Name: transformation of probiotic yeast and their recovery from gastrointestinal immune tissues following oral gavage in mice
Uploaded: 2016-02-08T14:00:00.000+00:00
Duration: 12 min 12 s
Description: Watch this Scientific Journal Video about Transformation of Probiotic Yeast and Their Recovery from Gastrointestinal Immune Tissues Following Oral Gavage in Mice at JoVE.com

저자는 트레이시 J. 어린 양에게 수여 면역학 및 면역 주사와 NIH 새로운 혁신 상 (1DP2AI112242-01)에 대한 아동 센터를 통해 자금을 인정합니다. 저자는 또한 rad1 S.의 관대 한 기여 나탈리아 P. Degtyareva 감사합니다 cerevisiae의.

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	<li>Edwards-Ingram, L., Gitsham, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genotypic+and+physiological+characterization+of+Saccharomyces+boulardii,+the+probiotic+strain+of+Saccharomyces+cerevisiae.">Genotypic and physiological characterization of Saccharomyces boulardii, the probiotic strain of Saccharomyces cerevisiae.</a> Applied and environmental microbiology. 73 (8), 2458-2467 (2007).</li><li>Hudson, L. E., Fasken, M. B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Functional+Heterologous+Protein+Expression+by+Genetically+Engineered+Probiotic+Yeast+Saccharomyces+boulardii.">Functional Heterologous Protein Expression by Genetically Engineered Probiotic Yeast Saccharomyces boulardii.</a> PloS one. 9 (11), 1-12 (2014).</li><li>Guthrie, C., Fink, G. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Methods in Enzymology, Volume 149: Guide to Yeast Genetics and Molecular Biology. , (1991).</li><li>DiCarlo, J. E., Norville, J. E., Mali, P., Rios, X., Aach, J., Church, G. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genome+engineering+in+Saccharomyces+cerevisiae+using+CRISPR-Cas+systems.">Genome engineering in Saccharomyces cerevisiae using CRISPR-Cas systems.</a> Nucleic acids research. 41 (7), 4336-4343 (2013).</li><li>Sander, J. D., Joung, J. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=CRISPR-Cas+systems+for+editing,+regulating+and+targeting+genomes.">CRISPR-Cas systems for editing, regulating and targeting genomes.</a> Nature biotechnology. 32 (4), 347-355 (2014).</li><li>Lorenz, M. C., Muir, R. S., Lim, E., McElver, J., Weber, S. C., Heitman, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gene+disruption+with+PCR+products+in+Saccharomyces+cerevisiae.">Gene disruption with PCR products in Saccharomyces cerevisiae.</a> Gene. 158, 113-117 (1995).</li><li>Hamedi, H., Misaghi, A., Modarressi, M. H., Salehi, T. Z., Khorasanizadeh, D., Khalaj, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Generation+of+a+Uracil+Auxotroph+Strain+of+the+Probiotic+Yeast+Saccharomyces+boulardii+as+a+Host+for+the+Recombinant+Protein+Production.">Generation of a Uracil Auxotroph Strain of the Probiotic Yeast Saccharomyces boulardii as a Host for the Recombinant Protein Production.</a> Avicenna Journal of Medical Biotechnology. 5 (1), 29-34 (2013).</li><li>Hinnen, A., Hicks, J. B., Fink, G. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transformation+of+yeast.">Transformation of yeast.</a> Proceedings of the National Academy of Sciences of the United States of America. 75 (4), 1929-1933 (1978).</li><li>Hashimoto, H., Morikawa, H., Yamada, K., Kimura, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+novel+method+for+transformation+of+intact+yeast+cells+by+electroinjection+of+plasmid+DNA.">A novel method for transformation of intact yeast cells by electroinjection of plasmid DNA.</a> Appl Microbiol Biotechnol. 21, 336-339 (1985).</li><li>Becker, D., Guarente, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-efficiency+transformation+of+yeast+by+electroporation.">High-efficiency transformation of yeast by electroporation.</a> Methods in Enzymology, Volume 149: Guide to Yeast Genetics and Molecular Biology. , 182-187 (1991).</li><li>Benatuil, L., Perez, J. M., Belk, J., Hsieh, C. -. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+improved+yeast+transformation+method+for+the+generation+of+very+large+human+antibody+libraries.">An improved yeast transformation method for the generation of very large human antibody libraries.</a> Protein engineering, design &amp; selection : PEDS. 23 (4), 155-159 (2010).</li><li>Ito, H., Fukuda, Y., Murata, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transformation+of+intact+yeast+cells+treated+with+alkali+cations.">Transformation of intact yeast cells treated with alkali cations.</a> Journal of Bacteriology. 153 (1), 166-168 (1983).</li><li>Kawai, S., Hashimoto, W., Murata, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transformation+of+Saccharomyces+cerevisiae+and+other+fungi:+methods+and+possible+underlying+mechanism.">Transformation of Saccharomyces cerevisiae and other fungi: methods and possible underlying mechanism.</a> Bioengineered bugs. 1 (6), 395-403 (2010).</li><li>Zheng, H. Z., Liu, H. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Yeast+transformation+process+studied+by+fluorescence+labeling+technique.">Yeast transformation process studied by fluorescence labeling technique.</a> Bioconjugate Chemistry. 16, 250-254 (2005).</li><li>Gietz, R. D., Woods, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+transformation+of+yeast.">Genetic transformation of yeast.</a> BioTechniques. 30 (4), 816-831 (2001).</li><li>JoVE Science Education Database. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+ELISA+Method.+.">The ELISA Method. .</a> Basic Methods in Cellular and Molecular Biology. , (2015).</li><li>Gallagher, S., Chakavarti, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Immunoblot+analysis.">Immunoblot analysis.</a> Journal of visualized experiments : JoVE. (16), (2008).</li><li>Horn, C. C., Kimball, B. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Why+Can't+Rodents+Vomit?+A+Comparative+Behavioral,+Anatomical,+and+Physiological+Study.">Why Can't Rodents Vomit? A Comparative Behavioral, Anatomical, and Physiological Study.</a> PLoS ONE. 8 (4), (2013).</li><li>Hoggatt, A. F., Hoggatt, J., Honerlaw, M., Pelus, L. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+spoonful+of+sugar+helps+the+medicine+go+down:+a+novel+technique+to+improve+oral+gavage+in+mice.">A spoonful of sugar helps the medicine go down: a novel technique to improve oral gavage in mice.</a> Journal of the American Association for Laboratory Animal Science : JAALAS. 49 (3), 329-334 (2010).</li><li>Johnson, M., Shayne, C. G., Kemper, C. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+rat.">The rat.</a> Animal Models in Toxicology. , 50-193 (2006).</li><li>Brown, A. P., Dinger, N., Levine, B. