Name: the mplex protocol for multi-omic analyses of soil samples
Uploaded: 2018-05-30T14:00:00
Description: Watch this Scientific Journal Video about The MPLEx Protocol for Multi-omic Analyses of Soil Samples at JoVE.com

작가 네이 선 존슨 수치 준비에 그의 원조에 대 한 감사 하 고 싶습니다. 이 연구는 미국 에너지 부의 사무실의 생물학과 환경 연구 (게놈 과학 프로그램), 전환 (민트) 실험실 감독 연구 개발에 Microbiomes에 의해 투자 되는 팬-omics 프로그램에 의해 지원 되었다 퍼시픽 노스 웨스트 국립 연구소, 국가 연구소의 건강 국립 연구소의 환경 건강 과학 (R01 ES022190)에서 이니셔티브와 NIH (P42 ES027704). KEBJ는 R21 HD084788 개발 하 고 새로운 멀티-omic 추출 기법 검증 재정 지원에 대 한 감사 하 고 싶습니다. 이 작품에는 W. R. 윌 환경 분자 과학 실험실 (EMSL), 미상 국가 과학적인 사용자 시설 평화로운 북 서 국립 연구소 (PNNL)에서 수행 되었다. PNNL은 Battelle 계약 드-AC06-76RL01830에서 DOE에 대 한 운영 하는 멀티 프로그램 국립 연구소 이다.

참고: 매우 젖은 토양 이전 효과에 손해 없이 추출 추출의 lyophilized 수 있습니다. 젖은 토양 또한 사용 될 수 있지만 특정 비율에 시 약을 추가할 때 고려해 야 합니다.
참고:이 좋습니다 추출, 두 50 mL 튜브 (최대 50 mL 튜브 당 10 g 토양) 사이 분할 한다 당 건조 한 토양 중량의 20 g을 사용 하 여. 추출 또는 사용 가능한 샘플에 따라 다름 아래로 확장할 수 있습니다.
<p class="jove_co...

<ol>
	<li>Hultman, J., 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=Multi-omics+of+permafrost,+active+layer+and+thermokarst+bog+soil+microbiomes.">Multi-omics of permafrost, active layer and thermokarst bog soil microbiomes.</a> Nature. 521, 208-212 (2015).</li><li>White, R. 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=Moleculo+long-read+sequencing+facilitates+assembly+and+genomic+binning+from+complex+soil+metagenomes.">Moleculo long-read sequencing facilitates assembly and genomic binning from complex soil metagenomes.</a> mSystems. 1, (2016).</li><li>White, R. A., Callister, S. J., Moore, R. J., Baker, E. S., Jansson, 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=The+past,+present+and+future+of+microbiome+analyses.">The past, present and future of microbiome analyses.</a> Nat Protoc. 11, 4-8 (2016).</li><li>Ritchie, M. D., Holzinger, E. R., Li, R., Pendergrass, S. A., Kim, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Methods+of+integrating+data+to+uncover+genotype-phenotype+interactions.">Methods of integrating data to uncover genotype-phenotype interactions.</a> Nat Rev Genet. 16, 85-97 (2015).</li><li>Jansson, J. K., Baker, E. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+multi-omic+future+for+microbiome+studies.">A multi-omic future for microbiome studies.</a> Nat Microbiol. 1, (2016).</li><li>Domon, B., Aebersold, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Options+and+considerations+when+selecting+a+quantitative+proteomics+strategy.">Options and considerations when selecting a quantitative proteomics strategy.</a> Nat Biotechnol. 28, 710-721 (2010).</li><li>Marx, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Targeted+proteomics.">Targeted proteomics.</a> Nat Methods. 10, 19-22 (2013).</li><li>Roberts, L. D., Souza, A. L., Gerszten, R. E., Clish, C. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Targeted+metabolomics.">Targeted metabolomics.</a> Curr Protoc Mol Biol. , (2012).</li><li>Folch, J., Lees, M., Sloane Stanley, G. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+simple+method+for+the+isolation+and+purification+of+total+lipides+from+animal+tissues.">A simple method for the isolation and purification of total lipides from animal tissues.</a> J Biol Chem. 226, 497-509 (1957).</li><li>Nakayasu, E. S., 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=MPLEx:+a+Robust+and+Universal+Protocol+for+Single-Sample+Integrative+Proteomic,+Metabolomic,+and+Lipidomic+Analyses.">MPLEx: a Robust and Universal Protocol for Single-Sample Integrative Proteomic, Metabolomic, and Lipidomic Analyses.