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Student Number 952211010
Author Yi-Ting Ke(_t@)
Author's Email Address kortinn@yahoo.com.tw
Statistics This thesis had been viewed 2020 times. Download 0 times.
Department Graduate Institute of Systems Biology and Bioinformatics
Year 2008
Semester 1
Degree Master
Type of Document Master's Thesis
Language English
Title Characterization of rice group 3 late embryogenesis abundant (LEA) genes expression during development stages and stresses
Date of Defense 2009-01-23
Page Count 117
Keyword
  • development
  • rice
  • stress
  • Abstract Late embryogenesis abundant (LEA) proteins were initially found in the stage of embryo maturation. According to amino acid sequence similarities and conserved motifs, the LEA proteins can be separated into more than five groups. It is known that abscisic acid (ABA), salt, drought or cold stress can induce expression of the lea genes, conferring stress tolerance to plant cells. LEA proteins have been shown to involve in binding and replacement of water molecule, ion sequestration, maintaining of protein or cell membrane structure, and development regulation. Group 3 LEA proteins are characterized by 11-mer amino acid conserved tandem repeats (TEAAKQKAAET) and considered as the stress proteins. Barley HVA1, a member of group 3 LEA proteins, has been reported that it can improve stress tolerance in transgenic plants. However, its exact physiological functions still remain unclear. In this study we used the sequence BLAST to search Group 3 LEA protein orthologues in rice genome database and identify five lea3-like genes. They are AP004018/BAD19162, AP003381/BAB86507, AC073556/AAL84288, AC098833/AAU43988, AP000836/BAD81113 (gene/protein accession number). Here we were trying to characterize their gene sequence organization and expression patterns during developmental stages and environmental stresses. The results showed that these LEA genes played important roles on regulation of plant growth and response for varied stresses, and tissue-specific expression in root, stem, leaf, and seed. Furthermore, cellular localization analysis of the AP004018-GFP, AP003381-GFP, AC073556-GFP, AC098833-GFP, and AP000836-GFP fusion proteins in onion epidermal cells indicated that rice group 3 LEA proteins could be in nucleus (AP003381, AC073556 under ABA or 4J treatment), cytoplasm (AC073556, AC098833 under ABA; AP004018, AC073556, AC098833 under NaCl; AP004018, AC098833 under 4J treatment), or membrane structure (AC073556 under ABA and NaCl; AP004018, AP003381, AC073556, and AC098833 under 4J treatment ), whereas these fusion proteins localize in either nucleus or cytoplasm under non-stress conditions.
    Table of Content Table of contentsKKKKKKKKKKKKKKKKKKKKKKKKKKKKKKI
    Chinese abstractKKKKKKKKKKKKKKKKKK.K.KKKK..KKK..KIII
    English abstract KKKKKKKKKKKKKKKKKK.KKKKK...KKKK.V
    List of abbreviationsKKKKKKKKKKKKKKKKKKKKKKKKK..K....VII
    Chapter 1. IntroductionKKKKKKKKKKKKK..KKK.KK..KKKKKK...1
     Environment and stressesKKKKKKK.KKKK.KKK..KKKK.KKK.1
     Properties of LEA proteinsKKKKKKK..K.KKK.KK..KKKKKK.K...2
     The classification of LEA proteinsKKKKKKKKK...KKKK..KKKKK.2
     Physiological functions of LEA proteinsKKKKKKKKKKKKKKKKK5
     Specific aim of this thesisKKKKKKKKKKKKKKKKKKKKKKK8
    Chapter 2. Materials and MethodsKKKKKKKKKKKKKKKKKKKK10
    2.1 Bioinformatics analysis and data miningKKKKKKKKKKKKKKKK.K.10