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Stress+produced+by+gavage+administration+in+the+rat.">Stress produced by gavage administration in the rat.</a> Contemporary topics in laboratory animal science / American Association for Laboratory Animal Science. 39, 17-21 (2000).</li><li>Jacoby, R., Fox, J., Davisson, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biology+and+diseases+of+mice.">Biology and diseases of mice.</a> Laboratory animal medicine. , 35-133 (2002).</li><li>Schulz, O., Pabst, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Antigen+sampling+in+the+small+intestine.">Antigen sampling in the small intestine.</a> Trends in immunology. 34 (4), 155-161 (2013).</li><li>McDole, J. R., Wheeler, L. W., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Goblet+cells+deliver+luminal+antigen+to+CD103++dendritic+cells+in+the+small+intestine.">Goblet cells deliver luminal antigen to CD103+ dendritic cells in the small intestine.</a> Nature. 483 (7389), 345-349 (2012).</li><li>Farache, J., Koren, I., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Luminal+bacteria+recruit+CD103++dendritic+cells+into+the+intestinal+epithelium+to+sample+bacterial+antigens+for+presentation.">Luminal bacteria recruit CD103+ dendritic cells into the intestinal epithelium to sample bacterial antigens for presentation.</a> Immunity. 38 (3), 581-595 (2013).</li><li>Burke, D., Dawson, D., Stearns, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Methods in yeast genetics: a Cold Spring Harbor Laboratory course manual. , (2000).</li><li>Boeke, J. D., LaCroute, F., Fink, G. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+positive+selection+for+mutants+lacking+orotidine-5'-phosphate+decarboxylase+activity+in+yeast:+5-fluoro-orotic+acid+resistance.">A positive selection for mutants lacking orotidine-5'-phosphate decarboxylase activity in yeast: 5-fluoro-orotic acid resistance.</a> Molecular &amp; general genetics : MGG. 197 (2), 345-346 (1984).</li><li>Chattoo, B. B., Sherman, F., Azubalis, D. A., Fjellstedt, T. A., Mehnert, D., Ogur, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Selection+of+lys2+mutants+of+the+yeast+Saccharomyces+cerevisiae+by+the+utilization+of+alpha-aminoadipate.">Selection of lys2 mutants of the yeast Saccharomyces cerevisiae by the utilization of alpha-aminoadipate.</a> Genetics. 93, 51-65 (1979).</li><li>Toyn, J. H., Gunyuzlu, P. L., White, W. H., La Thompson, ., Hollis, G. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+counterselection+for+the+tryptophan+pathway+in+yeast:+5-fluoroanthranilic+acid+resistance.">A counterselection for the tryptophan pathway in yeast: 5-fluoroanthranilic acid resistance.</a> Yeast. 16 (6), 553-560 (2000).</li><li>Singh, A., Sherman, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+and+physiological+characterization+of+met15+mutants+of+Saccharomyces+cerevisiae:+a+selective+system+for+forward+and+reverse+mutations.">Genetic and physiological characterization of met15 mutants of Saccharomyces cerevisiae: a selective system for forward and reverse mutations.</a> Genetics. 81, 75-97 (1975).</li><li>Singh, A., Sherman, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characteristics+and+relationships+of+mercury+resistant+mutants+and+methionine+auxotrophs+of+yeast.">Characteristics and relationships of mercury resistant mutants and methionine auxotrophs of yeast.</a> Journal of Bacteriology. 118 (3), 911-918 (1974).</li><li>McCoy, S. L., Hausman, F. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantification+of+DNA+binding+to+cell-surfaces+by+flow+cytometry.">Quantification of DNA binding to cell-surfaces by flow cytometry.</a> Journal of Immunological Methods. 241, 141-146 (2000).</li><li>Gietz, R. D., Schiestl, R. H., Willems, A. R., Woods, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Studies+on+the+transformation+of+intact+yeast+cells+by+the+LiAc/SS-DNA/PEG+procedure.">Studies on the transformation of intact yeast cells by the LiAc/SS-DNA/PEG procedure.</a> Yeast. 11, 355-360 (1995).</li><li>Zhang, Z., Chisti, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Plasmid+stability+in+recombinant+saccharomyces+cerevisiae.">Plasmid stability in recombinant saccharomyces cerevisiae.</a> Science. 14 (4), 401-435 (1996).</li><li>Lorang, J. M., Tuori, R. P., Martinez, J. P., Sawyer, T. L., Redman, R. S., Rollins, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Green+Fluorescent+Protein+Is+Lighting+Up+Fungal+Biology.">Green Fluorescent Protein Is Lighting Up Fungal Biology.</a> Applied and Environmental Microbiology. 67 (5), 1987-1994 (2001).</li><li>Thompson, J. R., Register, E., Curotto, J., Kurtz, M., Kelly, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+improved+protocol+for+the+preparation+of+yeast+cells+for+transformation+by+electroporation.">An improved protocol for the preparation of yeast cells for transformation by electroporation.</a> Yeast. 14, 565-571 (1998).</li><li>Damsch, S., Eichenbaum, G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gavage-related+reflux+in+rats:+identification,+pathogenesis,+and+toxicological+implications+(review).">Gavage-related reflux in rats: identification, pathogenesis, and toxicological implications (review).</a> Toxicologic pathology. 39, 348-360 (2011).</li><li>Desai, M., Labhasetwar, V., Amidon, G., Levy, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gastrointestinal+uptake+of+biodegradable+nanoparticles:+effect+of+particle+size.">Gastrointestinal uptake of biodegradable nanoparticles: effect of particle size.</a> Pharma. 13 (12), (1994).</li><li>Shakweh, M., Ponchel, G., Fattal, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Particle+uptake+by+Peyer's+patches:+a+pathway+for+drug+and+vaccine+delivery.">Particle uptake by Peyer's patches: a pathway for drug and vaccine delivery.</a> Expert opinion on drug delivery. 1 (1), (2004).</li><li>Szymanski, E. P., Kerscher, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Budding+yeast+protein+extraction+and+purification+for+the+study+of+function,+interactions,+and+post-translational+modifications.">Budding yeast protein extraction and purification for the study of function, interactions, and post-translational modifications.</a> Journal of visualized experiments : JoVE. (80), e50921 (2013).</li><li>Hashimoto, S., Ogura, M., Aritomi, K., Hoshida, H., Nishizawa, Y., Akada, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isolation+of+Auxotrophic+Mutants+of+Diploid+Industrial+Yeast+Strains+after+UV+Mutagenesis.">Isolation of Auxotrophic Mutants of Diploid Industrial Yeast Strains after UV Mutagenesis.</a> Applied and environmental microbiology. 71 (1), 312-319 (2005).</li></ol>

