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Microbiota Analysis Using an Illumina MiSeq Platform to Sequence 16S rRNA Genes 利用Illumina MiSeq平台对16S rRNA基因进行微生物群分析
Current protocols in mouse biology Pub Date : 2017-06-19 DOI: 10.1002/cpmo.29
Alexis Rapin, Céline Pattaroni, Benjamin J. Marsland, Nicola L. Harris
{"title":"Microbiota Analysis Using an Illumina MiSeq Platform to Sequence 16S rRNA Genes","authors":"Alexis Rapin,&nbsp;Céline Pattaroni,&nbsp;Benjamin J. Marsland,&nbsp;Nicola L. Harris","doi":"10.1002/cpmo.29","DOIUrl":"10.1002/cpmo.29","url":null,"abstract":"<p>The microbiota have been shown to play an important role in diverse biological processes including immunity, metabolism, and digestion. Assessing the exact composition of the microbiota has proven challenging due to the often unknown growth specificities of its members, and culture-based approaches typically fail to capture the complete diversity of microorganisms present. Next Generation Sequencing (NGS) methods provide an efficient means to gather information about cultured and uncultured members of the microbiota. This article provides a method to characterize bacterial communities in terms of species composition using high-throughput sequencing. Briefly, by extracting the entire DNA content of a microbiota sample and performing a targeted high-throughput sequencing of the 16S rRNA gene, a phylogenetic marker for prokaryotes, prediction of the composition of the entire bacterial community is made possible. © 2017 by John Wiley &amp; Sons, Inc.</p>","PeriodicalId":37980,"journal":{"name":"Current protocols in mouse biology","volume":"7 2","pages":""},"PeriodicalIF":0.0,"publicationDate":"2017-06-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpmo.29","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"35100251","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 33
Non-Typeable Haemophilus influenzae Infection of the Junbo Mouse 君波小鼠不可分型流感嗜血杆菌感染
Current protocols in mouse biology Pub Date : 2017-03-02 DOI: 10.1002/cpmo.24
Michael T. Cheeseman, Derek W. Hood
{"title":"Non-Typeable Haemophilus influenzae Infection of the Junbo Mouse","authors":"Michael T. Cheeseman,&nbsp;Derek W. Hood","doi":"10.1002/cpmo.24","DOIUrl":"10.1002/cpmo.24","url":null,"abstract":"<p>Acute otitis media, inflammation of the middle ear bulla, is the most common bacterial infection in children. For one of the principal otopathogens, non-typeable <i>Haemophilus influenzae</i> (NTHi), animal models allow us to investigate host-microbial interactions relevant to the onset and progression of infection and to study treatment of middle ear disease. We have established a robust model of NTHi middle ear infection in the <i>Junbo</i> mouse. Intranasal inoculation with NTHi produces high rates of bulla infection and high bacterial titers in bulla fluids; bacteria can also spread down the respiratory tract to the mouse lung. An innate immune response is detected in the bulla of <i>Junbo</i> mice following NTHi infection, and bacteria are maintained in some ears at least up to day 56 post-inoculation. The <i>Junbo</i>/NTHi infection model facilitates studies on bacterial pathogenesis and antimicrobial intervention regimens and vaccines for better treatment and prevention of NTHi middle ear infection. © 2017 by John Wiley &amp; Sons, Inc.</p>","PeriodicalId":37980,"journal":{"name":"Current protocols in mouse biology","volume":"7 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2017-03-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpmo.24","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"34776796","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 2
Cre-loxP-Mediated Recombination: General Principles and Experimental Considerations cre - loxp介导的重组:一般原理和实验考虑
Current protocols in mouse biology Pub Date : 2017-03-02 DOI: 10.1002/cpmo.22
Micheal A. McLellan, Nadia A. Rosenthal, Alexander R. Pinto
