夜间高温和白天高温导致棉花雄性不育的比较研究
发布时间：2021-03-06 01:17
棉花是重要的经济作物,随着全球气温的逐年升高,棉花的产量受到高温胁迫的威胁越来越严重。高温胁迫对植物生殖器官的影响要远大于营养器官,且雄性生殖器官对温度的敏感程度高于雌性生殖器官,所以高温胁迫下的雄性不育更应受到关注。因不同的高温类型会引起不同的雄性不育表型,为了分析不同高温类型导致雄性败育发生的机理,我们设置了四种温度组合条件（白天常温/晚间常温,NN;白天高温/晚间常温,HN;白天常温/晚间高温,NH;白天高温/晚间高温,HH）进行处理,并对处理后的棉花三个发育时期（四分体时期、绒毡层降解时期、花药开裂期）的花药进行组织细胞水平,生理生化水平及转录水平分析。研究发现,高夜温对于棉花雄性育性的影响要远远大于高日温,其造成的雄性不育是引起棉花减产的主要原因之一。具体研究结果如下:1)高温会导致花器官明显变小,花丝变短,花粉不育,其中全天高温（HH）造成的不育最明显,且高夜温（NH）引起的花粉不育比高日温（HN）要明显。通过花药切片观察发现,在绒毡层降解时期,全天高温条件下的小孢子出现明显皱缩,而在白天高温/晚间正常和白天正常/晚间高温条件下有部分小孢子出现皱缩,程度要轻于全天高温。2)...
【文章来源】：华中农业大学湖北省 211工程院校 教育部直属院校
【文章页数】：136 页
【学位级别】：博士
【文章目录】：
摘要
ABSTRACT
ABBREVIATIONS
CHAPTER I
1 INTRODUCTION
1.1 High Temperature（HT）stress
1.2 High temperature stress and its effect on crops
1.3 High temperature stress and its effect on Cotton
1.4 High temperature and flowering time
1.5 High temperature and fertility
1.6 High temperature affects pollen development
1.7 High temperature and its effect on carbohydrate
1.8 High temperature and its effects on lipids
1.9 High temperature and its effect on ROS in the anthers
1.10 High temperature and hormones
1.10.1 Auxin and male sterility
1.10.2 Gibberellin（GA）and male sterility
1.10.3 Jasmonic Acid（JA）and male sterility
1.11 CRISPR/Cas9 system in plants
1.11.1 Mechanism of CRISPR-Cas9 system
1.11.2 Applications of CRISPR/Cas9 in Crop improvement
1.12 Aims and overview of the present study
CHAPTER II
2.MATERIALS AND METHODS
2.1 Plant material
2.2 Pollen viability,Tissue sectioning and observation
2.3 RNA sequencing
2.4 Data analysis
2.5 Quantitative Reverse Transcriptase PCR（q RT-PCR）
2.6 Fatty acid extraction and measurement
2.7 Transmission electron microscopy（TEM）
2.8 Measurement of ABA,IAA,and JA
2.9 Quantification of H2O2
2.10 Starch,sucrose and fructose measurement
2.11 Vector construction and transformation in cotton
2.11.1 PCR conditions:
2.11.2 Second PCR and purification
2.11.3 PCR conditions:
2.11.4 Enzyme digestion of the expression vector:
2.11.5 In-fusion ligation
2.11.6 Heat shock and electroporation
2.12 Plant material,growth conditions,and treatments
2.13 Tissue culturing and generation of homozygous transgenic lines
2.14 DNA extraction and Southern blotting
2.15 Hi-TOM sequencing
2.16 Application of exogenous Me JA to buds
2.17 Reactive oxygen species（ROS）staining
2.18 Statistical analysis
CHAPTER III
3.RESULTS
3.1 Phenotypic studies of the high temperature（HT）sensitive line H05 under day and night HT stress
3.2 High night temperature causes pollen abortion
3.3 Differentially expressed genes（DEGs）in H05 anthers under different HT stress combinations
3.4 HT stress disrupts cotton anthers lipid metabolism pathways
3.5 Day and night HT affect fatty acid metabolism differently
3.6 Day and night HT drive Alpha-linolenic acid metabolism to JA in ADS stage
3.7 Effects of HT on the glycerolipid metabolism pathway genes
3.8 Day and night HT affects the lipid and sugar contents in anthers
3.9 Study of selected genes from the Jasmonic Acid（JA）pathway
3.9.1 Structural characterization of Allene oxide cyclase4（AOC4）and Acyl-Co A oxidase2（ACO2）
3.9.2 Transformation and plant regeneration
3.9.3 Selection of Gh AOC4 and Gh ACO2 mutant lines
3.9.4 Phenotypic study and histological sectioning of Gh AOC4 and Gh ACO2 mutant lines anthers
3.9.5 Transmission electron microscopy（TEM）of the mutant lines anthers
3.9.6 Gh AOC4 and Gh ACO2 gene disrupts the JA pathway and caused pollen abortion
3.9.7 High temperature（HT）decreases JA accumulation in the anthers of cotton HT-sensitive line anthers and causes male sterility
3.9.8 Decreased JA biosynthesis causes the accumulation of H2O2 in the anther and pollen of the Gh AOC4 mutant
CHAPTER IV
4.DISCUSSION
4.1 Lipid and fatty acid changes are associated with male sterility under both day and night HT stress
4.2 Day and Night HT disrupt the glycerolipid and JA pathway genes,leading to male sterility
4.3 Disruption of hormones during day and night HT leads to male sterility
4.4 Knock out of Gh AOC4 and Gh ACO2 leads to male sterility