    2.2 Plant materialsKKKKKKKKKKKKKKK...KKKKKKKKKKKK..11
    2.3 Total RNA extractionKKKKKKKKKKKKKK...KKKKKKKKKK....11
    2.4 Reverse transcriptional polymerase chain reactionKKKKKKKKKKKK...13
    2.5 Polymerase chain reaction and transformationKKKKKKKKKKKKKK..14
    2.6 Preparation of plasmid DNAKKKKKKK.KKKKKKKKKKKKKK.K18
    2.7 Gel elutionKKKKKKKKKKK.KKKKKKKKKKKKKKKKKK...19
    2.8 Transient expression of promoter activities to different stressesKKKK.KK...20
    2.9 Functional localization analysisKKKKKKKKKKKKKKKKKKKKK.24
    Chapter 3. ResultsKKKKKKKKKKKKKKKKK..KKK.KKKKKKK28
    3.1 Orthologues of HVA1 in rice share same motif sequence and are ABA inducible
    KKKK...KKKKKKKKKKKKKKKKKKKKKKKKKKKKK28
    3.2 Characterization for gene expression of rice group 3 lea genes during
    development stages and stressesKKKKKKKKKKKKKKKKKK......29
    3.3 Analysis of the promoters of rice group 3 lea genes under stressesKK...31
    3.4 LEA-GFP fusion proteins showed different subcellular localization under stress
    treatmentsKKKKKKKKKKKKKKKKKKKKKKKKKKKKK...32
    Chapter 4. DiscussionKKKKKKK.KKKKKKKKKKKKKKKKKK33
     Conserved amino acids were identified in the motif of rice group 3 LEA proteins
    KKKKKKKKKKKKKKKKKKKKKKKKKKKKKKKKK..33
     Analysis of phylogenic tree and evolution of rice group 3 lea genesKKKK.33
     Rice CBF2 gene (RCBF2) as a marker.KKKKKKKKKKKKKKKK..34
     Physiological role of LEAsKKKKKKKKKKKKKKKKKKKKKK.34
     Prospects of LEAsKKKKKKKKKKKKKKKKKKKKKKKKK..35
    Chapter 5. Tables and FiguresKKKKKKKKKKKKKKKKKKKKKK36
    Table 1. Selected genes in this studyKKKKKKKKKKKKKKKKKKKK36
    Table 2. List of primers used in this studyKKKKKKKKKKKKKKKKKK38
    Table 3. Analysis of cis-acting regulatory elements on promoter regions of rice lea 3
    genesKKKKKKKKKKKKKKKKKKKKKKKKKKKK.......40
    Table 4. Profiles of gene expression patternKKKKKKKKKKKKKKKKK42
    Figure 1. Responsiveness of rice lea 3 genes to ABA treatmentKKKKKKKKK46
    Figure 2. Multiple sequence alignment of group 3 LEA proteinsKKKKKKKKK.47
    Figure 3. Analysis of hydropathicity and intrinsic disorder tendency structureKKK49
    Figure 4. Overview of rice Tainung 67 (TNG67) growth and development stagesK53
    Figure 5. Expression patterns of rice lea 3 genes during developmental stages and
    stress treatmentsKKKKKKKKKK..KKKKKKKKKKKKK55
    Figure 6. Promoter activities of rice lea 3 genes analyze by Arabidopsis protoplast
    cellsKKKKKKKKKKKKKKKKKKKKKKKKKKKKKK66
    Figure 7. Subcellular localization of rice LEA- GFP fusion proteins analyze by onion
    epidermal cellsKKKKKKKKKKKKKKKKKKKKKKKKK69
    Chapter 6. ReferencesKKKKKKKKKKKKKKKKKKKKKKKKKK75
    Chapter 7. AppendixKKKKKKKKKKKKKKKKKKKKKKKKKKK80
    I. Gene expression profile: electrophoresis images and normalized dataK80
    (A) Rice grains imbibitions in water with different time courseKKKKKKKKK80
    (B) 20-day-old rice seedling in different stress with time courseKKKKKKKK81
    (C) Rice tillering stage in different stress and time courseKKKKKKKKKK88
    (D) Rice panicle stage in different stress and time courseKKKKKKKKK95
    (E) Seed maturation stage expression dataKKKKKKKKKKKKKK.K102
    II. The promoter region sequences of five lea genesKKKKKKKKKKKKK.104
    III. Description of Cis-acting elementsKKKKKKKKKKKKKKKKKKK..105
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    Date of Submission 2009-02-03

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