奶牛瘤胃尿素分解菌溶糊精琥珀酸弧菌Z6完整基因组与氮利用
发布时间：2021-11-17 06:47
溶糊精琥珀酸弧菌（Succinivibrio dextrinosolvens）属于革兰氏阴性厌氧细菌,是反刍动物瘤胃中的常见栖息细菌,并且在高谷物日粮条件下数量较多。溶糊精琥珀酸弧菌能发酵淀粉水解产物糊精和麦芽糖,某些溶糊精琥珀酸弧菌菌株能产生高活性脲酶。因此,研究瘤胃溶糊精琥珀酸弧菌的基因组特征及其代谢尿素氮机制,对于丰富瘤胃微生物氮代谢研究具有重要补充作用。通过厌氧细菌分离技术,从奶牛瘤胃中分离获得一株溶糊精琥珀酸弧菌,命名为SD-Z6菌株。通过三代Oxford高通量测序,分析溶糊精琥珀酸弧菌SD-Z6的全基因组序列,并且与该菌种中其他菌株基因组进行比较。同时,检测溶糊精琥珀酸弧菌SD-Z6的形态特征和生理生化。利用尿素、氨基酸和硫酸铵三种氮源,分析溶糊精琥珀酸弧菌SD-Z6的氮代谢,并利用转录组方法分析溶糊精琥珀酸弧菌SD-Z6功能基因的变化。结果表明,溶糊精琥珀酸弧菌SD-Z6的全基因组大小为3.47 Mbp,G+C含量为38.9%。总共含2993个编码序列,占98%,其中鉴定出21个rRNA基因和69个tRNA。COG功能注释表明,调节碳水化合物（10.6%）和氨基酸（9%）...
【文章来源】：中国农业科学院北京市
【文章页数】：116 页
【学位级别】：博士
【文章目录】：
摘要
Abstract
Abbreviations
Chapter1.Literature Review
1.1.Introduction
1.2.True protein as nitrogen sources for ruminants
1.2.1.Degradation of protein in the rumen
1.3.Non– protein nitrogen containing compound for ruminants
1.4.Urea as protein sources for ruminants
1.5.Metabolism of Urea/Non-protein nitrogen compound in ruminants
1.6.Ammonia assimilation and emission
1.6.1.Glutamate dehydrogenase
1.6.2.Glutamine synthetase
1.6.3.Glutamate synthase
1.7.Factors affecting urea utilization and ammonia assimilation
1.7.1.Effects of nitrogen Sources
1.7.2.Effects of nitrogen concentration
1.7.3.Effect of Energy on ammonia assimilation and NPN utilization
1.7.4.Effects of enzyme activity inhibitors on Nitrogen metabolism
1.8.Isolation,identification and characterization of ruminal bacteria
1.8.1.Culture-dependent methods
1.8.2.Culture-independent methods
1.8.3.MALDI-TOF mass spectrometry methods
1.9.Phenotypic characteristics and importance of Succinivibrio dextrinosolvens
Chapter2.Enrichment,Isolation and Characterization of a ureolytic Succinivibrio dextrinosolvens strain Z6 from rumen of dairy cows
2.1.Summary
2.2.Introduction
2.3.Material Methods
2.3.1.Media components and Preparation
2.3.2.Rumen fluid sample collection
2.3.3.Enrichment and isolation cultures
2.3.4.DNA isolation and PCR Amplification
2.3.5.Matrix assisted laser desorption ionization time of flight mass spectrometry(MALDI-TOF MS)analysis
2.3.6.Phenotypic and Chemotaxonomic Characterization
2.4.Result
2.4.1.Isolation and enrichment
2.4.2.Phenotypic and chemotaxonomic characterization
2.5.Discussion
2.6.Conclusion
Chapter3.Complete genome sequence and identification of gene related to nitrogen metabolism of Succinivibrio dextrinosolvens strain Z6
3.1.Summary
3.2.Introduction
3.3.Material and methods
3.3.1.Bacterial Culture Conditions
3.3.2.Genomic DNA Extraction
3.3.3.Whole genome sequencing and analysis
3.4.Results
3.4.1.Complete genome of S.dextrinosolvens Z6
3.4.2.Comparative Genome Analysis of S.dextrinosolvens strains
3.4.3.Genes involved in nitrogen metabolism in S.dextrinosolvens Z6
3.5.Discussion
3.6.Conclusion
Chapter4.Effects of different nitrogen sources on the transcriptional regulation of nitrogen metabolism and the corresponding expression of genes to enzymatic activities in Succinivibrio dextrinosolvens strain Z6
4.1.Summary
4.2.Introduction
4.3.Material and methods
4.3.1.Bacteria and culture conditions
4.3.2.Experimental design
4.3.3.Nitrogen Metabolism
4.3.4.Sampling technique
4.3.5.Chemical and Enzyme activity analysis
4.3.6.Transcriptomic analysis
4.3.7.KEGG pathway enrichment
4.3.8.Validation of DEGs using quantitative real time PCR
4.3.9.Statistical analysis
4.3.10.Data availability
4.4.Results
4.4.1.Nitrogen Metabolism of S.dextrinosolvens Z6 in different nitrogen sources
4.4.2.Different nitrogen sources utilization and Ammonia concentration
4.4.3.Ammonia assimilation of S.dextrinosolvens Z6 in different nitrogen sources
4.4.4.RNA sequencing and read mapping
4.4.5.Quantitative Real Time PCR validation