저자는 공개 아무것도 없어.

함께, 프로토콜은 여기에 장에 이종 치료 단백질의 전달을위한 개발과 영양 요 바이오 틱 효모 균주의 시험에 필요한 필수 단계를 설명합니다. 이 조작 및 재조합 바이오 틱 효모의 테스트는 개별 실험실은 익숙하지 않을 수있는 기술과 자원을 필요로한다. 많은 선행 연구가 여러 효모와 마우스 균주에 대해 위의 프로토콜을 설명하고 있지만 따라서, 이러한 방법은 저자의 지식이 상세한, 통합 형...

프로 바이오 틱 미생물을 효​​율적이고 경제적으로 위장관에 이종 단백질을 전달하는 흥미로운 잠재적 인 수단이다. 이 생물은 아직 1 식민지화하지 않는 위장관 통과 생존 제어 투약이 가능하고 약물 표현에 노출을 제한 할 수있다. 또한, 쉽게 대규모 이종 단백질을 생산하는 이들 미생물을 설계하는 능력은 그들을 합성 전달 입자 경제적 인 대안을 렌더링한다. 최근 2 boulardii에 생균제 효모 사카의 영양 요 구성 균주를 사용하여 입증하지만, 이러한 접근법의 개발은, 효모 및 분자 생물학에서 동물 처리 기술 및 면역 학적 방법에 이르기까지 전통적으로 특정 연구에서 결합되지 실험 기법에 대한 지식을 필요로한다. 본원에 기재된 각각의 방법은 그 자체로 신규 아니다 따라서 비록실험 프로토콜이 원고의 목적은 쥐 위장관에 약물 전달 비히클로서 생균제 효모 실험 테스트에 필요한 기술을 통합 도입을 제공하는 것이다. 제공에 필수적인 프로토콜의 편집이다 : 쉽게 유 전적으로 조작 할 수있는 효모의 영양 요 구성 돌연변이 균주의 1) 생성; 효모 배양 2) 변환은 이종 단백질을 발현하는 단계; 구강 투여를 통해 장에 재조합 효모의 3) 관리; 자신의 이종 단백질 발현의 쥐 소장 및 평가에서 가능한 재조합 바이오 틱 효모의 4) 복구. 다수의 양성 및 음성 선택 방법 효모 종의 조작에 존재하지만 첫째, 이러한 영양 요 구성 마커를 사용함으로써 부정적 선택 효율 효모 형질 전환 및 선택 될 수있는 용이성을 모두 증가시킨다. 공동 항생제를 사용하여 형질 전환 체의 긍정적 인 선택,ntrast 크게 효모 조작의 비용을 증가시킨다. 또한, 항생제 함유 고체 배지에 효모의 선택은 밖으로 합성 드롭 고체 배지 (게시되지 않은 관찰)에 영양 요 효모의 선택에 대해 변형되지 않은 배경 식민지의 증가 성장을 할 수 있습니다. 영양 요 구성 효모가 필수 아미노산 또는 우라실의 합성에 중요한 효소가 결여 균주이다. 효모가 필수 대사 물질이 부족 미디어 밖으로 합성 드롭에 도금 할 때 따라서 음의 선택이 가능 누락 된 대사 또는 대사 유전자로 보충하면 효모는 증가 할 수 있습니다. 많은 일반적으로 사용되는 사카로 마이 세스 세레 비지에 실험실 균주는 이미 사실 영양 요 구성 돌연변이 3입니다. 공업, 임상, 바이오 틱 및 효모 그러나, 일반적으로 필요한 모든 영양소를 합성하는 능력 prototrophic이다. 효모를보다 효율적으로 유전자 조작을 사용하려면, 영양 요 구성 유전자를 선택적으로 타겟팅 할 수 있습니다항생제없이 선택 될 수있는 균주를 생성한다. 6 - 영양 요 구성 마커 유전자의 구체적인 타겟팅 CRISPR / Cas9 4 타겟으로 최근에 상동 재조합에 의존 PCR 매개 유전자 파괴를 통해 달성 될 수있다. 또한, UV 돌연변이 유발 빠르게도 7 기술적으로 어려운 여러 플라스미드와 함께하는 형질 전환 효모 균주의 영양 요 구성 돌연변이 체를 생성 할 수 있습니다. PCR 타겟팅 및 CRISPR / Cas9 광범위 다른 설명되었지만,이 원고 한 음성 선별보다는 효모 형질 전환 체의 양성 항생제 선택할 수 영양 요 구성 균주를 제작하는 UV 돌연변이 유발 접근법을 설명하는 상세한 프로토콜 부분에 제시 하였다. 이종 단백질의 경구 전달을 위해 같은 영양 요 구성 균주의 사용에 필요한 다음 단계는 플라스미드 DNA로 형질 전환 효모이다. 찬성의 첫 번째 성공적인 변환 이후1978 사카로 마이 세스 세레 비지에 (8)에 대해보고 된 spheroplasts t는 수많은 변형 효율을 증가 효모 종은 유 전적으로 변형 될 수있는 것을 특징으로 완화되었다. S.로 DNA의 성공적인 전환을위한 전기 사용 cerevisiae를 먼저 1985 9에서 설명하고, 이후지지 삼투 셀 (10)에 1 M 소르비톨 배양 첨가를 통해 개선되었다. 전기 효율은 또한 효모 종 및 균주, 세포 수와 성장 단계, 전기 체적, 전계 강도, 및 특정 버퍼 (11)에 의존하는 것으로 나타났다. 이 특별한 장비를 필요로하지 않기 때문에 원래 이토 외. (12)에 의해 기재된 리튬 아세테이트 (LiOAc) 변환은, 가장 일반적으로 사용되는 프로토콜 사이에서 변환된다. 