{"title":"Cre-loxP-Mediated Recombination: General Principles and Experimental Considerations","authors":"Micheal A. McLellan,&nbsp;Nadia A. Rosenthal,&nbsp;Alexander R. Pinto","doi":"10.1002/cpmo.22","DOIUrl":"10.1002/cpmo.22","url":null,"abstract":"<p>The cre-<i>lox</i>P–mediated recombination system (the “cre-<i>lox</i>P system”) is an integral experimental tool for mammalian genetics and cell biology. Use of the system has greatly expanded our ability to precisely interrogate gene function in the mouse, providing both spatial and temporal control of gene expression. This has been largely due to the simplicity of its use and its adaptability to address diverse biological questions. While the use of the cre-<i>lox</i>P system is becoming increasingly widespread, in particular because of growing availability of conditional mouse mutants, many considerations need to be taken into account when utilizing the cre-<i>lox</i>P system. This review provides an overview of the cre-<i>lox</i>P system and its various permutations. It addresses the limitations of cre-<i>lox</i>P technology and related considerations for experimental design, and it discusses alternative strategies for site-specific genetic recombination and integration. © 2017 by John Wiley &amp; Sons, Inc.</p>","PeriodicalId":37980,"journal":{"name":"Current protocols in mouse biology","volume":"7 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2017-03-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpmo.22","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"34776793","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 110
Exploration of Inflammatory Bowel Disease in Mice: Chemically Induced Murine Models of Inflammatory Bowel Disease (IBD) 小鼠炎症性肠病的探索:化学诱导的炎症性肠病(IBD)小鼠模型
Current protocols in mouse biology Pub Date : 2017-03-02 DOI: 10.1002/cpmo.20
Raffaella Maria Gadaleta, Oihane Garcia-Irigoyen, Antonio Moschetta
{"title":"Exploration of Inflammatory Bowel Disease in Mice: Chemically Induced Murine Models of Inflammatory Bowel Disease (IBD)","authors":"Raffaella Maria Gadaleta,&nbsp;Oihane Garcia-Irigoyen,&nbsp;Antonio Moschetta","doi":"10.1002/cpmo.20","DOIUrl":"10.1002/cpmo.20","url":null,"abstract":"<p>Inflammatory bowel disease (IBD) is a chronic multifactorial inflammatory disorder characterized by periods of activation and remission of intestinal inflammation, with potentially severe complications, that can lead to mortality. Experimental animal models of intestinal inflammation are crucial for understanding the pathogenesis of Crohn's disease (CD) and ulcerative colitis (UC), the two major human IBD phenotypes. Animal models have been instrumental in unveiling the molecular background of IBD, and although a single model is not able to capture the complexity of this disease, each of them provided valuable insight into its different aspects. Chemically induced models of intestinal inflammation, mainly dextran sodium sulfate (DSS)- and 2,4,6-trinitrobenzenesulfonic acid (TNBS)-induced colitis, are widely used. This article describes DSS- and TNBS-induced colitis models and their relevance to IBD pathophysiology and pre-clinical therapeutic management. © 2017 by John Wiley &amp; Sons, Inc.</p>","PeriodicalId":37980,"journal":{"name":"Current protocols in mouse biology","volume":"7 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2017-03-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpmo.20","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"34776795","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 31
A Mouse Model for Ocular Surface Staphylococcus aureus Infection 小鼠眼表金黄色葡萄球菌感染模型的建立
Current protocols in mouse biology Pub Date : 2017-03-02 DOI: 10.1002/cpmo.23
Zhiyong Zhang, Osama Abdel-Razek, Guirong Wang
{"title":"A Mouse Model for Ocular Surface Staphylococcus aureus Infection","authors":"Zhiyong Zhang,&nbsp;Osama Abdel-Razek,&nbsp;Guirong Wang","doi":"10.1002/cpmo.23","DOIUrl":"10.1002/cpmo.23","url":null,"abstract":"<p>Creation of an appropriate animal model that accurately reflects the disease and host immune response to bacterial infection in humans is a major challenge in ocular-surface infection research. For decades, mice have been the ideal small animal model for ocular-surface infection research because of the availability and relatively low cost of various genetic backgrounds, targeted defects, and immunologic reagents. By employing different combinations of mouse and bacterial strains, murine infection models can be used to explore a complete picture of bacterial infection and innate immunity of the ocular surface. A murine model of <i>Staphylococcus aureus</i> infection under normal ocular circumstances is presented here as a convenient and tractable model system in which to study mammalian host responses to pathogens. © 2017 by John Wiley &amp; Sons, Inc.</p>","PeriodicalId":37980,"journal":{"name":"Current protocols in mouse biology","volume":"7 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2017-03-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpmo.23","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"34776797","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 3
