Role of Pectinesterase Isoforms During Tomato Fruit Ripening

番茄果实成熟中的果胶甲酯酶

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Author: Wen Bo
Language: English
ISBN/ISSN: 9787511624116
Published on: 2016-01
Soft Cover

Role of Pectinesterase Isoforms During Tomato Fruit Ripening,《番茄果实成熟中的果胶甲酯酶》主要介绍的是作者在英国研究期间从事园艺植物番茄生理生化、遗传育种、果实采后贮藏加工等生物学的研究与实验等方面所取得的新成果。《番茄果实成熟中的果胶甲酯酶》具有实用价值和参考价值。温波博士:安徽农业大学园艺学院副教授,硕士生导师,2003年5月至2004年5月在英国诺丁汉大学植物科学系做访问研究;2005至2009年在英国诺丁汉大学生物科学学院攻读博士;2009至2012年在卡迪夫大学从事博士后研究;2013年至今在安徽农业大学园艺学院工作。


CHAPTER1 INTRODUCTION
1.1Fruit
1.1.1 General introduction
1.1.2 Tomato
1.2 Fruit riipening
1.2.1 Colour, flavour and texture changes in fruit ripening
1.3 Plant cell wall
1.3.1 General introduction
1.3.2 Cell wall components
1.3.3 Cell wall structure
1.4 Cell wall enzymes
1.4.1 Polygalacturonase(PG)
1.4.2 Pectate lyases
1.4.3 β—Galactosidase
1.4.4 Expansin
1.5 Pectinmethylesterase
1.5.1 PE proteins
1.5.2 PE modes of action and regulation
1.5.3 PE genes and isoforms
1.5.4 Tomato PE antisense lines
CHAPTER 2 MATERIALS AND METHODS
2.1 Plant material
2.1.1 Tomato growth and maintenance
2.1.2 Crossing tomato
2.2 Chemicals
2.3 Fruit texture analysis
2.4 Biochemical methods
2.4.1 Extraction of crude protein from tomato fruits
2.4.2 Bio—Rad (Bradford) protein assay
2.4.3 Pectinesterase assay
2.4.4 Heparin affmity chromatography
2.4.5 PE profile assay
2.4.6 Concentration ofpooled PE isoforms
2.4.7 Totalleafprotein extraction for SDS PAGE analysis
2.4.8 Making acetone insoluble solid (AIS)
2.4.9 Determination ofdegree ofesterification (DE) of the pectin by titration
2.4.10 Extraction of CDTA Soluble Pectin
2.4.11 Determination of Neutral Sugars
2.4.12 Determination of uronic acid
2.5 Molecular biological methods
2.5.1 DNA extraction from tomato leaves
2.5.2 DNA measurement
2.5.3 Polymerase chain reaction (PCR)
2.5.4 Restriction enzyme digestion
2.5.5 Agarose gel electrophoresis
2.5.6 DNA sequence (MWG value read)
2.5.7 PCR product purification by Perfectprep Gel Cleanup Kit (Eppendorf)
2.6 Immunochemistry and immunolabelling
2.6.1 Immunodot assay
2.6.2 Immunolocalization
2.7 Statistics
2.8 Capillary Electrophoresis(CE)
2.8.1 Enzyme digests
2.8.2 CE
CHAPTER 3 GENERATION OF DOUBLE ANTISENSE PLANTS
3.1 Plant generation
3.2 Generation of the double antisense plants
3.3 PE Isoform profiles ofdouble antisense plants
3.3.1 PE extraction method optimisation
3.3.2 PE profiles in tomato fruit
3.4 Summary
CHAPTER 4 CHARACTERIZATION OF PE SINGLE AND DOUBLE ANTISENSE PLANTS
4.1 Fruit PE activity and morphology analysis
4.1.1 Generation of wild—type controls
4.1.2 Fruit PE activities
4.1.3 Fruit morphology and softening
4.2 Cell wall component analysis
4.2.1 UA in acetone insoluble solids (AIS) and CDTA soluble pectin 4.2.2 DE ofpectinin AIS
4.2.3 Neutral sugar analysis by GC
4.3 Capillary electrophoresis (CE) ofpectin
4.3.1 DE of CDTA soluble pectin
4.3.2 CE profiles of PG digested CDTA soluble pectin
4.4 Characterization of antisense lines using monoclonal antibodies
4.4.1 Immuno—dot blot analysis of CDTA soluble pectin
4.4.2 Immunolocalization: Fruit pericarp
4.4.3 Immunolocalization: Stem
4.5 Summary
4.5.1 Biochemical and morphological study
4.5.2 Immunological study
CHAPTER 5 USE OF ANTISENSE LINES FOR PEISOFORMS ANALYSIS
5.1 Introduction
5.2 Effect of pH on PE activity
5.3 PEisoform profiling
5.3.1 Fruit
5.3.2 Stem
5.3.3 Leaf
5.4 Summary
CHAPTER 60VERALL DISCUSSION
6.1 Role of PE during fruit ripening
6.2 PE isoforms in vegetative tissues
6.3 Unstable PEl expression in double antisense lines
6.4 Future work
REFERENCES





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