Functional Characterization of Tomato Curl3 Gene in Metaboli
發(fā)布時(shí)間：2024-06-01 18:57
油菜素內(nèi)酯(BRs)在植物生長(zhǎng)發(fā)育中起著重要作用。然而,在園藝作物中,BR信號(hào)介導(dǎo)的植物生長(zhǎng)調(diào)控的分子生物學(xué)證據(jù)仍然不足。BR信號(hào)轉(zhuǎn)導(dǎo)與其他植物激素之間的互作共同調(diào)控植物生長(zhǎng)發(fā)育。激素、環(huán)境信號(hào)和發(fā)育過(guò)程之間具有復(fù)雜互作機(jī)制,共同調(diào)控植物生長(zhǎng)發(fā)育。本研究將有助于了解BR信號(hào)在植物生長(zhǎng)發(fā)育調(diào)控中的作用。在本研究中,為了闡明BR信號(hào)傳導(dǎo)對(duì)番茄生長(zhǎng)發(fā)育的可能影響,我們基于CRISPR系統(tǒng)創(chuàng)建了BR不敏感型番茄curl3突變體。但是該突變體與對(duì)照的高度差異,我們同時(shí)從TGRC引進(jìn)一個(gè)在curl3位點(diǎn)的等位基因突變體(abs1)。與curl3突變體相比,abs1突變體表型變化相對(duì)微弱。在本研究中,我們闡明了Sl Curl3(番茄中關(guān)鍵BR信號(hào)轉(zhuǎn)導(dǎo)基因)在調(diào)節(jié)植物生長(zhǎng)發(fā)育、初級(jí)代謝、激素平衡中的功能。主要結(jié)果如下:1)在本研究中,從番茄中分離了一個(gè)BRI1(油菜素內(nèi)酯不敏感1基因)的同源基因Curl3,并基于CRISPR創(chuàng)建了敲除突變體,對(duì)其功能進(jìn)行研究。在植物中BR信號(hào)突變體的功能喪失會(huì)導(dǎo)致其生理生化產(chǎn)生重要變化。curl3突變體表現(xiàn)出極度矮化,伴隨著卷曲葉片以及生化和代謝途徑的改變。此外,在對(duì)...
【文章頁(yè)數(shù)】：110 頁(yè)
【學(xué)位級(jí)別】：博士
【文章目錄】：
Abstract
摘要
List of Abbreviations
1 Introduction
1.1 Plant Hormones
1.2 Brassinolide Hormone
1.3 Brassinolide Crosstalk with other Hormones
1.4 Brassinolide Perception by BRI1
1.5 BR-Dependent Gene Expression
1.6 BR-Signaling Mutant
1.7 Objectives of This Research
2 Brassinolide Signaling Disruption Regulates Primary Metabolism in Accordance with Phytohormone Production
2.1 Materials and Methods
2.1.1 Plant Material
2.1.2 Gene Isolation
2.1.3 CRISPR/Cas9 Gene Editing
2.1.4 Morphological Characterization and Anatomical Observations
2.1.5 Hormone Quantification
2.1.6 Tomato Primary Metabolite Profiling
2.1.7 Quantitative Gene Expression Profiling
2.1.8 Statistical and Bioinformatics Analysis
2.2 Results
2.2.1 Sequence analysis of Curl3 from Solanum lycopersicum
2.2.2 CRISPR based Gene Editing
2.2.3 Altered Expression of BR Signaling Components
2.2.4 Morphological Characterization and Anatomical Observations of curl3 Mutant
2.2.5 BR-Insensitivity Alters Phytohormone Production in Transgenic Tomato Plants
2.2.6 Response of Primary Metabolites
2.2.7 Altered Expression of Metabolite Related Genes in curl3 Mutant
2.3 Discussion
2.4 Conclusion
3 Altered brassinolide sensitivity1 Transcriptionally Inhibits Chlorophyll Synthesis and Photosynthesis Capacity in Tomato
3.1 Materials and Methods
3.1.1 Plant Material and DNA Extraction
3.1.2 RNA-seq Library Preparation and Sequencing
3.1.3 Determination of Leaf Morphological Traits and Chlorophyll Accumulation
3.1.4 Chlorophyll Fluorescence Measurements
3.1.5 Photosynthetic Measurements
3.1.6 Gene Expression Quantification
3.1.7 Extraction and Quantification of Endogenous Phytohormones
3.1.8 Data Analysis
3.2 Results
3.2.1 abs1 Mutant Characterization
3.2.2 Altered Leaf Development in abs1 Mutant
3.2.3 BR-insensitive Mutant Modulates Endogenous Phytohormone Production
3.2.4 Reduced Chlorophyll Synthesis in abs1 Mutant
3.2.5 Chlorophyll Fluorescence Activity was Inhibited in abs1 Mutant