</a> mSystems. 1, (2016).</li><li>Bligh, E. G., Dyer, W. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+rapid+method+of+total+lipid+extraction+and+purification.">A rapid method of total lipid extraction and purification.</a> Can J Biochem Physiol. 37, 911-917 (1959).</li><li>Pomraning, K. R., 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=Multi-omics+analysis+reveals+regulators+of+the+response+to+nitrogen+limitation+in+Yarrowia+lipolytica.">Multi-omics analysis reveals regulators of the response to nitrogen limitation in Yarrowia lipolytica.</a> BMC Genomics. 17, 138 (2016).</li><li>Tisoncik-Go, J., 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=Integrated+Omics+Analysis+of+Pathogenic+Host+Responses+during+Pandemic+H1N1+Influenza+Virus+Infection:+The+Crucial+Role+of+Lipid+Metabolism.">Integrated Omics Analysis of Pathogenic Host Responses during Pandemic H1N1 Influenza Virus Infection: The Crucial Role of Lipid Metabolism.</a> Cell Host Microbe. 19, 254-266 (2016).</li><li>Kyle, J. E., 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=Uncovering+biologically+significant+lipid+isomers+with+liquid+chromatography,+ion+mobility+spectrometry+and+mass+spectrometry.">Uncovering biologically significant lipid isomers with liquid chromatography, ion mobility spectrometry and mass spectrometry.</a> Analyst. 141, 1649-1659 (2016).</li><li>Lovelace, E. S., 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=Silymarin+Suppresses+Cellular+inflammation+by+inducing+reparative+stress+signaling.">Silymarin Suppresses Cellular inflammation by inducing reparative stress signaling.</a> J Nat Prod. 78, 1990-2000 (1990).</li><li>Kim, Y. M., 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=Diel+metabolomics+analysis+of+a+hot+spring+chlorophototrophic+microbial+mat+leads+to+new+hypotheses+of+community+member+metabolisms.">Diel metabolomics analysis of a hot spring chlorophototrophic microbial mat leads to new hypotheses of community member metabolisms.</a> Front Microbiol. 6, 209 (2015).</li><li>Pomraning, K. R., 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=Comprehensive+Metabolomic,+Lipidomic+and+microscopic+profiling+of+Yarrowia+lipolytica+during+lipid+accumulation+identifies+targets+for+increased+lipogenesis.">Comprehensive Metabolomic, Lipidomic and microscopic profiling of Yarrowia lipolytica during lipid accumulation identifies targets for increased lipogenesis.</a> PLoS One. 10, e0123188 (2015).</li><li>Huang, E. L., 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=The+fungus+gardens+of+leaf-cutter+ants+undergo+a+distinct+physiological+transition+during+biomass+degradation.">The fungus gardens of leaf-cutter ants undergo a distinct physiological transition during biomass degradation.</a> Environ Microbiol Rep. 6, 389-395 (2014).</li><li>Deatherage Kaiser, B. L., 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=A+Multi-Omic+View+of+Host-Pathogen-Commensal+Interplay+in+Salmonella-Mediated+Intestinal+Infection.">A Multi-Omic View of Host-Pathogen-Commensal Interplay in Salmonella-Mediated Intestinal Infection.</a> PLoS One. 8, e67155 (2013).</li><li>Kim, Y. M., 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=Salmonella+modulates+metabolism+during+growth+under+conditions+that+induce+expression+of+virulence+genes.">Salmonella modulates metabolism during growth under conditions that induce expression of virulence genes.