Regulation of Somatic Embryo Development in Norway Spruce

Methods Mol Biol. 2020;2122:241-255. doi: 10.1007/978-1-0716-0342-0_17.

Abstract

Somatic embryogenesis in Norway spruce combined with reverse genetics can be used as a model to study the regulation of embryo development in conifers. The somatic embryo system includes a sequence of developmental stages, which are similar in morphology to their zygotic counterparts. The system can be sufficiently synchronized to enable the collection and study of a large number of somatic embryos at each developmental stage.Here we describe a protocol for establishing transgenic cell lines in which genes of interest are upregulated or downregulated. Furthermore, we present methods for comparing embryo morphology and development in transgenic and control cell lines, including phenotyping the embryos, histological analysis, and tracking embryo development. The expression pattern of different genes is determined by GUS reporter assays.

Keywords: Morphological analysis; Norway spruce; Reporter gene expression; Somatic embryogenesis; Tracking embryo development; Transgenic cell lines.

MeSH terms

  • Cell Line
  • Embryo Culture Techniques / methods
  • Gene Expression Regulation, Plant
  • Genes, Reporter
  • Microscopy, Confocal / methods
  • Phenotype
  • Picea / embryology*
  • Picea / genetics
  • Plants, Genetically Modified / embryology
  • Plants, Genetically Modified / genetics
  • Seeds / embryology*
  • Seeds / genetics