3.2.6 Photosynthetic Capacity was Inhibited in abs1 Mutant
3.2.7 Validation of DEGs
3.3 Discussion
3.4 Conclusion
4 Altered brassinolide sensitivity1 Impaired Floral Organ and Pollen Development
4.1 Materials and Methods
4.1.1 Plant Material and Growth Conditions
4.1.2 Quantitative Gene Expression Profiling
4.1.3 Extraction and Quantification of Phytohormones
4.1.4 Morphological Characterization and Anatomical Observations
4.1.5 Pollen Viability Assay
4.1.6 Seed Germination Assay
4.1.7 Statistical Analysis
4.2 Results
4.2.1 Temporal expression of Curl3 During Floral Development
4.2.2 abs1 Modulate BR and GA Related Genes and Endogenous Concentrations
4.2.3 abs1 Mutant Modulate Flower Development in Tomato
4.2.4 abs1 Mutant Modulate Flower Anatomical Structure in Tomato
4.2.5 BR Signaling Disruption Reduces Male Fertility
4.2.6 BR Signaling Disruption Reduces Seed Formation
4.3 Discussion
4.4 Conclusion
5 Altered brassinolide sensitivity1 Alters Fruit Size in Association with Phytohormones Modulation in Tomato
5.1 Materials and Methods
5.1.1 Plant material and Gene Isolation
5.1.2 Quantitative Gene Expression Profiling
5.1.3 Morphological Characterization and Anatomical Observations
5.1.4 Extraction and Quantification of Phytohormones
5.1.5 Statistical Analysis
5.2 Results
5.2.1 Temporal expression of Curl3 During Fruit Development
5.2.2 The Altered Transcript Level of BR Signaling Components in abs1 Mutant Fruit
5.2.3 BR Signaling Disruption Causes a Decrease in Phytohormone Production
5.2.4 BR Signaling Disruption Decreased Fruit Size of abs1 Mutant Fruit
5.2.5 BR Signaling Disruption Influences Cell Division and Expansion During Early Developmental Stages
5.2.6 The Altered Transcript Level of Fruit Size-Determining Genes in abs1 Mutant Fruit
5.3 Discussion
5.4 Conclusion
6 Summary and Future Perspectives
References
Appendices
Acknowledgements
本文編號(hào)：3986332
【文章頁(yè)數(shù)】：110 頁(yè)
【學(xué)位級(jí)別】：博士
【文章目錄】：
Abstract
摘要
List of Abbreviations
1 Introduction
1.1 Plant Hormones
1.2 Brassinolide Hormone
1.3 Brassinolide Crosstalk with other Hormones
1.4 Brassinolide Perception by BRI1
1.5 BR-Dependent Gene Expression
1.6 BR-Signaling Mutant
1.7 Objectives of This Research
2 Brassinolide Signaling Disruption Regulates Primary Metabolism in Accordance with Phytohormone Production
2.1 Materials and Methods
2.1.1 Plant Material
2.1.2 Gene Isolation
2.1.3 CRISPR/Cas9 Gene Editing
2.1.4 Morphological Characterization and Anatomical Observations
2.1.5 Hormone Quantification
2.1.6 Tomato Primary Metabolite Profiling
2.1.7 Quantitative Gene Expression Profiling
2.1.8 Statistical and Bioinformatics Analysis
2.2 Results
2.2.1 Sequence analysis of Curl3 from Solanum lycopersicum
2.2.2 CRISPR based Gene Editing
2.2.3 Altered Expression of BR Signaling Components
2.2.4 Morphological Characterization and Anatomical Observations of curl3 Mutant