</a> Mol Biosyst. 9, 1522-1534 (2013).</li><li>Ansong, C., 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=A+multi-omic+systems+approach+to+elucidating+Yersinia+virulence+mechanisms.">A multi-omic systems approach to elucidating Yersinia virulence mechanisms.</a> Mol Biosyst. 9, 44-54 (2013).</li><li>Bordbar, 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=Model-driven+multi-omic+data+analysis+elucidates+metabolic+immunomodulators+of+macrophage+activation.">Model-driven multi-omic data analysis elucidates metabolic immunomodulators of macrophage activation.</a> Mol Syst Biol. 8, 558 (2012).</li><li>Hu, Z. 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=Metabolomic+response+of+human+skin+tissue+to+low+dose+ionizing+radiation.">Metabolomic response of human skin tissue to low dose ionizing radiation.</a> Mol Biosyst. 8, 1979-1986 (2012).</li><li>Perera, R., 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=Dengue+virus+infection+perturbs+lipid+homeostasis+in+infected+mosquito+cells.">Dengue virus infection perturbs lipid homeostasis in infected mosquito cells.</a> PLoS Pathog. 8, e1002584 (2012).</li><li>Gao, X., 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=A+reversed-phase+capillary+ultra-performance+liquid+chromatography-mass+spectrometry+(UPLC-MS)+method+for+comprehensive+top-down/bottom-up+lipid+profiling.">A reversed-phase capillary ultra-performance liquid chromatography-mass spectrometry (UPLC-MS) method for comprehensive top-down/bottom-up lipid profiling.</a> Anal Bioanal Chem. 402, 2923-2933 (2012).</li><li>Sorensen, C. M., 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=Perturbations+in+the+lipid+profile+of+individuals+with+newly+diagnosed+type+1+diabetes+mellitus:+Lipidomics+analysis+of+a+Diabetes+Antibody+Standardization+Program+sample+subset.">Perturbations in the lipid profile of individuals with newly diagnosed type 1 diabetes mellitus: Lipidomics analysis of a Diabetes Antibody Standardization Program sample subset.</a> Clin Biochem. 43, 948-956 (2010).</li><li>Diamond, D. L., 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=Temporal+proteome+and+lipidome+profiles+reveal+hepatitis+C+virus-associated+reprogramming+of+hepatocellular+metabolism+and+bioenergetics.">Temporal proteome and lipidome profiles reveal hepatitis C virus-associated reprogramming of hepatocellular metabolism and bioenergetics.</a> PLoS Pathog. 6, e1000719 (2010).</li><li>Alquier, T., 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=Deletion+of+GPR40+impairs+glucose-induced+insulin+secretion+in+vivo+in+mice+without+affecting+intracellular+fuel+metabolism+in+islets.">Deletion of GPR40 impairs glucose-induced insulin secretion in vivo in mice without affecting intracellular fuel metabolism in islets.</a> Diabetes. 58, 2607-2615 (2009).</li><li>Ding, J., 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=Application+of+the+accurate+mass+and+time+tag+approach+in+studies+of+the+human+blood+lipidome.">Application of the accurate mass and time tag approach in studies of the human blood lipidome.</a> J Chromatogr B Analyt Technol Biomed Life Sci. 871, 243-252 (2008).