추가의 분석은 세포를 중간 로그 상에 수집되는 경우 LiOAc 효모 변환 효율을 크게 증가시키는 것으로 나타났다열 성장은 42 ° C에서 12 폴리에틸렌 글리콜 (PEG)과 DNA의 존재 하에서 충격된다. PEG와 전체 그대로 효모의 배양이 가능 세포막에 DNA의 첨부 파일을 개선을 통해뿐만 아니라 막 (13)에 다른 효과를 통해, 효율적인 변환을 위해 필수적이다. 리튬 자체도 손상 전지 (14)의 투과성을 증가시킨다. 대부분의 실험실 S. 있지만 세레 비지에 균주 쉽게 LiOAc 변형 3을 이용하여 변환 될 수 있고, 다른 효모 종은 더욱 효율적으로 다른 프로토콜을 사용하여 변환 될 수있다. 피 키아 파스 토리스, 예를 들면, 가장 효율적으로 변환 오히려 LiOAc 변환 (13)보다 전기를 통한 것이다. 이것은 중요하다, 따라서, 복수의 테스트 유 전적으로지지 않은 효모 균주를 수정을 시도 할 때와 변화의 방법은 배양 기간과 시약의 농도를 최적화 할 수 있습니다. 이 원고 따라서 LiOAc 그럴 모두 설명영양 요구 돌연변이와 야생 유형 S.의 변환을위한 기술로 ansformation 및 전기 boulardii에. 관심있는 독자는 효모 변형, 다른 프로토콜 및 행동 13, 15의 가능한 메커니즘의 추가 논의의 진화의 철저한 설명은 최근 리뷰에 관한 것이다. 플라스미드가 쉽게 검출 가능한 단백질을 코딩하는 효모로 형질 전환은 적절한 발현 이종 단백질의 기능을 보장하기 위해 다운 스트림 테스트 또한 필수적이다. 무수한 다른 단백질 치료 연구의 궁극적 인 목적 및 면역, ELISA, 및 다른 기술에 의해 단백질이 검출 가능한 항체에 따라 선택 될 수있다. 이러한 기술에 대한 프로토콜은 완전히 다른 16,17 설명되었지만, 표준 곡선과 비교하여 형질 전환 효모에서 이종 단백질 생산의 수준을 결정하기 위해 사용될 수있다. 목적 데모 및 표시 할효모 생물학 매우 일반적으로 사용되는 단백질의 성공적인 제조 방법이 원고는 형광 현미경을 사용하여 연속적인 검출이 가능 플라스미드 인코딩 녹색 형광 단백질 (GFP)로 형질 전환을 제공한다. 부품 셋, 넷에 설명 된대로 이종 단백질, 적절한 관리 및 위장관 조직 내에서 이러한 미생물의 검출 인 표현 바이오 틱 미생물의 생산에 똑같이 중요. 구강 투여를 통해 재조합 효모의 관리를 직접 C57BL / 6 마우스 자연스럽게 18 구토 능력이있는에서 위,에 효모의 제어 수량의 전달을 위해 수 있습니다. 그러나, 부적절한 동물 처리 및 위관은 식도 손상 및 천공, 위 천공, 기관 관리, 흡인 성 폐렴 (19, 20)로 이어질 수 있습니다. 빈약 한 기술과 경험 부족은 또한 쥐의 면역 반응 및 실험 resu의 변동성을 증가시킬 수있다구강 투여에 21,22 동물 응력에 기인 한 LTS. 적절한 기술의 연습 따라서뿐만 아니라 동물의 불편을 감쇠 할 수 있습니다뿐만 아니라, 실험 결과의 정밀도를 높일 수 있습니다. 이 원고는 설명하고 재조합 효모의 제어 용량의 관리를 위해 동물 처리 및 구강 투여를 보여줍니다. 마지막으로, 효모 및 이종 단백질의 존재를 림프 조직을 분석하여 재조합 효모의 성공적인 전달을 확인하는 것이 중요하다. 가장 용이하고 예측 효모의 존재를 검사 할 수 위장관 면역 조직 파이어 패치이다. 파이어 패치는 점막 면역 반응의 유도 (23)의 키 사이트입니다 소장을 따라 차 림프 기관입니다. 내강에서 항원 microfold 통해 transcellularly 상피 (M) 세포를 전달하고 묶 노출 시킴 따라서 파이어 패치로 출시제시 세포 D 항원 내강 내용 장내한다. 장 상피를 가로 지르는 입자 흡수도 술잔 세포에 의해 달성 될 수 있지만, 이러한 셀은 차지 입자 직경이 24 미만 0.02 ㎛ 인 것으로 나타났다. Transepithelial 수상 돌기는 CD103 + 수지상 세포 (DC)도 25 루멘 장에서 작은 입자를 차지 연장; 그러나, CD103 + DC가 세균보다 큰 입자를 가지고 있음을 보여주는보고는 현재 존재하지 않는다. 따라서, 그대로 생균제 효모 직경 3-6 μm의 평균 크기 사이의, M 세포에 의해 용해 및 파이어 패치로 전송 될 가능성이 가장 높은. 여기에 설명 된 절차가 쉽게 바이오 틱 균의 흡수를 평가하기 위해 적응 될 수 있지만, 수집 및 생존 재조합 효모 파이어 패치 선별 프로토콜이다. 요약하면, 배달 재조합 효모 생균제 평가소장에 rapeutic 단백질은 동물의 취급 및 면역학에 분자 생물학에 걸친 실험 기술의 능력을 필요로한다. 1 여기를 제시 프로토콜) 쉽게 부정적인 항생제없이 선택할 수있는 세대 및 영양 요 효모 균주의 선별, 2) 다른 프로토콜은 효모를 변환 및 이종 단백질의 발현을 활성화, 3) 적절한 동물 처리 기술과 경구 위관의 시위 재조합 효모의 위장 납품 및 파이어 판 절제술 4) 프로토콜과 가능한 재조합 효모 및 기능 이종 단백질을 스크리닝. 결합하여, 이러한 프로토콜은 생성 및 위장관에 이종 단백질 치료제를 제공 할 수있는 생균제 효모의 테스트를 허용한다.

