小地老虎性信息素合成激活肽受体（PBANR）的克
发布时间：2021-05-20 09:24
小地老虎Agrotis ipsilon是一种破坏力很强的迁飞性鳞翅目害虫,危害世界范围内的多种作物。性信息素合成激活肽受体（PBANR）,调节多种鳞翅目昆虫性信息素的合成。能够与调控性信息素合成中扮演重要角色的GPCRs结合。在鳞翅目成虫中,交配能够引起雌虫短时间终止性信息素的产生。本文通过RACE-PCR方法,从小地老虎雌虫性腺中克隆获得小地老虎PBANR全长基因,命名为Aips-PBANR,该基因由3539个碱基组成。蛋白序列比对表明,Aips-PBANR与鳞翅目其他昆虫的PBANR具有较高的序列一致性。利用RT-PCR方法,从小地老虎雌虫性腺和雄虫生殖器中均检测到Aips-PBANR基因的表达。qRT-PCR该基因在成虫不同组织和日龄的表达特征进行研究。结果表明,Aips-PBANR在雌虫性腺中的表达量最高,在雄虫的生殖器中也检测到较低的表达;羽化后第四天上午2:00基因表达量最高;另外,qRT-PCR结果表明,交配能够明显抑制Aips-PBANR基因的表达,因此进一步证明交配能够抑制昆虫性信息素的产生。RNA干扰技术（RNAi）作为害虫防治的新方法具有很大的应用潜力。然而,在不...
【文章来源】：中国农业科学院北京市
【文章页数】：138 页
【学位级别】：博士
【文章目录】：
中文摘要
Abstract
Abbreviations
CHAPTER 1 Introduction and review of literature
1.1. Black Cutworm
1.2. Distribution of Black cutworm
1.3. Life History and Developmental Stages of BCW
1.3.1. Life History of BCW
1.3.2. Egg Stage
1.3.3. Larval Stage
1.3.4. Pupal Stage
1.3.5. Adult Stage
1.4. Factors and Damages by BCW
1.5. Significance and Migration of BCW
1.6. Economic Importance
1.7. Management of BCW
1.7.1. Survey and Trapping
1.7.2. Chemical Control
1.7.3. Cultural and Mechanical Control
1.8. Molecular Techniques used the present study
1.8.1. Rapid Amplification of cDNA Ends (RACE)
1.8.2. Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR)
1.8.3. Quantitative Real-Time PCR (qRT-PCR)
1.8.4. Basic Principals of Real Time PCR
1.8.5. Chemistries of Real Time PCR
1.8.6. DNA Binding Dye Chemistry
1.8.7. Probe Based Chemistry
1.8.8. Primer and Probe designing for qRT-PCR
1.8.9. The qRT-PCR Reaction Performance
1.8.10. PCR Reaction Efficiency
1.8.11. PCR Reaction Precision
1.8.12. PCR Reaction Sensitivity
1.9. Objectives
CHAPTER 2 Molecular Cloning, differential expression and mating interruption of Pheromone Biosynthesis Activating Neuropeptide Receptor, a sex pheromone gene in blackcutworm, Agrotis ipsilon (Hufnagel) (Lepidoptera:Noctuidae) adults
2.1. Introduction
2.2. Materials and methods
2.2.1. Ethics Statement
2.2.2. Insect Material and Rearing Method
2.2.3. Pheromone Gland Dissection Method
2.2.4. Total RNA Extraction
2.2.5. Synthesis of cDNA
2.2.6. Tissue distribution, different time rythms and mating effect studies
2.2.7. Molecular Cloning and sequencing of A. ipsilon PBANR
2.2.8. Full length PBANR acquired by Rapid amplification of cDNA ends(RACE)
2.2.9. Basic Bioinformatic Analysis
2.2.10. Aips-PBANR Expression by Reverse transcription PCR (RT-PCR)
2.2.11. Expression of Aips-PBANR by Quantitative Real-Time PCR
2.2.12. Statistical Analysis
2.3. Results
2.3.1. Cloning and Sequencing of Aips-PBANR
2.3.2. Phylogenetic Relationship of Aips-PBANR
2.3.3. RT-PCR analysis of Aips-PBANR expression
2.3.4. Quantitative RT-PCR (qRT-PCR) Analysis of Aips-PBANR
2.4. DISCUSSION
CHAPTER 3 Knockdown of pheromone biosynthesis-activating neuropeptide receptor (PBANR) in black cutworm, Agrotis ipsilon (Hufnagel) (Lepidoptera:Noctuidae) adults by RNAinterference
3.1. Introduction
3.1.1. Sex Pheromone
3.1.2. RNA interference (RNAi)
3.1.3. Need for RNAi
3.1.4. RNA interference by siRNA
3.1.5. RNA interference by dsRNA
3.1.6. Different dsRNA Uptake methods used in RNAi
3.1.7. Advantages and disadvantages of RNAi by dsRNA
3.1.8. Important factors influencing RNAi through dsRNA
3.1.8.1 Concentration of dsRNA