</li><li>Rasmussen, A. L., 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=Systems+virology+identifies+a+mitochondrial+fatty+acid+oxidation+enzyme,+dodecenoyl+coenzyme+A+delta+isomerase,+required+for+hepatitis+C+virus+replication+and+likely+pathogenesis.">Systems virology identifies a mitochondrial fatty acid oxidation enzyme, dodecenoyl coenzyme A delta isomerase, required for hepatitis C virus replication and likely pathogenesis.</a> J Virol. 85, 11646-11654 (2011).</li><li>Manza, L. L., Stamer, S. L., Ham, A. J., Codreanu, S. G., Liebler, D. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sample+preparation+and+digestion+for+proteomic+analyses+using+spin+filters.">Sample preparation and digestion for proteomic analyses using spin filters.</a> Proteomics. 5, 1742-1745 (2005).</li><li>Wisniewski, J. R., Zougman, A., Nagaraj, N., Mann, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Universal+sample+preparation+method+for+proteome+analysis.">Universal sample preparation method for proteome analysis.</a> Nat Methods. 6, 359-362 (2009).</li><li>Zhou, J. Y., 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=Simple+sodium+dodecyl+sulfate-assisted+sample+preparation+method+for+LC-MS-based+proteomics+applications.">Simple sodium dodecyl sulfate-assisted sample preparation method for LC-MS-based proteomics applications.</a> Anal Chem. 84, 2862-2867 (2012).</li><li>Anderson, J. C., 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=Decreased+abundance+of+type+III+secretion+system-inducing+signals+in+Arabidopsis+mkp1+enhances+resistance+against+Pseudomonas+syringae.">Decreased abundance of type III secretion system-inducing signals in Arabidopsis mkp1 enhances resistance against Pseudomonas syringae.</a> Proc Natl Acad Sci U S A. 111, 6846-6851 (2014).</li><li>Chourey, K., 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=Direct+cellular+lysis/protein+extraction+protocol+for+soil+metaproteomics.">Direct cellular lysis/protein extraction protocol for soil metaproteomics.</a> J Proteome Res. 9, 6615-6622 (2010).</li><li>Kim, S., Gupta, N., Pevzner, P. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spectral+probabilities+and+generating+functions+of+tandem+mass+spectra:+a+strike+against+decoy+databases.">Spectral probabilities and generating functions of tandem mass spectra: a strike against decoy databases.</a> J Proteome Res. 7, 3354-3363 (2008).</li><li>Kim, S., Pevzner, P. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=MS-GF++makes+progress+towards+a+universal+database+search+tool+for+proteomics.">MS-GF+ makes progress towards a universal database search tool for proteomics.</a> Nat Commun. 5, 5277 (2014).</li><li>Cole, J. K., 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=Phototrophic+biofilm+assembly+in+microbial-mat-derived+unicyanobacterial+consortia:+Model+systems+for+the+study+of+autotroph-heterotroph+interactions.">Phototrophic biofilm assembly in microbial-mat-derived unicyanobacterial consortia: Model systems for the study of autotroph-heterotroph interactions.</a> Front Microbiol. 5, (2014).</li><li>Isaacson, T., 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=Sample+extraction+techniques+for+enhanced+proteomic+analysis+of+plant+tissues.">Sample extraction techniques for enhanced proteomic analysis of plant tissues.</a> Nat Protoc. 1, 769-774 (2006).</li></ol>

그것은 특정 방법, 예를 들어 세포 단계, 적응 시킬 수 있다 그래서 모든 실험실 같은 사용 가능한 장비 해야한다 참고 해야 합니다. 그러나 여기 우리 사용 vortexing sonicating, 큰 50 mL 비드 때리는 사용 하 여 작동 합니다. Lyophilizer-105 ° C의 온도 수집기와 사용할 수 없는 경우 샘플 질소 스트림 아래 건조 수 있습니다. 또한 토양 유형을 크게 다르며 모래, 갯벌, 찰 흙,이 탄, 및 옥 토 (등), 포함 ?...