<table><tbody><tr><td>SmartSpec 3000 Spectrophotometer</td><td>BioRad</td><td>170-2501</td><td>Example of spectrophotometer for determining cell concentration and OD&lt;sub&gt;600&lt;/sub&gt; of yeast cultures</td></tr><tr><td>New Brunswick Roller Drum</td><td>Eppendorf</td><td>M1053-4004</td><td>Example of roller drum for yeast culture incubation</td></tr><tr><td>UV Stratalinker 2400</td><td>Stratagene</td><td>400075-03</td><td>Example stratalinker</td></tr><tr><td>Stuart Colony Counter SC6PLUS</td><td>11983044</td><td>Fisher Scientific</td><td>Plate stand with magnification records colony count upon sensing pressure from pen</td></tr><tr><td>Scienceware Colony Counter&amp;nbsp;</td><td>F378620002</td><td>Bel-Art Scienceware</td><td>Hand held colony counter pen</td></tr><tr><td>Replica plating device</td><td>Fisherbrand</td><td>09-718-1</td><td>Example of replica plating stand and pads&amp;nbsp;</td></tr><tr><td>Velveteen squares</td><td>Fisherbrand</td><td>09-718-2</td><td></td></tr><tr><td>L shaped sterile cell spreaders&amp;nbsp;</td><td>Fisherbrand</td><td>14665230</td><td></td></tr><tr><td>Deoxyribonucleic acid, single stranded from salmon testes</td><td>Sigma-Aldrich</td><td>D7656-1ML</td><td>Example carrier DNA for yeast LiOAc transformation</td></tr><tr><td>Gavage needles</td><td>Braintree Scientific</td><td>N-PK&amp;nbsp;002</td><td>For mice 15-20 g, the suggested needle is a 22 gauge (1.25 mm ball), 1 in long, straight reusable gavage needle. For mice weighing greater than 20&amp;nbsp;g, 20 gauge or larger straight or curved gavage needles may be used</td></tr><tr><td>1 ml sterile slip-tip disposable tuberculin syringe</td><td>Becton Dickinson</td><td>BD 309659</td><td></td></tr><tr><td>Blunt forceps such as Electron Microscopy Sciences 7&quot; (178 mm) serrated tip, broad grip forceps</td><td>Electron Microscopy Sciences</td><td>77937-28</td><td>Example of blunt forceps needed for dissection&amp;nbsp;</td></tr><tr><td>Straight and curved dissection scissors</td><td>Electron Microscopy Sciences</td><td>72966-02 and 72966-03</td><td>Examples of scissors needed for dissection</td></tr><tr><td>IMDM</td><td>Life technologies</td><td>12440053</td><td></td></tr></tbody></table>

transformation of probiotic yeast and their recovery from gastrointestinal immune tissues following oral gavage in mice

Development of recombinant oral therapy would allow for more direct targeting of the mucosal immune system and improve the ability to combat gastrointestinal disorders. Adapting probiotic yeast in particular for this approach carries several advantages. These strains have not only the potential to synthesize a wide variety of complex heterologous proteins but are also capable of surviving and protecting those proteins during transit through the intestine. Critically, however, this approach requires expertise in many diverse laboratory techniques not typically used in tandem. Furthermore, although individual protocols for yeast transformation are well characterized for commonly used laboratory strains, emphasis is placed here on alternative approaches and the importance of optimizing transformation for less well characterized probiotic strains. Detailing these methods will help facilitate discussion as to the best approaches for testing probiotic yeast as oral drug delivery vehicles and indeed serve to advance the development of this novel strategy for gastrointestinal therapy.