4.5 Gh ACO2 and Gh AOC4 are responsible for the sterile pollen and anther indehiscence
CONCLUSION
REFERENCES
APPENDICES
APPENDIX 1
APPENDIX 2
APPENDIX 3
APPENDIX 4
APPENDIX 5
APPENDIX 6
APPENDIX 7
APPENDIX 8
ACKNOWLEDGMENT
List of Publications
本文编号：3066187
【文章来源】：华中农业大学湖北省 211工程院校 教育部直属院校
【文章页数】：136 页
【学位级别】：博士
【文章目录】：
摘要
ABSTRACT
ABBREVIATIONS
CHAPTER I
1 INTRODUCTION
1.1 High Temperature（HT）stress
1.2 High temperature stress and its effect on crops
1.3 High temperature stress and its effect on Cotton
1.4 High temperature and flowering time
1.5 High temperature and fertility
1.6 High temperature affects pollen development
1.7 High temperature and its effect on carbohydrate
1.8 High temperature and its effects on lipids
1.9 High temperature and its effect on ROS in the anthers
1.10 High temperature and hormones
1.10.1 Auxin and male sterility
1.10.2 Gibberellin（GA）and male sterility
1.10.3 Jasmonic Acid（JA）and male sterility
1.11 CRISPR/Cas9 system in plants
1.11.1 Mechanism of CRISPR-Cas9 system
1.11.2 Applications of CRISPR/Cas9 in Crop improvement
1.12 Aims and overview of the present study
CHAPTER II
2.MATERIALS AND METHODS
2.1 Plant material
2.2 Pollen viability,Tissue sectioning and observation
2.3 RNA sequencing
2.4 Data analysis
2.5 Quantitative Reverse Transcriptase PCR（q RT-PCR）
2.6 Fatty acid extraction and measurement
2.7 Transmission electron microscopy（TEM）
2.8 Measurement of ABA,IAA,and JA
2.9 Quantification of H2O2
2.10 Starch,sucrose and fructose measurement
2.11 Vector construction and transformation in cotton
2.11.1 PCR conditions:
2.11.2 Second PCR and purification
2.11.3 PCR conditions:
2.11.4 Enzyme digestion of the expression vector:
2.11.5 In-fusion ligation
2.11.6 Heat shock and electroporation
2.12 Plant material,growth conditions,and treatments
2.13 Tissue culturing and generation of homozygous transgenic lines
2.14 DNA extraction and Southern blotting
2.15 Hi-TOM sequencing
2.16 Application of exogenous Me JA to buds
2.17 Reactive oxygen species（ROS）staining
2.18 Statistical analysis
CHAPTER III
3.RESULTS
3.1 Phenotypic studies of the high temperature（HT）sensitive line H05 under day and night HT stress
3.2 High night temperature causes pollen abortion
3.3 Differentially expressed genes（DEGs）in H05 anthers under different HT stress combinations
3.4 HT stress disrupts cotton anthers lipid metabolism pathways
3.5 Day and night HT affect fatty acid metabolism differently
3.6 Day and night HT drive Alpha-linolenic acid metabolism to JA in ADS stage
3.7 Effects of HT on the glycerolipid metabolism pathway genes
3.8 Day and night HT affects the lipid and sugar contents in anthers
3.9 Study of selected genes from the Jasmonic Acid（JA）pathway
3.9.1 Structural characterization of Allene oxide cyclase4（AOC4）and Acyl-Co A oxidase2（ACO2）
3.9.2 Transformation and plant regeneration
3.9.3 Selection of Gh AOC4 and Gh ACO2 mutant lines
3.9.4 Phenotypic study and histological sectioning of Gh AOC4 and Gh ACO2 mutant lines anthers
3.9.5 Transmission electron microscopy（TEM）of the mutant lines anthers
3.9.6 Gh AOC4 and Gh ACO2 gene disrupts the JA pathway and caused pollen abortion
3.9.7 High temperature（HT）decreases JA accumulation in the anthers of cotton HT-sensitive line anthers and causes male sterility
3.9.8 Decreased JA biosynthesis causes the accumulation of H2O2 in the anther and pollen of the Gh AOC4 mutant
CHAPTER IV
4.DISCUSSION
4.1 Lipid and fatty acid changes are associated with male sterility under both day and night HT stress
4.2 Day and Night HT disrupt the glycerolipid and JA pathway genes,leading to male sterility
4.3 Disruption of hormones during day and night HT leads to male sterility
4.4 Knock out of Gh AOC4 and Gh ACO2 leads to male sterility
4.5 Gh ACO2 and Gh AOC4 are responsible for the sterile pollen and anther indehiscence
CONCLUSION
REFERENCES
APPENDICES
APPENDIX 1
APPENDIX 2
APPENDIX 3
APPENDIX 4
APPENDIX 5
APPENDIX 6
APPENDIX 7
APPENDIX 8
ACKNOWLEDGMENT
List of Publications
本文编号：3066187
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