Measurement of Behavioral Taste Responses in Mice: Two‐Bottle Preference, Lickometer, and Conditioned Taste‐Aversion Tests 小鼠味觉行为反应的测量:两瓶偏好、舔液计和条件味觉厌恶测试
Current protocols in mouse biology Pub Date : 2016-12-01 DOI: 10.1002/cpmo.18
D. Gaillard, J. Stratford
{"title":"Measurement of Behavioral Taste Responses in Mice: Two‐Bottle Preference, Lickometer, and Conditioned Taste‐Aversion Tests","authors":"D. Gaillard, J. Stratford","doi":"10.1002/cpmo.18","DOIUrl":"https://doi.org/10.1002/cpmo.18","url":null,"abstract":"The natural like and dislike of foods based on taste is one of the most easily observed behaviors in animals. Animals eat palatable foods and reject aversive foods, which makes measurement of taste perception possible using various behavioral techniques. Three different methods to accurately measure taste behavior are described here. First, two‐bottle preference tests evaluate whether a taste compound (tastant) is preferred over water. Second, lickometer tests quantify the like and dislike for multiple concentrations of the same tastant or multiple tastants at the same time. Finally, conditioned taste aversion tests accurately determine the perceived taste threshold for palatable tastants. Together, these diverse methods enable researchers to observe and measure behavioral taste responses in mice to any tastant. © 2016 by John Wiley & Sons, Inc.","PeriodicalId":37980,"journal":{"name":"Current protocols in mouse biology","volume":"6 1","pages":"380 - 407"},"PeriodicalIF":0.0,"publicationDate":"2016-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpmo.18","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"50901513","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 17
Methodological Considerations for Optimizing and Validating Behavioral Assays 优化和验证行为分析的方法学考虑
Current protocols in mouse biology Pub Date : 2016-12-01 DOI: 10.1002/cpmo.17
S. J. Sukoff Rizzo, J. Silverman
{"title":"Methodological Considerations for Optimizing and Validating Behavioral Assays","authors":"S. J. Sukoff Rizzo, J. Silverman","doi":"10.1002/cpmo.17","DOIUrl":"https://doi.org/10.1002/cpmo.17","url":null,"abstract":"Preclinical animal models are indispensable tools for translational research for which behavioral characterization and phenotyping are essential to testing hypotheses and for evaluating the potential of novel therapeutic agents to treat diseases. The methods employed for comprehensive behavioral phenotyping and pharmacological experiments are complex and should be conducted exclusively by trained technicians with demonstrated proficiency. The ultimate goal is to identify disease‐relevant and translational behavioral endpoints that are robust, reliable, and reproducible, and that can be employed to evaluate potential of novel therapeutic agents to treat disease. The intent of the present article is to provide a pragmatic outline for establishing and optimizing behavioral assays and phenotyping batteries, ensuring that the assays and the data are reliable such that they can be reproduced within and across technicians and laboratories and, more importantly, that the data is translatable to the clinic. © 2016 by John Wiley & Sons, Inc.","PeriodicalId":37980,"journal":{"name":"Current protocols in mouse biology","volume":"6 1","pages":"364 - 379"},"PeriodicalIF":0.0,"publicationDate":"2016-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpmo.17","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"50901449","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 35
Histopathological Evaluation of Skeletal Muscle with Specific Reference to Mouse Models of Muscular Dystrophy 肌肉萎缩症小鼠模型骨骼肌的组织病理学评价
Current protocols in mouse biology Pub Date : 2016-12-01 DOI: 10.1002/cpmo.19
R. Terry, D. Wells