UV 조사 다음 생존 곡선의 생성은 별개 집락 형성 단위 (CFU)를 형성 할 수 있도록 희석하여 효모 세포의 도금을 필요로한다. 상기와 같이 수집 된 각 500 μL 샘플을 약 5 × 106 세포를 포함 그러나, 접시 당 100보다 큰 식민지를 정확하게 구별하기가 어렵다. 희석되지 않은 샘플과 조사 셀 직렬 1시 10분 희석 따라서도 1에서 설명한 바와 같이 CFU는 각각 UV 조?...

Watch this Scientific Journal Video about Transformation of Probiotic Yeast and Their Recovery from Gastrointestinal Immune Tissues Following Oral Gavage in Mice at JoVE.com

Transformation of Probiotic Yeast and Their Recovery from Gastrointestinal Immune Tissues Following Oral Gavage in Mice

개발 변환 관리 및 생균제 효모 사카 boulardii에의 외래 단백질 발현을 테스트 할 수있는 기술의 통일 된 설명은 여기에 제시 하였다.

마우스의 구강 투여에 따라 생균제 효모의 변환 및 위장 면역 조직에서 그들의 복구

Development of recombinant oral therapy would allow for more direct targeting of the mucosal immune system and improve the ability to combat ...

transformation-probiotic-yeast-their-recovery-from-gastrointestinal

Research

JoVE Journal

Immunology and Infection

12.3K 조회수.  Emory University School of Medicine.  개발 변환 관리 및 생균제 효모 사카 boulardii에의 외래 단백질 발현을 테스트 할 수있는 기술의 통일 된 설명은 여기에 제시 하였다. 

비디오: Transformation of Probiotic Yeast and Their Recovery from Gastrointestinal Immune Tissues Following Oral Gavage in Mice

Quantitative Polymerase Chain Reaction-based Analyses of Murine Intestinal Microbiota After Oral Antibiotic Treatment

The gut microbiota has a central influence on human health. Microbial dysbiosis is associated with many common immunopathologies such as inflammatory bowel disease, asthma and arthritis. Thus, understanding the mechanisms underlying microbiota-immune system crosstalk is of crucial importance. Antibiotic administration, while aiding pathogen clearance, also induces drastic changes in the size and composition of intestinal bacterial communities which can have an impact on human health. Antibiotic treatment in mice recapitulates the impact and long-term changes in human microbiota from antibiotic treated patients, and enables investigation of the mechanistic links between changes in microbial communities and immune cell function. While several methods for antibiotic treatment of mice have been described, some of them induce severe dehydration and weight-loss complicating the interpretation of the data. Here, we provide two protocols for oral antibiotic administration which can be used for long-term treatment of mice without inducing major weight-loss. These protocols make use of a combination of antibiotics that target both Gram-positive and Gram-negative bacteria and can be provided either ad libitum in the drinking water or by oral gavage. Moreover, we describe a method for the quantification of microbial density in fecal samples by qPCR which can be used to validate the efficacy of the antibiotic treatment. The combination of these approaches provides a reliable methodology for the manipulation of the intestinal microbiota and the study of the effects of antibiotic treatment in mice.

Here we provide detailed protocols for the oral administration of antibiotics to mice, collection of fecal samples, DNA extraction and quantification of fecal bacteria by qPCR.

구두 항생제 치료 후 Murine 장 Microbiota의 정량적 중 합 효소 연쇄 반응-기반 분석

The gut microbiota has a central influence on human health. Microbial dysbiosis is associated with many common immunopathologies such as inflammatory ...

연구

면역학 및 감염병학

Probiotic Studies in Neonatal Mice Using Gavage

Adult mouse models have been widely used to understand the mechanism behind disease progression in humans. The applicability of studies done in adult mouse models to neonatal diseases is limited. To better understand disease progression, host responses and long-term impact of interventions in neonates, a neonatal mouse model likely is a better fit. The sparse use of neonatal mouse models can in part be attributed to the technical difficulties of working with these small animals. A neonatal mouse model was developed to determine the effects of probiotic administration in early life and to specifically assess the ability to establish colonization in the newborn mouse intestinal tract. Specifically, to assess probiotic colonization in the neonatal mouse, Lactobacillus plantarum (LP) was delivered directly into the neonatal mouse gastrointestinal tract. To this end, LP was administered to mice by feeding through intra-esophageal (IE) gavage. A highly reproducible method was developed to standardize the process of IE gavage that allows an accurate administration of probiotic dosages while minimizing trauma, an aspect particularly important given the fragility of newborn mice. Limitations of this process include possibilities of esophageal irritation or damage and aspiration if gavaged incorrectly. This approach represents an improvement on current practices because IE gavage into the distal esophagus reduces the chances of aspiration. Following gavage, the colonization profile of the probiotic was traced using quantitative polymerase chain reaction (qPCR) of the extracted intestinal DNA with LP specific primers. Different litter settings and cage management techniques were used to assess the potential for colonization-spread. The protocol details the intricacies of IE neonatal mouse gavage and subsequent colonization quantification with LP.

This study details the process of gavaging precise amounts of probiotics to neonatal mice. The experimental set-up was optimized to include but is not limited to probiotic dosing, methods of administration, and quantification of bacteria in the intestines.

Gavage를 사용 하 여 신생아 쥐에 있는 Probiotic 연구

Adult mouse models have been widely used to understand the mechanism behind disease progression in humans. The applicability of studies done in adult ...

