Abstract
Key messageiTRAQ based proteomic identified key proteins and provided new insights into the molecular mechanisms underlying somatic embryogenesis in cotton.Somatic embryogenesis, which involves cell dedifferentiation and redifferentiation, has been used as a model system for understanding molecular events of plant embryo development in vitro. In this study, we performed comparative proteomics analysis using samples of non-embryogenic callus (NEC), embryogenic callus (EC) and somatic embryo (SE) using the isobaric tags for relative and absolute quantitation (iTRAQ) technology. In total, 5892 proteins were identified amongst the three samples. The majority of these proteins (93.4%) were found to have catalytic activity, binding activity, transporter activity or structural molecular activity. Of these proteins, 1024 and 858 were differentially expressed in NEC versus EC and EC versus SE, respectively. Compared to NEC, EC had 452 and 572 down- and up-regulated proteins, respectively, and compared to EC, SE had 647 and 221 down- and up-regulated proteins, respectively. KEGG (Kyoto Encyclopedia of Genes and Genomes) analysis indicated that genetic information transmission, plant hormone transduction, glycolysis, fatty acid biosynthesis and metabolism, galactose metabolism were the top pathways involved in somatic embryogenesis. Our proteomics results not only confirmed our previous transcriptomic results on the role of the polyamine metabolic pathways and stress responses in cotton somatic embryogenesis, but identified key proteins important for cotton somatic embryogenesis and provided new insights into the molecular mechanisms underlying somatic embryogenesis in cotton.
Highlights
Upland cotton (Gossypium hirsutum L.) is one of the most important economic crops, and the earliest commercialized transgenic crop worldwide
Total RNA was extracted from non-embryogenic callus (NEC), embryogenic calli (EC) and somatic embryo (SE) of Xinluzao 33 using a modified cetyltrimethyl ammonium bromide (CTAB) method (Luo et al 2003) and was stored at − 80 °C before use
The quality of total RNA was checked on a 1% (w/v) ethidium bromide-stained agarose gel. 1 μg aliquot of total RNA was used for the first-strand cDNA synthesis with the M-MLV reverse transcriptase (TaKaRa) following the manufacturer’s instructions
Summary
Upland cotton (Gossypium hirsutum L.) is one of the most important economic crops, and the earliest commercialized transgenic crop worldwide. Somatic embryogenesis (SEM) provides a technique for gene transformation, and serves as a model system for study of zygotic embryo development in plants (Yang and Zhang 2010; Zimmerman 1993). These efforts identified genes differentially expressed in somatic embryos, highlighted the pathways likely to be involved in SEM and discovered DNA or protein markers for SEM (Mantiri et al 2008). A comparison of proteome dynamics between globular and cotyledonary embryos indicated that stress response, hormone homeostasis, and respiration and photosynthesis determined somatic embryo differentiation (Ge et al 2014) These studies described the expression changes of a large number of genes or proteins during SEM and provided abundant clues for elucidation of the molecular mechanisms underlying SEM.
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