for early selection of embryogenic cultures with high embryogenic potential in brazilian pine (Araucaria angustifolia (BERT) O. KTZE)

Background Brazilian pine (Araucaria angustifolia (Bert) O. Ktze) is the only native conifer species with economic importance in Brazil. Recently, due to intensive exploitation Brazilian pine was included in the official list of endangered Brazilian plants. Biotechnology tools, like somatic embryogenesis (SE), may become a potentially useful tool for mass clonal propagation and ex situ conservation of commercial and endangered plant species, especially conifers. SE involves the coordinated execution of four steps (embryogenic culture (EC) induction, proliferation, maturation, and plant regeneration). As observed for other conifers, the presence of well-developed early somatic embryos (SE) in EC of Brazilian pine can be considered the pre-requisite for embryo maturation in a medium supplemented with abscisic acid (ABA) and osmotic agents. However, in some genotypes even the presence of bipolar SE does not guarantee embryo maturation. Since SE morphology cannot be used as the only factor for EC selection, the development of molecular markers for early detection of embryogenic cultures responsive to maturation promoters (ABA and osmotic agents) is highly desirable. Polyamines (putrescine (Put), spermidine (Spd), and spermine (Spm)) have been classified as plant growth regulators and hormonal second-messengers playing a critical role in various growth and developmental processes in plants, such as the differentiation and development of somatic embryos. The relation Put/Spd has demonstrated the best answers for predicting embryogenic potential in different plant species. Apart from polyamines quantification, the analysis of gene expression has been used to detect the expression of embryogenesis regulating genes like somatic embryogenesis receptor kinase (SERK), wuschel-related WOX (WOX), and ABA insensitive-1 (ABI1) during conifer somatic embryogenesis. In order to develop molecular markers for early detection of EC with high embryogenic potential in Brazilian pine, we measured the polyamine content (free and conjugate) and the expression of three embryogenesis-regulating genes (SERK, WOX and ABI1) during the proliferation phase of ECs with different maturation capabilities.