Physiological And Proteomic Studies Of Maize Seed Vigor

The viability (vigor) of maize (Zea mays) cv Zhengdan 958 was evaluated in term of germination rate, TTC staining and controlled deterioration test (CDT). Proteomic changes of maize seeds with different vigor were compared in order to identify the potential protein marker for seed viabilily. The main results are represented below.1 Physiological papameters changed obviously after CDT.Seed vigor reduced with CDT days. The germination potential and rate of seeds aged for 1 d decreased by 7% and 3%, respectively; the germination potential and rate of the seeds aged for 2 d and 3 d sharply decreased by 74% and 64%, respectively; aged 6 d seeds was not able to germinate. Moreover, germination potential was more sensitive than germination rate with respect to CDT. TTC staining showed that natural and artificial aged seeds could be categorized into deep, light and non-stained types. However, compared with germination test, TTC method gave a little high vitality of seeds.MDA content increased gradually with CDT days, while soluble sugar content in seeds after CDT for 2 d was highest. Anti-oxidation system of SOD, CAT, and POD activity decreased with CDT days. SOD and CAT activities declined with CDT days, while POD activity did not change significantly in the first 2 d of CDT. Similarly, soluble protein content decreased during CDT.2 Seed proteome, especially the embryo proteome, changed markedly after CDT compared to controls.SDS-PAGE showed that CDT promoted protein degradation, and also induced some protein in amount and protein synthesis. Endosperm and embryo differred in protein expression. Many proteins in the embryo changed in abundance, and some new protein bands emerged. Particularly, in embryos with different vigor,14 kDa protein changed dramatically.Given the significant difference in embryo protein profiles between aged-for-6d seeds and controls, embryo protein were extracted and combared using 2-DE (two-dimensional electrophoresis). We found that 4 protein spots, among ca.750 spots, changed 3 times in abundance, which were subjected to mass spectrometry for protein identification. Homology alignment showed that these four proteins are LEA EMB564 (Swissprot accession no. P46517), expressed protein (NCBI accession no. NP₀01142211), lipoprotein (Swissprot accession no. ACG48482), and glucose and ribotol dehydrogenase (NCBI accession no. NP₀01140439).3 EMB564 in scutellum is a potential protein marker for mazie seed vigor.To further localize EMB564, plumule, radicle and scutellum were separated from embryos, and protein was extracted for each parts and used for electrophoretic analysis. SDS-PAGE showed that EMB564 in the plumule, radicle and scutellum were different in abundance between aged and control seeds. Scutellum is the major part of embryo and easy to be separared, so in germination experiments, scutellum proteome of different seed vigor were studied comparatively. During seed germination, EMB564 was relative abundant in the scutellum of weak vitality, while the band was almost not visible in the embryos with high vitality.The hydrophilicity and hydrophobicity, homology and the secondary, tertiary structures of EMB564 were predicted using bioinformatic tools. After homologous alignment with known LEA sequences, EMB564 was identified as a member of group-1 LEA family. The 3-D structure of EMB564 resembled DNA-binding proteins, implying that EMB564 functions via binding DNA during seed maturation. Currently, physiological functions of almost LEA proteins are not yet clear. This thesis, for the first time, provided physiological and proteomic data that EMB564 is related to seed vigor, and possibly used as the molecular marker for seed vigor of maize cv Zhengdan 958.