Epsilon Archive for Student Projects

Abstract

Somatic embryogenesis (SE) is an in vitro clonal propagation method with great potential for conifer tree propagation but hampered by low yields of plants relative the zygotic embryogenesis (ZE) process in seeds. Previous studies with transcript profiling indicates that the somatic embryo does not fully mature to the same extent as the zygotic embryo. The underlying reasons for the suboptimal somatic embryogenesis process is not understood. Here the involvement of miRNA in the regulation of the two embryogenesis processes was investigated by comparing mRNA and miRNA expression data from both ZE and SE in search for miRNA target genes of known importance to embryogenesis.
In the primary study, mRNA and miRNA expression datasets from Picea abies ZE and SE samples were analysed to assess potential differences between ZE and SE in miRNA gene regulation. By filtering for strong miRNA – mRNA matches using miRNAs exclusively identified in ZE, 46 specific target genes could be identified of which five genes were notably different in expression when comparing heatmaps for the specific developmental stages in ZE and SE previously identified as key steps differing between ZE and SE. The two genes with the most interesting differential expression pattern, PA_chr08_G003877 and PA_chr12_G000025, were selected for Weighted Gene Co-expression Network Analysis and Gene Ontology enrichment searches. The results suggest that these genes are important for proper embryo development, however further functional validation of these candidate genes by a transgenic approach is required.
A subtask to this study was to review the expression patterns of the conserved microRNAs miR156 and miR529 during ZE and SE. These miRNAs are known to control the SQUAMOSA BINDING PROTEIN‐LIKE (SPL) genes, which drive adult development and flowering and are of key importance for efforts to reduce the cone-setting age of Picea abies. In the future, miR156 and miR529 may be utilized to impact the flowering process by molecular modifications which takes place during SE with methods like the miRNA mimicry method.
MikroRNA156 and miR529, known to control SPL expression, were analysed during ZE and SE and revealed patterns of combined miR156/529 negative regulation of the SPL genes. A higher abundance of miRNAs 156 and 529 variants could be detected in the SE data, possibly linked to the stress-inducing in vitro environment.
This study provides a short list of candidate miRNAs controlling potential key regulatory genes in the embryogenesis process in Picea abies. It also provides an overview of the miR156 and miR529 expression patterns during ZE and SE.