Abstract
In vitro organogenesis, one of the most common pathways leading to in vitro plant regeneration, is widely used in biotechnology and the fundamental study of plant biology. Although previous studies have constructed a complex regulatory network model for Arabidopsis in vitro organogenesis, no related study has been reported in ramie. To generate more complete observations of transcriptome content and dynamics during ramie in vitro organogenesis, we constructed a reference transcriptome library and ten digital gene expression (DGE) libraries for illumina sequencing. Approximately 111.34 million clean reads were obtained for transcriptome and the DGE libraries generated between 13.5 and 18.8 million clean reads. De novo assembly produced 43,222 unigenes and a total of 5,760 differentially expressed genes (DEGs) were filtered. Searching against the Kyoto Encyclopedia of Genes and Genomes Pathway database, 26 auxin related and 11 cytokinin related DEGs were selected for qRT-PCR validation of two ramie cultivars, which had high (Huazhu No. 5) or extremely low (Dazhuhuangbaima) shoot regeneration abilities. The results revealed differing regulation patterns of auxin and cytokinin in different genotypes. Here we report the first genome-wide gene expression profiling of in vitro organogenesis in ramie and provide an overview of transcription and phytohormone regulation during the process. Furthermore, the auxin and cytokinin related genes have distinct expression patterns in two ramie cultivars with high or extremely low shoot regeneration ability, which has given us a better understanding of the in vitro organogenesis mechanism. This result will provide a foundation for future phytohormone research and lead to improvements of the ramie regeneration system.
Highlights
In vitro organogenesis, one of the most common pathways leading to in vitro plant regeneration, is used in biotechnology and the fundamental study of plant biology [1,2]
Adventitious shoot buds regenerated from most of the Huazhu NO. 5 (H5) petiole explants (Figure 1E), whereas few developed on the DZ petiole explants after 35 days of culturing (Figure 1J)
From the 5,760 differentially expressed genes (DEGs) in H5 libraries, we identified 248 transcription factors (TFs) mRNAs in 36 published TF families in the Database of Arabidopsis Transcription Factors (DATF) [49]
Summary
One of the most common pathways leading to in vitro plant regeneration, is used in biotechnology and the fundamental study of plant biology [1,2]. Organogenesis is a multistep process consisting of callus formation, adventitious organ formation and micropropagation using axillary or apical meristem containing tissues [3]. Morphological, biochemical and molecular methods have been used to investigate the mechanisms behind phytohormone signaling and gene regulation during in vitro organogenesis in several plants, such as Arabidopsis [5], poplar [3] and almond [6]. Previous studies have constructed a complex regulatory network model for in vitro organogenesis, for Arabidopsis [2,7,8]. The results from our experiments will build on these previous studies
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