4.4.6.Transcriptomic analysis of genes in response to different nitrogen sources
4.4.7.KEGG pathway enrichment analysis of differentially expressed gene
4.5.Discussion
4.6.Conclusion
Chapter5.General conclusion
References
Appendix
Acknowledgements
Author Resume
本文编号：3500411
【文章来源】：中国农业科学院北京市
【文章页数】：116 页
【学位级别】：博士
【文章目录】：
摘要
Abstract
Abbreviations
Chapter1.Literature Review
1.1.Introduction
1.2.True protein as nitrogen sources for ruminants
1.2.1.Degradation of protein in the rumen
1.3.Non– protein nitrogen containing compound for ruminants
1.4.Urea as protein sources for ruminants
1.5.Metabolism of Urea/Non-protein nitrogen compound in ruminants
1.6.Ammonia assimilation and emission
1.6.1.Glutamate dehydrogenase
1.6.2.Glutamine synthetase
1.6.3.Glutamate synthase
1.7.Factors affecting urea utilization and ammonia assimilation
1.7.1.Effects of nitrogen Sources
1.7.2.Effects of nitrogen concentration
1.7.3.Effect of Energy on ammonia assimilation and NPN utilization
1.7.4.Effects of enzyme activity inhibitors on Nitrogen metabolism
1.8.Isolation,identification and characterization of ruminal bacteria
1.8.1.Culture-dependent methods
1.8.2.Culture-independent methods
1.8.3.MALDI-TOF mass spectrometry methods
1.9.Phenotypic characteristics and importance of Succinivibrio dextrinosolvens
Chapter2.Enrichment,Isolation and Characterization of a ureolytic Succinivibrio dextrinosolvens strain Z6 from rumen of dairy cows
2.1.Summary
2.2.Introduction
2.3.Material Methods
2.3.1.Media components and Preparation
2.3.2.Rumen fluid sample collection
2.3.3.Enrichment and isolation cultures
2.3.4.DNA isolation and PCR Amplification
2.3.5.Matrix assisted laser desorption ionization time of flight mass spectrometry(MALDI-TOF MS)analysis
2.3.6.Phenotypic and Chemotaxonomic Characterization
2.4.Result
2.4.1.Isolation and enrichment
2.4.2.Phenotypic and chemotaxonomic characterization
2.5.Discussion
2.6.Conclusion
Chapter3.Complete genome sequence and identification of gene related to nitrogen metabolism of Succinivibrio dextrinosolvens strain Z6
3.1.Summary
3.2.Introduction
3.3.Material and methods
3.3.1.Bacterial Culture Conditions
3.3.2.Genomic DNA Extraction
3.3.3.Whole genome sequencing and analysis
3.4.Results
3.4.1.Complete genome of S.dextrinosolvens Z6
3.4.2.Comparative Genome Analysis of S.dextrinosolvens strains
3.4.3.Genes involved in nitrogen metabolism in S.dextrinosolvens Z6
3.5.Discussion
3.6.Conclusion
Chapter4.Effects of different nitrogen sources on the transcriptional regulation of nitrogen metabolism and the corresponding expression of genes to enzymatic activities in Succinivibrio dextrinosolvens strain Z6
4.1.Summary
4.2.Introduction
4.3.Material and methods
4.3.1.Bacteria and culture conditions
4.3.2.Experimental design
4.3.3.Nitrogen Metabolism
4.3.4.Sampling technique
4.3.5.Chemical and Enzyme activity analysis
4.3.6.Transcriptomic analysis
4.3.7.KEGG pathway enrichment
4.3.8.Validation of DEGs using quantitative real time PCR
4.3.9.Statistical analysis
4.3.10.Data availability
4.4.Results
4.4.1.Nitrogen Metabolism of S.dextrinosolvens Z6 in different nitrogen sources
4.4.2.Different nitrogen sources utilization and Ammonia concentration
4.4.3.Ammonia assimilation of S.dextrinosolvens Z6 in different nitrogen sources
4.4.4.RNA sequencing and read mapping
4.4.5.Quantitative Real Time PCR validation
4.4.6.Transcriptomic analysis of genes in response to different nitrogen sources
4.4.7.KEGG pathway enrichment analysis of differentially expressed gene
4.5.Discussion
4.6.Conclusion
Chapter5.General conclusion
References
Appendix
Acknowledgements
Author Resume
本文编号：3500411
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