Visualization of Candida albicans in the Murine Gastrointestinal Tract Using Fluorescent In Situ Hybridization

Candida albicans is a fungal component of the gut microbiota in humans and many other mammals. Although C. albicans does not cause symptoms in most colonized hosts, the commensal reservoir does serve as a repository for infectious disease, and the presence of high fungal titers in the gut is associated with inflammatory bowel disease. Here, we describe a method to visualize C. albicans cell morphology and localization in a mouse model of stable gastrointestinal colonization. Colonization is established using a single dose of C. albicans in animals that have been treated with oral antibiotics. Segments of gut tissue are fixed in a manner that preserves the architecture of luminal contents (microorganisms and mucus) as well as the host mucosa. Finally, fluorescent in situ hybridization is performed using probes against fungal rRNA to stain for C. albicans and hyphae. A key advantage of this protocol is that it allows for simultaneous observation of C. albicans cell morphology and its spatial association with host structures during gastrointestinal colonization.

Visualization of Candida albicans in the Murine Gastrointestinal Tract Using Fluorescent In Situ Hybridization

The purpose of this protocol is to visualize Candida albicans cell shape and localization in the mammalian gastrointestinal tract.

시투 혼성화에서 형광을 사용하여 뮤린 위장에서 칸디다 알비칸스의 시각화

Candida albicans is a fungal component of the gut microbiota in humans and many other mammals. Although C. albicans does not cause symptoms in most ...

Therapeutic Evaluation of Fecal Microbiota Transplantation in an Interleukin 10-Deficient Mouse Model

With the development of microecology in recent years, the relationship between intestinal bacteria and inflammatory bowel disease (IBD) has attracted considerable attention. Accumulating evidence suggests that dysbiotic microbiota plays an active role in triggering or worsening the inflammatory process in IBD and that fecal microbiota transplantation (FMT) is an attractive therapeutic strategy since transferring a healthy microbiota to IBD patient could restore the appropriate host-microbiota communication. However, the molecular mechanisms are unclear, and the efficacy of FMT has not been very well established. Thus, further studies in animal models of IBD are necessary. In this method, we applied FMT from wild-type C57BL/6J mice to IL-10 deficient mice, a widely used mouse model of colitis. The study elaborates on collecting fecal pellets from the donor mice, making the fecal solution/suspension, administering the fecal solution, and monitoring the disease. We found that FMT significantly mitigated the cardiac impairment in IL-10 knockout mice, underlining its therapeutic potential for IBD management.

The interaction of genetic susceptibility, mucosal immunity, and intestinal microecological environment is involved in the pathogenesis of inflammatory bowel disease (IBD). In this study, we applied fecal microbiota transplantation to IL-10 deficient mice and investigated its impact on colonic inflammation and heart function.

인터루킨 10-결핍 마우스 모델에서 분변 미생물 이식의 치료 평가

With the development of microecology in recent years, the relationship between intestinal bacteria and inflammatory bowel disease (IBD) has attracted ...

T4 Phage를 사용한 Murine Models의 Gut Phage-Bacteria Dynamics 탐색

T4 Bacteriophage and E. coli Interaction in the Murine Intestine: A Prototypical Model for Studying Host-Bacteriophage Dynamics In Vivo

Bacteriophages (phages) are viruses that infect bacteria with species- and strain-level specificity and are the most abundant biological entities across all known ecosystems. Within bacterial communities, such as those found in the gut microbiota, phages are implicated in regulating microbiota population dynamics and driving bacterial evolution. There has been renewed interest in phage research in the last decade, in part due to the host-specific killing capabilities of lytic phages, which offer a promising tool to counter the increasing threat of antimicrobial resistant bacteria. Furthermore, recent studies demonstrating that phages adhere to intestinal mucus suggest they may have a protective role in preventing bacterial invasion into the underlying epithelium. Importantly, like bacterial microbiomes, disrupted phageomes have been associated with worsened outcomes in diseases such as inflammatory bowel disease. Previous studies have demonstrated that phages can modulate the microbiome of animals and humans through fecal filtrate transplants, benefiting the host&#39;s health. With this recent wave of research comes the necessity to establish and standardize protocols for studying phages in the context of the gut microbiome. This protocol provides a set of procedures to study isolated T4 phages and their bacterial host, Escherichia coli,&#160;in the context of the murine gastrointestinal tract. The methods described here outline&#160;how to start from a phage lysate, administer it to mice and assess effects on bacterial host and phage levels.&#160;This protocol can be modified and applied to other phage-bacterial pairs and provides a starting point for studying host-phage dynamics in vivo.

저자 스포트라이트: Gnotobiotic Mice의 박테리오파지 유도 면역 반응 조사

Bacteriophages (phages), viruses that infect bacteria, are an integral component of the gut microbiome. Though these symbiotic inhabitants drive bacterial fitness and population dynamics, little is understood about how they impact gut homeostasis and disease. This protocol studies isolated T4 phages within a mouse model, adaptable to other phage-bacterial pairs.

쥐 장에서 T4 박테리오파지와 대장균 상호작용: 생체 내 숙주-박테리오파지 역학 연구를 위한 프로토타입 모델

Bacteriophages (phages) are viruses that infect bacteria with species- and strain-level specificity and are the most abundant biological entities across ...