토양 미생물 지역 사회 평가 탄소 순환, 기후 변화를 이해 하기 위한 중요 한 의미를 갖는다. 그러나 최근 연구는 어려움을, 다양 한 토양 유형 microbiota에 대 한 시퀀스 게놈의 부족 및 감지 하는 단백질의 많은 것의 알 수 없는 기능 등을 강조 했다. 이러한 도전 결과1,,23현재까지 알려진 가장 복잡 한 미생물 지역 사회 토양 때문. 멀티-omic 분석, metagenomic, metatranscriptomic, metaproteomic, metabolomic, 및 lipidomic 연구에서 결과 결합 하는 최근 미생물 존재, 동안에 더 큰 이해를 얻기 위해 수많은 토양 연구에서 구현 되었습니다. 때문에 환경 섭1,,45분자 변화에 대 한 포괄적인 정보를 얻는. 멀티-omic 연구와 함께 한 가지 문제는 질량 분석 (MS)-기반으로 metaproteomic, metabolomic, 및 lipidomic 측정은 일반적으로 각 omic MS 호환6,7 에 대 한 특정 추출 과정 필요 , 8 , 9.이 정확한 절차 확인의 구현을 매우 어렵거나 불가능 한 때 샘플의 한정 된 수량만 사용할 수. 이러한 과제는 우리 동시 대사 산물, 단백질 및 지질 추출 (MPLEx) 방법 작은 샘플 볼륨 또는 대 중을 사용 하 여 정확도 개선 하 고 제공 하는 모든 3 개의 분석에 대 한 빠른 샘플 준비 조사를 자극합니다 10. 데이트 하 고, 이러한 목표를 모두 달성할 수 있는 대체 토양 추출 절차가 필요 하지 않습니다.
글로벌 멀티-omic 단일 토양 샘플의 분석을 사용 하려면 클로 프롬, 메탄올, 물 분판 기반으로 하는 유기 용 매 추출 프로토콜 사용된10이었다. 이 메서드는 원래 총 지질 기사9,11 에 대 한 개발 및 최근에 단일 샘플12,13 에서에서 대사 산물, 단백질과 지질 동시 추출에 대 한 개정 했다 14,15,16,17,18,19,20,,2122, 23,,2425,26,27,28,29,30, 적은 샘플 수량을 사용 하 고 실험적 변화10. MPLEx 프로토콜에서 클로 프롬은 별개 조각으로 샘플 성분의 triphasic 화학 별거를 위한 기초를 제공 하는 물과 혼합할 수 있는. 최고 수성 단계 따라서 친수성 대사, 단백질 디스크 그리고 지질 층 하단 클로 프롬 단계 (그림 1)에서 다음을 포함 합니다. MPLEx 대부분 토양에 적용 될 때 미 립 자 파편 샘플링 튜브의 맨 아래에 축적 하 고 모든 레이어는 수집 된 후 삭제 될 수 있습니다. 그러나 각 토양 유형 다를 수,, 및이 탄 등 높은 유기 토양, 토양 파편 중간 계층에 샘플링 튜브의 바닥에 떨어지지 않습니다. MPLEx 1) 작은 샘플 양이 멀티-omic 분석, 2) 멀티-omic 기사 같은 샘플 감소에서 사용 될 수 있습니다와 같은 동일한 샘플에서 유형 여러 분자를 분리 하면 몇 가지 장점이 제공 전반적인 실험 가변성, 그리고 샘플의 3) 큰 숫자 높은 처리량 연구10훨씬 더 빨리 준비 될 수 있습니다. 함께 이러한 혜택은 토양 샘플 및 그들의 복잡 한 미생물 지역 사회를 평가 하기 위한 더 나은 측정 기능을 제공 하기 위한 중요 합니다.

