Physiology Mechanism And Differential Protein Of Fraxinus Mandshurica Somatic Embrygenesis Acompanied Explant Browning

Manchrian ash(Fraxinus mandshurica Rupr.) is a hardwood species in the genus Fraxinus in Oleaceae and one of the three valuable hardwood species in Northeast China.In our previous study on somatic embryogenesis of F.mandshurica,we found an interesting phenomenon that the somatic embryos regenerated on the browned explants mostly,which indicated that the browning of explants played an important role on somatic embryogenesis of F.mandshurica.Thus,we carried out the research to study the internal relation between explant browning and somatic embryogenesis by plant growth regulators(PGRs) regulation, physiological and biochemical analysis and proteomics analysis used F.mandshurica immature zygotic embryos cotyledon as material and compared with Syringa reticulate var. mandshurica which is also a species in Oleaceae.The purpose is to open out the relationship between explant browning and somatic embryogenesis and provide a basis for exact regulation of explants and culture condition and provide an academic basis for tree somatic embryogenesis as well.The results were as follows:(1) Explant browning rate of F.mandshurica during somatic embryogenesis was 67.03% and the somatic embryogenesis rate was 37.22%,the somatic embryogenesis rate of nonbrowned explants was 34.06%which was higher than that of non-browned one(3.16%).(2) Somatic embryos of S.reticulate were induced by using immature single cotyledon as explants and the best induction medium,maturation medium,germination medium and rooting medium were determined.The browning rate of explants was 30.00%according to the best induction medium,which is lower than that of F.mandshurica,and the somatic embryos produced on non-browned explants mostly.(3) Polyphenol content and Pholyphenol Oxidase(PPO) activity between the two species were not significant,but the browning rate were significant,which might relate with the district distributing of the two substances.(4) The PGRs affected explant browning and somatic embryogenesis of F.mandshurica significantly,and also of S.reticulate somatic embryogenesis,but not significantly on explant browning;explants browning of the two species affected somatic embryogenesis significantly, but the effects were different.(5) The difference of SOD,CAT and POD activity in different explants showed that the browned explants had higher potential to produce somatic embryos than that of non-browned explants was because of its higher H₂O₂ content,which promoted somatic embryogenesis; polyphenol and PPO played a role on somatic embryogenesis but the mechanism was unknown.(6) Species and content of protein were different in different explants;the results of 2-DE image analysis and MS/MS identification indicated that molecular weight and pI of experimental and theoretical were different in some protein spots;in identified protein spots, osmotin-like protein,heat shock protein and peroxidase are likely to relate with explant browning and others just as 7S globulin,dehydrin as well as osmotin-like protein are likely to relate with somatic embryogenesis.