2.2.5 BR-Insensitivity Alters Phytohormone Production in Transgenic Tomato Plants
2.2.6 Response of Primary Metabolites
2.2.7 Altered Expression of Metabolite Related Genes in curl3 Mutant
2.3 Discussion
2.4 Conclusion
3 Altered brassinolide sensitivity1 Transcriptionally Inhibits Chlorophyll Synthesis and Photosynthesis Capacity in Tomato
3.1 Materials and Methods
3.1.1 Plant Material and DNA Extraction
3.1.2 RNA-seq Library Preparation and Sequencing
3.1.3 Determination of Leaf Morphological Traits and Chlorophyll Accumulation
3.1.4 Chlorophyll Fluorescence Measurements
3.1.5 Photosynthetic Measurements
3.1.6 Gene Expression Quantification
3.1.7 Extraction and Quantification of Endogenous Phytohormones
3.1.8 Data Analysis
3.2 Results
3.2.1 abs1 Mutant Characterization
3.2.2 Altered Leaf Development in abs1 Mutant
3.2.3 BR-insensitive Mutant Modulates Endogenous Phytohormone Production
3.2.4 Reduced Chlorophyll Synthesis in abs1 Mutant
3.2.5 Chlorophyll Fluorescence Activity was Inhibited in abs1 Mutant
3.2.6 Photosynthetic Capacity was Inhibited in abs1 Mutant
3.2.7 Validation of DEGs
3.3 Discussion
3.4 Conclusion
4 Altered brassinolide sensitivity1 Impaired Floral Organ and Pollen Development
4.1 Materials and Methods
4.1.1 Plant Material and Growth Conditions
4.1.2 Quantitative Gene Expression Profiling
4.1.3 Extraction and Quantification of Phytohormones
4.1.4 Morphological Characterization and Anatomical Observations
4.1.5 Pollen Viability Assay
4.1.6 Seed Germination Assay
4.1.7 Statistical Analysis
4.2 Results
4.2.1 Temporal expression of Curl3 During Floral Development
4.2.2 abs1 Modulate BR and GA Related Genes and Endogenous Concentrations
4.2.3 abs1 Mutant Modulate Flower Development in Tomato
4.2.4 abs1 Mutant Modulate Flower Anatomical Structure in Tomato
4.2.5 BR Signaling Disruption Reduces Male Fertility
4.2.6 BR Signaling Disruption Reduces Seed Formation
4.3 Discussion
4.4 Conclusion
5 Altered brassinolide sensitivity1 Alters Fruit Size in Association with Phytohormones Modulation in Tomato
5.1 Materials and Methods
5.1.1 Plant material and Gene Isolation
5.1.2 Quantitative Gene Expression Profiling
5.1.3 Morphological Characterization and Anatomical Observations
5.1.4 Extraction and Quantification of Phytohormones
5.1.5 Statistical Analysis
5.2 Results
5.2.1 Temporal expression of Curl3 During Fruit Development
5.2.2 The Altered Transcript Level of BR Signaling Components in abs1 Mutant Fruit
5.2.3 BR Signaling Disruption Causes a Decrease in Phytohormone Production
5.2.4 BR Signaling Disruption Decreased Fruit Size of abs1 Mutant Fruit
5.2.5 BR Signaling Disruption Influences Cell Division and Expansion During Early Developmental Stages
5.2.6 The Altered Transcript Level of Fruit Size-Determining Genes in abs1 Mutant Fruit
5.3 Discussion
5.4 Conclusion
6 Summary and Future Perspectives
References
Appendices
Acknowledgements
本文編號(hào)：3986332
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