<table>
	<tbody>
		<tr>
			<td>Chloroform</td>
			<td>Sigma-Aldrich</td>
			<td>650498</td>
			<td>Stored at -20&#176;C !Caution chloroform has acute potential health effects, skin irritation and possible chemical burns, irritation to the respiratory system, may affect the kidneys, liver, heart. Wear suitable protective glasses, clothing and gloves, work in a fume hood.</td>
		</tr>
		<tr>
			<td>Methanol</td>
			<td>Sigma-Aldrich</td>
			<td>34860</td>
			<td>Stored at -20&#176;C !Caution Methanol may cause respiratory tract, skin and eye irritation, may damage the nerves, kidneys and liver. Wear suitable protective glasses, clothing and gloves, work in a fume hood.</td>
		</tr>
		<tr>
			<td>Purified water from Millipore</td>
			<td>Milli-Q</td>
			<td></td>
			<td>Water purification system.</td>
		</tr>
		<tr>
			<td>Sodium dodecyl sulfate</td>
			<td>Sigma-Aldrich</td>
			<td>L6026</td>
			<td>!Caution SDS causes acute toxicity and is flammable. It is a skin, eye and airway irritant. Wear gloves and safety glasses.</td>
		</tr>
		<tr>
			<td>Soil protein extraction kit</td>
			<td>MoBio, NoviPure Soil Protein Extraction Kit, Qiagen</td>
			<td>30000-20</td>
			<td></td>
		</tr>
		<tr>
			<td>DL-dithiothreitol</td>
			<td>Sigma-Aldrich</td>
			<td>43815</td>
			<td></td>
		</tr>
		<tr>
			<td>1M Trizma HCL</td>
			<td>Sigma-Aldrich</td>
			<td>T2694</td>
			<td></td>
		</tr>
		<tr>
			<td>Trichloroacetic acid</td>
			<td>Sigma-Aldrich</td>
			<td>T0699</td>
			<td>!Caution TCA is caustic, toxic and may cause skin burns. Wear gloves and safety glasses.</td>
		</tr>
		<tr>
			<td>Acetone</td>
			<td>Sigma-Aldrich</td>
			<td>650501</td>
			<td>Stored at -20&#176;C !Caution Acetone may cause respiratory tract and skin and eye irritation. Flammable liquid and vapor. Wear safety glasses gloves and a lab coat, work in a fume hood.</td>
		</tr>
		<tr>
			<td>Urea</td>
			<td>Sigma-Aldrich</td>
			<td>208884</td>
			<td>!Caution Urea is an eye and skin irritant, use gloves and safety glasses</td>
		</tr>
		<tr>
			<td>Ammonium bicarbonate</td>
			<td>Fluka</td>
			<td>09830</td>
			<td></td>
		</tr>
		<tr>
			<td>Trypsin</td>
			<td>Promega</td>
			<td>V528A</td>
			<td>20&#181;g vials</td>
		</tr>
		<tr>
			<td>Bicinchoninic acid protein assay kit</td>
			<td>Pierce</td>
			<td>23227</td>
			<td></td>
		</tr>
		<tr>
			<td>Ammonium Formate</td>
			<td>Sigma-Aldrich</td>
			<td>09735</td>
			<td></td>
		</tr>
		<tr>
			<td>Acetonitrile</td>
			<td>Sigma-Aldrich</td>
			<td>34998</td>
			<td>!Caution Acetonitrile is a skin and eye irritant. Highly flammable. Wear gloves and safety glasses. Work in a fume hood.</td>
		</tr>
		<tr>
			<td>Trifluoroacetic acid</td>
			<td>Sigma-Aldrich</td>
			<td>T6508</td>
			<td>!Caution TFA is extremely hazardous in case of skin contact, eye contact, ingestion and inhalation. May produce tissue damage particularly on mucous membranes of eyes, mouth and respiratory tract. Skin contact may produce burns. Wear gloves, lab coat, safety glasses and work in a fume hood.</td>