{"title":"Histopathological Evaluation of Skeletal Muscle with Specific Reference to Mouse Models of Muscular Dystrophy","authors":"R. Terry, D. Wells","doi":"10.1002/cpmo.19","DOIUrl":"https://doi.org/10.1002/cpmo.19","url":null,"abstract":"The muscular dystrophies are a diverse group of degenerative diseases for which many mouse models are available. These models are frequently used to assess potential therapeutic interventions and histological evaluation of multiple muscles is an important part of this assessment. Histological evaluation is especially useful when combined with tests of muscle function. This unit describes a protocol for necropsy, processing, cryosectioning, and histopathological evaluation of murine skeletal muscles, which is applicable to both models of muscular dystrophy and other neuromuscular conditions. Key histopathological features of dystrophic muscle are discussed using the mdx mouse (a model of Duchenne muscular dystrophy) as an example. Optimal handling during dissection, processing and sectioning is vital to avoid artifacts that can confound or prevent future analyses. Muscles carefully processed using this protocol are suitable for further evaluation using immunohistochemistry, immunofluorescence, special histochemical stains, and immuoblotting. © 2016 by John Wiley & Sons, Inc.","PeriodicalId":37980,"journal":{"name":"Current protocols in mouse biology","volume":"6 1","pages":"343 - 363"},"PeriodicalIF":0.0,"publicationDate":"2016-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpmo.19","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"50901623","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 6
Measurement of Intestinal and Peripheral Cholesterol Fluxes by a Dual‐Tracer Balance Method 双示踪平衡法测量肠道和外周胆固醇通量
Current protocols in mouse biology Pub Date : 2016-12-01 DOI: 10.1002/cpmo.16
O. Ronda, Theo H. Dijk, H. Verkade, A. Groen
{"title":"Measurement of Intestinal and Peripheral Cholesterol Fluxes by a Dual‐Tracer Balance Method","authors":"O. Ronda, Theo H. Dijk, H. Verkade, A. Groen","doi":"10.1002/cpmo.16","DOIUrl":"https://doi.org/10.1002/cpmo.16","url":null,"abstract":"Long‐term elevated plasma cholesterol levels put individuals at risk for developing atherosclerosis. Plasma cholesterol levels are determined by the balance between cholesterol input and output fluxes. Here we describe in detail the methodology to determine the different cholesterol fluxes in mice. The percentage of absorbed cholesterol is calculated from a stable isotope–based double‐label method. Cholesterol synthesis is calculated from MIDA after 13C‐acetate enrichment. Cholesterol is removed from the body via the feces. The fecal excretion route is either biliary or non‐biliary. The non‐biliary route is dominated by trans‐intestinal cholesterol efflux, or TICE. Biliary excretion of cholesterol is measured by collecting bile. Non‐biliary excretion is calculated by computational modeling. In this article, we describe methods and procedures to measure and calculate dietary intake of cholesterol, fractional cholesterol absorption, fecal neutral sterol output, biliary cholesterol excretion, TICE, cholesterol synthesis, peripheral fluxes, and whole‐body cholesterol balance. © 2016 by John Wiley & Sons, Inc.","PeriodicalId":37980,"journal":{"name":"Current protocols in mouse biology","volume":"6 1","pages":"408 - 434"},"PeriodicalIF":0.0,"publicationDate":"2016-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpmo.16","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"50901772","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 18
Islet Insulin Secretion Measurements in the Mouse 小鼠胰岛胰岛素分泌测量
Current protocols in mouse biology Pub Date : 2016-09-01 DOI: 10.1002/cpmo.14
Alison Hugill, Kenju Shimomura, Roger D. Cox
{"title":"Islet Insulin Secretion Measurements in the Mouse","authors":"Alison Hugill,&nbsp;Kenju Shimomura,&nbsp;Roger D. Cox","doi":"10.1002/cpmo.14","DOIUrl":"10.1002/cpmo.14","url":null,"abstract":"<p>This article describes detailed protocols for in vitro measurements of insulin function and secretion in isolated mouse islets for the analysis of glucose homeostasis. We specify a method of enzyme digestion and hand picking to isolate and release the greatest number of high quality islets from the pancreas of the mouse. We describe an effective method for generating dynamic measurements of insulin secretion using a perifusion assay including a detailed protocol for constructing a peristaltic pump and tubing assembly. In addition we describe an alternative and simple technique for measuring insulin secretion using static incubation of isolated islets. © 2016 by John Wiley &amp; Sons, Inc.</p>","PeriodicalId":37980,"journal":{"name":"Current protocols in mouse biology","volume":"6 3","pages":""},"PeriodicalIF":0.0,"publicationDate":"2016-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpmo.14","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"34354380","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 5
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