Preparation and Maintenance of Bioexclusion IsoPositive Cage Experiment for Human Fecal Transplantation into Germ-Free Mice

Germ-free mice are an important investigation tool for understanding the contribution of microorganisms in host health and disease, enabling assessment of the specific role of individuals, defined or complex groups of microorganisms in host response. Traditionally bred and reared in flexible-film or semi-rigid isolators, germ-free mouse husbandry and experimental manipulation are costly and require numerous trained staff and a large space footprint in animal housing facilities. The IsoPositive caging system allows for experimental manipulation of germ-free mice in individual, hermetically-sealed, positive-pressure isolator cages (isocages), reducing cost and enabling greater flexibility in experimental manipulations. 
Here, a protocol is described for transferring germ-free mice from breeding isolators to isocages and subsequent fecal transfer from human donor stool into mice to create stable long-term gut "humanized" mice for future studies. The materials and preparation needed for the utilization of the isocage system are described, including the use of chlorine-dioxide sterilant chemical sterilant to clean cages, supplies, equipment, and personal protective equipment. The methods for confirming the germ-free status of transferred mice and how to determine contamination in the caging system are discussed. The procedure for husbandry, including bedding, food, and water supply, is further discussed. The protocol for human fecal slurry preparation and gavage into germ-free mice to create gut "humanized" mice, along with stool collection to monitor the microbial community composition of these mice, are described. An experiment illustrates that two weeks post-human fecal transplant allows for stable colonization of donor microbiota in the murine hosts, enabling subsequent experimental usage. Furthermore, the collection of humanized mouse feces in viability preservation media, enabling use in further functional experiments, is described. Overall, these methods allow for the safe and effective establishment of humanized mouse communities in experimental gnotobiotic cages for further manipulation.

This protocol describes the best practices for germ-free mouse transfer to and housing in experimental single-cage isolators (isocages) while maintaining sterile conditions. Methods for fecal transplant into germ-free mice and the collection of viable bacteria from these gut "humanized" mice for further applications are discussed.

무균 마우스에 인간 배설물 이식을 위한 Bioexclusion IsoPositive 케이지 실험의 준비 및 유지 관리

Germ-free mice are an important investigation tool for understanding the contribution of microorganisms in host health and disease, enabling assessment of ...

Fecal Microbiota Transplantation and Detection of Prevalence of IgA-Coated Bacteria in the Gut

Gut microbiota exert pleiotropic roles in human health and disease. Fecal microbiota transplantation (FMT) is an effective method to investigate the biological function of intestinal bacteria as a whole or at the species level. Several different FMT methods have been published. Here, we present an FMT protocol that successfully depletes gut microbiota in a matter of days, followed by transplantation of fecal microbiota from fresh or frozen donor intestinal contents to conventional mice. Real time-PCR is applied to test the efficacy of bacterial depletion. Sequencing of the 16S ribosomal RNA (rRNA) is then applied to test the relative abundance and identity of gut microbiota in recipient mice. We also present a flow cytometry-based detection method of immunoglobulin A (IgA)-coated bacteria in the gut.

We established an efficient way to deplete intestinal bacteria in three days, and subsequently transplant fecal microbiota via gavage of fecal fluid prepared from fresh or frozen intestinal contents in mice. We also present an optimized method to detect the frequency of IgA-coated bacteria in the gut.

Gut microbiota exert pleiotropic roles in human health and disease. Fecal microbiota transplantation (FMT) is an effective method to investigate the ...

생물학

Investigating the Alleviating Effects of Bacillus cereus Administration on Colitis through Gut Microbiota Modulation

Gut microbiota dysbiosis is thought to exert a role in the progression of colitis. However, the precise standards for probiotic administration in alleviating colitis remain undefined. Most analysis methods rely on limited diversity and abundance of gut microorganisms. Therefore, observational studies cannot establish causation. In this study, we applied antibiotic-induced pseudo-germ-free mice to investigate the role of gut microbiota in regulating the probiotic effects of Bacillus&#160;cereus (B. cereus)&#160;on dextran sulfate sodium (DSS)-induced colitis in mice. This process allows for evaluating the bidirectional regulating effect of B. cereus&#160;supplementation on health and provides stable and reproducible results. Here, the detailed protocols for B. cereus cultivation, gavage operation, stool collection, and antibiotic clearance treatment on colitis mice are provided. The optimization methods are also applicable for other chronic inflammatory-associated disorders. The results showed that B. cereus administration decreased body weight loss, colon length shortening, disease activity index, and histopathological scores. However, treatment with antibiotics suppressed the positive effect of B. cereus on colitis. These results indicate that gut microbiota are needed for the alleviating effects of B. cereus on colitis. Therefore, exploring the beneficial effects of probiotics in this research is a promising approach for developing novel treatment strategies for alleviating the symptoms of chronic inflammatory-associated disorders.

Investigating the Alleviating Effects of Bacillus cereus Administration on Colitis through Gut Microbiota Modulation

Here, we present a protocol to establish the antibiotic-induced pseudo-germ-free dextran sulfate sodium-induced colitis mouse model to investigate the role of gut microbiota in regulating the positive effects of Bacillus cereus on colitis.

마우스의 구강 투여에 따라 생균제 효모의 변환 및 위장 면역 조직에서 그들의 복구

요약

더 많은 비디오 탐색

이 비디오의 챕터

구두 항생제 치료 후 Murine 장 Microbiota의 정량적 중 합 효소 연쇄 반응-기반 분석

Gavage를 사용 하 여 신생아 쥐에 있는 Probiotic 연구

시투 혼성화에서 형광을 사용하여 뮤린 위장에서 칸디다 알비칸스의 시각화

인터루킨 10-결핍 마우스 모델에서 분변 미생물 이식의 치료 평가

쥐 장에서 T4 박테리오파지와 대장균 상호작용: 생체 내 숙주-박테리오파지 역학 연구를 위한 프로토타입 모델

쥐 장에서 T4 박테리오파지와 대장균 상호작용: 생체 내 숙주-박테리오파지 역학 연구를 위한 프로토타입 모델

쥐 장에서 T4 박테리오파지와 대장균 상호작용: 생체 내 숙주-박테리오파지 역학 연구를 위한 프로토타입 모델

무균 마우스에 인간 배설물 이식을 위한 Bioexclusion IsoPositive 케이지 실험의 준비 및 유지 관리

Fecal Microbiota Transplantation and Detection of Prevalence of IgA-Coated Bacteria in the Gut

Investigating the Alleviating Effects of Bacillus cereus Administration on Colitis through Gut Microbiota Modulation