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Proteomics and Gene Expression Analysis of a Leaf Color Mutant in Wheat

Author: SongSuJie
Tutor: ZhaoBaoCun; LiuLuXiang
School: Hebei Normal
Course: Genetics
Keywords: wheat(Triticum aestivum L.) leaf color mutant Space mutagenesis 2D-Difference gel electrophoresis(2D-DIGE) Chlorophyll fluorescence kinetic parameters Real-time PCR (qPCR)
CLC: S512.1
Type: Master's thesis
Year: 2012
Downloads: 50
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Abstract


Proteomics approach and gene expression analysis were used to characterize achlorophyll-deficient wheat mutant Mt6172,which derived from spaceflight mutagenesis,compared with its wild type H6172, in order to get a better understanding of the reveal themolecular mechanism of leaf color mutantion in Mt6172.The main results were as following:1. Through2D-DIGE and MALDI-TOF/MS analysis on total protein of seedling leafbetween mutant Mt6172and its wide type H6172,62differentially expressed proteins wereidentified, among them50proteins down-regulated and12proteins up-regulated.These62differentially expressed proteins can be classified into29types and11functional groups,including the parts of subunits or Assembly protein of PSI, PSII, NAD (P) H dehydrogenasecomplex, ATP synthase, Calvin-Benson cycle and antioxidant mechanism.87.1%of thedifferentially expressed proteins located in the chloroplast, and64.5%involved inphotosynthesis.2. Based on an improved method for separation of wheat chloroplast and extraction ofwheat chloroplast protein,2D-DIGE and MALDI-TOF/MS analysis were used to identify thechloroplast proteins of wheat seedling.54differentially expressed proteins were identified,among them34proteins down-regulated and20proteins up-regulated in the mutant. These54differentially expressed proteins can be classified into37types and15functional groups,including the parts of subunits or Assembly protein of PSI,PSII,NAD (P) H dehydrogenasecomplex.51.9%of the differentially expressed proteins were involved in photosynthesis and40.7%were located in the chloroplast thylakoid membrane and involved in the light reactionpathway.3. Analysis of the expression and regulation mode of13light-reactive protein-relatedgenes of differentially expressed proteins located in the chloroplast thylakoid membraneshowed that almost all light-reactive protein genes in the mutant were down-regulated. Thisresult was consistent with the above2D-DIGE proteomics analysis. But in the process ofproplastid differentiation, the expression level of most of the genes were increased in differentdegrees in the mutant, implying the possible different modes of expression and regulation oflight response-related genes in different growth conditions.4. Determination of chlorophyll fluorescence parameters showed that, with the growthof the exposure time, the photosynthetic capacity of wild type gradually recovered, but thephotosynthetic capacity of mutant was extremely weak even in the latter of the exposure time period and close to zero. The PSII activity of the wild type is gradually increased with thegrowth of exposure time, but the PSII activity in the mutant can hardly be detected. Thisresult suggested that after exposure in the light, the mutant thylakoid membrane structureformation was blocked, resulting the light reaction complex formation failed.It was concluded that chlorophyll-deficient mutation in Mt6172is an extremely complexprocess, the defect of light reactive protein complex of thylakoid membrane may be theimportant reason of mutation. This study result is helpful to further clarify the molecularmechanism of the wheat chlorophyll deficiency mutation derived from spaceflightmutagenesis.

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