3.1.8.2 Nucleotide sequence
3.1.8.3 Length of dsRNA fragment
3.1.8.4 Persistence of the silencing effect
3.1.8.5 Life stage of the target organism
3.2. Materials and methods
3.2.1. Insect Culture and Rearing Methods
3.2.2. Tissue Collection
3.2.3. RNA extraction and cDNA synthesis
3.2.4. Molecular cloning
3.2.5. Synthesis and Injection of siRNA and dsRNA
3.2.6. Quantitative Real-Time PCR (qRT-PCR) analysis
3.2.7. Data Analysis
3.3. Results
3.3.1. Cloning and Sequencing of Aips-PBANR
3.3.2. Quantitative RT-PCR (q RT-PCR) Analysis of Aips-PBANR expression
3.3.3. Effect of si RNA treatment on Aips-PBANR transcript level
3.3.4. Effect of dsRNA treatment on Aips-PBANR transcript level
3.4. Discussion
CHAPTER 4 The influence of different temperatures on mating traits, oviposition andlogivity of black cutworm, Agrotis ipsilon (Hufnagel) (Lepidoptera:Noctuidae)
4.1. Introduction
4.2. Materials and methods
4.2.1. Study Organism
4.2.2. Insect Colony Rearing
4.2.3. Temperature and Mating Conditions
4.2.4. Mating Observations
4.2.5. Oviposition, Fecundity and Adult Longivity
4.2.6. Statistical Data analysis
4.3. Results
4.3.1. Mating Activity
4.3.2. Oviposition Traits
4.3.3. Adult Longivity
4.4. Discussion
CHAPTER 5 Conclusions and Recommendations
References
Acknowledgements
Author biography
【参考文献】：
期刊论文
[1]我国2008年草地螟大发生种群空间分布特征[J]. 黄绍哲,罗礼智,姜玉英,唐继洪,张蕾. 植物保护. 2011(04)
[2]草地螟交配行为及能力[J]. 王凯,程云霞,江幸福,罗礼智. 应用昆虫学报. 2011(04)
[3]我国2008年草地螟大发生特征及成因分析[J]. 罗礼智,黄绍哲,江幸福,张蕾. 植物保护. 2009(01)
[4]甜菜夜蛾性信息素激活肽基因在雌成虫的不同发育时期的表达[J]. 许国庆,丛斌,罗礼智. 昆虫知识. 2006(01)
[5]温度对甜菜夜蛾生殖行为及生殖力的影响[J]. 王竑晟,徐洪富,崔峰,许永玉,周真. 生态学报. 2004(01)
[6]交配和温度对甜菜夜蛾（Spodoptera exigua）雌蛾性信息素产生的影响[J]. 董双林,杜家纬. 应用生态学报. 2002(12)
[7]饲养温度对粘虫飞行和生殖能力的影响[J]. 江幸福,罗礼智,胡毅. 生态学报. 2000(02)
[8]温度对草地螟成虫产卵和寿命的影响[J]. 罗礼智,李光博. 昆虫学报. 1993(04)
[9]小地老虎对几类杀虫剂的毒力反应及其抗药性变化[J]. 韩召军. 植物保护学报. 1986(02)
本文编号：3197517
【文章来源】：中国农业科学院北京市
【文章页数】：138 页
【学位级别】：博士
【文章目录】：
中文摘要
Abstract
Abbreviations
CHAPTER 1 Introduction and review of literature
1.1. Black Cutworm
1.2. Distribution of Black cutworm
1.3. Life History and Developmental Stages of BCW
1.3.1. Life History of BCW
1.3.2. Egg Stage
1.3.3. Larval Stage
1.3.4. Pupal Stage
1.3.5. Adult Stage
1.4. Factors and Damages by BCW
1.5. Significance and Migration of BCW
1.6. Economic Importance
1.7. Management of BCW
1.7.1. Survey and Trapping
1.7.2. Chemical Control
1.7.3. Cultural and Mechanical Control
1.8. Molecular Techniques used the present study
1.8.1. Rapid Amplification of cDNA Ends (RACE)
1.8.2. Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR)
1.8.3. Quantitative Real-Time PCR (qRT-PCR)
1.8.4. Basic Principals of Real Time PCR
1.8.5. Chemistries of Real Time PCR
1.8.6. DNA Binding Dye Chemistry
1.8.7. Probe Based Chemistry
1.8.8. Primer and Probe designing for qRT-PCR
1.8.9. The qRT-PCR Reaction Performance
1.8.10. PCR Reaction Efficiency
1.8.11. PCR Reaction Precision
1.8.12. PCR Reaction Sensitivity
1.9. Objectives
CHAPTER 2 Molecular Cloning, differential expression and mating interruption of Pheromone Biosynthesis Activating Neuropeptide Receptor, a sex pheromone gene in blackcutworm, Agrotis ipsilon (Hufnagel) (Lepidoptera:Noctuidae) adults
2.1. Introduction
2.2. Materials and methods
