Abstract
Somatic embryogenesis (SE), which resembles zygotic embryogenesis, is an essential component of the process of plant cell differentiation and embryo development. Although microRNAs (miRNAs) are important regulators of many plant develop- mental processes, their roles in SE have not been thoroughly investigated. In this study, we used deep-sequencing, computational, and qPCR methods to identify, profile, and describe conserved and novel miRNAs involved in longan (Dimocarpus longan) SE. A total of 643 conserved and 29 novel miRNAs (including star strands) from more than 169 miRNA families were identified in longan embryogenic tissue using Solexa sequencing. By combining computational and degradome sequencing approaches, we were able to predict 2063 targets of 272 miRNAs and verify 862 targets of 181 miRNAs. Target annotation revealed that the putative targets were involved in a broad variety of biological processes, including plant metabolism, signal transduction, and stimulus response. Analysis of stage- and tissue-specific expressions of 20 conserved and 4 novel miRNAs indicated their possible roles in longan SE. These miRNAs were dlo-miR156 family members and dlo-miR166c* associated with early embryonic culture developmental stages; dlo-miR26, dlo-miR160a, and families dlo-miR159, dlo-miR390, and dlo-miR398b related to heart-shaped and torpedo- shaped embryo formation; dlo-miR4a, dlo-miR24, dlo-miR167a, dlo-miR168a*, dlo-miR397a, dlo-miR398b.1, dlo-miR398b.2, dlo-miR808 and dlo-miR5077 involved in cotyledonary embryonic development; and dlo-miR17 and dlo-miR2089*-1 that have regulatory roles during longan SE. In addition, dlo-miR167a, dlo-miR808, and dlo-miR5077 may be required for mature embryo formation. This study is the first reported investigation of longan SE involving large-scale cloning, characterization, and expression profiling of miRNAs and their targets. The reported results contribute to our knowledge of somatic embryo miRNAs and provide insights into miRNA biogenesis and expression in plant somatic embryo development.
Highlights
Longan (Dimocarpus longan Lour.) is a tropical/subtropical fruit tree in the family Sapindaceae
The longan somatic embryogenesis (SE) system has been widely used as a model system for investigating in vitro and in vivo regulation of embryogenesis in woody plants, most studies of longan SE have focused on a few selected aspects related to cell biology, molecular biology, and proteomics [1,2,3,4,5]
To identify miRNAs involved in longan SE, a pooled sRNA library was generated from embryogenic cultures and subjected to Solexa sequencing
Summary
Longan (Dimocarpus longan Lour.) is a tropical/subtropical fruit tree in the family Sapindaceae. In this species, embryo development status strongly influences seed size, fruit quality, fruit set, and yield. To improve longan fruit quality and yield, researchers have focused much attention on mechanisms of longan embryonic development. The longan somatic embryogenesis (SE) system has been widely used as a model system for investigating in vitro and in vivo regulation of embryogenesis in woody plants, most studies of longan SE have focused on a few selected aspects related to cell biology, molecular biology, and proteomics [1,2,3,4,5]. There are no reported studies devoted to identification and expression of microRNAs (miRNAs)–an important group of plant regulators– during longan SE
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