Gene Expression And Proteomics Analysis Of Intact Somatic Embryogenesis In An Elite Inbred Line Of Maize (Zea Mays L.)

Maize(Zea mays L.) is one of world’s most vital crops, the demand is increasing constantly. Plant transformation technology and traditional breeding technique have become the strategy to increase crop production particularly in maize. Plant somatic embryogenesis is a useful tool to propagate transgenic organisms. This research conducted a series of morpho-anatomical characterization, transcription and protein for somatic embryogenesis in maize. The main results are as follows:Histological analysis and scanning electron micrographs confirmed the different developmental stages of intact somatic embryos from monocotyledon explants. We found proembryos, globular-shaped embryo, pear-shaped embryo, scutiform embryo and mature embryo during somatic embryogenesis. And is similar to zygotic embryogenesis.c DNA-AFLP(c DNA-Amplified Fragment Length Polymorphism) for transcriptome profiling and differentially expressed gene cloning between embryogenic and non-embryogenic of the inbred maize line Y423 during the process of somatic embryogenesis. A total of 6583 TDFs were found, approximately 2297 differentially expressed TDFs were identified, including 1023 bands detected up-regulated and 1274 bands displayed down-regulated in embryonic calli compared to in non-embryonic calli. The functional annotation of about 117 candidate genes retrieved in the NCBI databases was carried out by using Go terminology. It consisted in three structured vocabularies describing genes, transcripts and proteins of any organism in terms of their associated biological process, cellular component and molecular function in a species-independent manner. In the biological process, the most of genes distributed in cellular process(25%), metabolic process(24%) and response to stimulus(11%). In the cellular component, the most of genes distributed in plastid(24%), nucleus(21%), mitochondrion(15%) and cytosol(13%). In the molecular function, the most of genes distributed in organic cyclic compound binding(26%), heterocyclic compound binding(26%) small molecule binding(14%), transferase activity(14%) and hydrolase activity(14%).The results showed that somatic embryogenesis is a complex biological process, and the genes need control and collaboration between them.Using RT-PCR, two full-length candidate genes were cloned which may play an important role during somatic embryogenesis, it was named as Zm SUF4 and Zm DRP3 A. The gene of Zm SUF4 encoding a 358-amino acids protein, domain analysis using smart showed that it had two Zn F_C2H2(C2H2-type(classical) zinc fingers(Znf)) domains, and the hydrophobic character prediction based on the Kyte–Doolittle scale indicates that Zm SUF4 hydrophilic with a grand average of hydropathicity at-0.233. The gene tree view showed that Zm SUF4 is closely related with Sorghum bicolor, Setaria italic and Oryza sativa japonica. The gene of Zm DRP3 A encoding a 821-amino acids protein, domain analysis using smart showed that it had one DYNc( Dynamin, GTPase) domain and a GED( Dynamin GTPase effector) domain, and the hydrophobic character prediction based on the Kyte–Doolittle scale indicates that Zm DRP3 A hydrophilic with a grand average of hydropathicity at-0.300. The gene tree view showed that Zm DRP3 A is closely related with Sorghum bicolor, Setaria italic and Oryza sativa.i TRAQ(Isobaric tags for relative and absolute quantitation) for proteomics profiling and enrichment of differentially expressed analysis between embryogenic and non-embryogenic of the inbred maize line Y423 during the process of somatic embryogenesis. A total of 5592 proteins were found, with close to 57% of the protein contains at least two peptides,approximately 632 differentially proteins were identified, including 366 up-regulated proteins and 266 down-regulated proteins detected in embryonic calli.The enrichment analysis for differentially proteins using COG(Cluster of Orthologous Groups of proteins), WEGO and pathway. In the COG analysis, the most of up-regulated proteins distributed in Carbohydrate transport and metabolism; Translation; ribosomal structure and biogenesis; Posttranslational modification, protein turnover,chaperones; and General function prediction only. The most of down-regulated proteins distributed in Energy production and conversion; Amino acid transport and metabolism; Lipid transport and metabolism; and General function prediction only. The Chromatin structure and dynamics; Cell cycle control, cell division, chromosome partitioning and Transcription only contained up-regulated proteins. In the WEGO analysis, the differentially proteins distributed in cellular component, molecular function and biological process. In the cellular component, the most of proteins distributed incell, cellpart and organelle. In the molecular function, the most of proteins distributed in binding and catalytic. In the biological process, the most of proteins distributed in cellular process and metabolic process. Extracellular region part and Multi\-organism process only contained up-regulated proteins. In the pathway analysis, the up-regulated proteins make more important role than the down-regulated proteins in Ribosome,. In ribosome, Starch and sucrose metabolism, and glycolysis. The increase of ribosome may contribute to the synthesis of proteins involved in the somatic embryogenesis. Starch and sucrose metabolism, and glycolysis play key roles in the somatic embryogenesis, proteins associated with the synthesis of beta-glucosidase, endoglucanase, enolase and phosphoenolpyruvate carboxykinase are highly expressed.