2.2.1. Ethics Statement
2.2.2. Insect Material and Rearing Method
2.2.3. Pheromone Gland Dissection Method
2.2.4. Total RNA Extraction
2.2.5. Synthesis of cDNA
2.2.6. Tissue distribution, different time rythms and mating effect studies
2.2.7. Molecular Cloning and sequencing of A. ipsilon PBANR
2.2.8. Full length PBANR acquired by Rapid amplification of cDNA ends(RACE)
2.2.9. Basic Bioinformatic Analysis
2.2.10. Aips-PBANR Expression by Reverse transcription PCR (RT-PCR)
2.2.11. Expression of Aips-PBANR by Quantitative Real-Time PCR
2.2.12. Statistical Analysis
2.3. Results
2.3.1. Cloning and Sequencing of Aips-PBANR
2.3.2. Phylogenetic Relationship of Aips-PBANR
2.3.3. RT-PCR analysis of Aips-PBANR expression
2.3.4. Quantitative RT-PCR (qRT-PCR) Analysis of Aips-PBANR
2.4. DISCUSSION
CHAPTER 3 Knockdown of pheromone biosynthesis-activating neuropeptide receptor (PBANR) in black cutworm, Agrotis ipsilon (Hufnagel) (Lepidoptera:Noctuidae) adults by RNAinterference
3.1. Introduction
3.1.1. Sex Pheromone
3.1.2. RNA interference (RNAi)
3.1.3. Need for RNAi
3.1.4. RNA interference by siRNA
3.1.5. RNA interference by dsRNA
3.1.6. Different dsRNA Uptake methods used in RNAi
3.1.7. Advantages and disadvantages of RNAi by dsRNA
3.1.8. Important factors influencing RNAi through dsRNA
3.1.8.1 Concentration of dsRNA
3.1.8.2 Nucleotide sequence
3.1.8.3 Length of dsRNA fragment
3.1.8.4 Persistence of the silencing effect
3.1.8.5 Life stage of the target organism
3.2. Materials and methods
3.2.1. Insect Culture and Rearing Methods
3.2.2. Tissue Collection
3.2.3. RNA extraction and cDNA synthesis
3.2.4. Molecular cloning
3.2.5. Synthesis and Injection of siRNA and dsRNA
3.2.6. Quantitative Real-Time PCR (qRT-PCR) analysis
3.2.7. Data Analysis
3.3. Results
3.3.1. Cloning and Sequencing of Aips-PBANR
3.3.2. Quantitative RT-PCR (q RT-PCR) Analysis of Aips-PBANR expression
3.3.3. Effect of si RNA treatment on Aips-PBANR transcript level
3.3.4. Effect of dsRNA treatment on Aips-PBANR transcript level
3.4. Discussion
CHAPTER 4 The influence of different temperatures on mating traits, oviposition andlogivity of black cutworm, Agrotis ipsilon (Hufnagel) (Lepidoptera:Noctuidae)
4.1. Introduction
4.2. Materials and methods
4.2.1. Study Organism
4.2.2. Insect Colony Rearing
4.2.3. Temperature and Mating Conditions
4.2.4. Mating Observations
4.2.5. Oviposition, Fecundity and Adult Longivity
4.2.6. Statistical Data analysis
4.3. Results
4.3.1. Mating Activity
4.3.2. Oviposition Traits
4.3.3. Adult Longivity
4.4. Discussion
CHAPTER 5 Conclusions and Recommendations
References
Acknowledgements
Author biography
【参考文献】：
期刊论文
[1]我国2008年草地螟大发生种群空间分布特征[J]. 黄绍哲,罗礼智,姜玉英,唐继洪,张蕾. 植物保护. 2011(04)
[2]草地螟交配行为及能力[J]. 王凯,程云霞,江幸福,罗礼智. 应用昆虫学报. 2011(04)
[3]我国2008年草地螟大发生特征及成因分析[J]. 罗礼智,黄绍哲,江幸福,张蕾. 植物保护. 2009(01)
[4]甜菜夜蛾性信息素激活肽基因在雌成虫的不同发育时期的表达[J]. 许国庆,丛斌,罗礼智. 昆虫知识. 2006(01)
[5]温度对甜菜夜蛾生殖行为及生殖力的影响[J]. 王竑晟,徐洪富,崔峰,许永玉,周真. 生态学报. 2004(01)
[6]交配和温度对甜菜夜蛾（Spodoptera exigua）雌蛾性信息素产生的影响[J]. 董双林,杜家纬. 应用生态学报. 2002(12)
[7]饲养温度对粘虫飞行和生殖能力的影响[J]. 江幸福,罗礼智,胡毅. 生态学报. 2000(02)
[8]温度对草地螟成虫产卵和寿命的影响[J]. 罗礼智,李光博. 昆虫学报. 1993(04)
[9]小地老虎对几类杀虫剂的毒力反应及其抗药性变化[J]. 韩召军. 植物保护学报. 1986(02)
本文编号：3197517
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