		</tr>
		<tr>
			<td>Methoxyamine hydrochloride</td>
			<td>Sigma-Aldrich</td>
			<td>226904</td>
			<td>!Caution Methoxyamine hydrochloride causes severe burns and serious damage to eyes, may cause sensitization by skin contact. Wear safety glasses, gloves and lab coat, work in a fume hood.</td>
		</tr>
		<tr>
			<td>Pyridine</td>
			<td>Sigma-Aldrich</td>
			<td>270970</td>
			<td>!Caution Pyridine can cause skin and eye irritation, central nervous system depression. Vapor may cause flash fire. Wear safety glasses, gloves and lab coat, work in a fume hood.</td>
		</tr>
		<tr>
			<td>N-Methyl-N-(trimethylsilyl)trifluoroacetamide with 1% trimethylchlorosilane</td>
			<td>Sigma-Aldrich</td>
			<td>69478</td>
			<td>!Caution MSTFA + 1% TMCS can cause skin corrosion, serious eye damage and specific target organ toxicity. Flammable liquid and vapor. Wear safety glasses, gloves and lab coat, work in a fume hood.</td>
		</tr>
		<tr>
			<td>Potassium chloride</td>
			<td>Sigma-Aldrich</td>
			<td>P9541</td>
			<td></td>
		</tr>
		<tr>
			<td>Milli-Q water purification system</td>
			<td>Millipore</td>
			<td></td>
			<td>model MPGP04001</td>
		</tr>
		<tr>
			<td>Vortex</td>
			<td>Scientific Industries</td>
			<td>SI-0236</td>
			<td>Vortex Genie 2</td>
		</tr>
		<tr>
			<td>Probe sonicator</td>
			<td>FisherBrand</td>
			<td></td>
			<td>model FB505</td>
		</tr>
		<tr>
			<td>Refrigerated centrifuge</td>
			<td>Eppendorf</td>
			<td></td>
			<td>model 5810R</td>
		</tr>
		<tr>
			<td>50mL tube swinging bucket rotor</td>
			<td>Eppendorf</td>
			<td>A-4-44</td>
			<td></td>
		</tr>
		<tr>
			<td>50mL fixed angle rotor</td>
			<td>Eppendorf</td>
			<td>FA-45-6-30</td>
			<td></td>
		</tr>
		<tr>
			<td>Balance</td>
			<td>OHAUS</td>
			<td></td>
			<td>model V22PWE150IT</td>
		</tr>
		<tr>
			<td>Serological pipette controller</td>
			<td>Eppendorf</td>
			<td>12-654-100</td>
			<td></td>
		</tr>
		<tr>
			<td>10mL, 25mL glass serological pipettes</td>
			<td>FisherBrand</td>
			<td>13-678-27F, 13-678-36D</td>
			<td></td>
		</tr>
		<tr>
			<td>Thermomixer with Thermotop</td>
			<td>Eppendorf</td>
			<td>5382000015, 5308000003</td>
			<td></td>
		</tr>
		<tr>
			<td>0.9 &#8211; 2.0 mm blend stainless steel beads</td>
			<td>NextAdvance</td>
			<td>SSB14B</td>
			<td></td>
		</tr>
		<tr>
			<td>0.15 mm garnet beads</td>
			<td>MoBio</td>
			<td>13122-500</td>
			<td></td>
		</tr>
		<tr>
			<td>Magnetic stir plate</td>
			<td>FisherBrand</td>
			<td>11-100-16SH</td>
			<td></td>
		</tr>
		<tr>
			<td>Magnetic stir bar</td>
			<td>FisherBrand</td>
			<td>14512130</td>
			<td></td>
		</tr>
		<tr>
			<td>pH paper strips, pH range 0&#8211;14</td>
			<td>FisherBrand</td>
			<td>M95903</td>
			<td></td>
		</tr>
		<tr>
			<td>15mL, 50mL conical polypropylene centrifuge tube</td>
			<td>Genesee Scientific</td>
			<td>21-103 21-108</td>
			<td>chloroform compatible</td>
		</tr>
		<tr>
			<td>50mL vortex attachment</td>
			<td>MoBio</td>
			<td>13000-V1-50</td>
			<td></td>
		</tr>
		<tr>
			<td>Ice bucket</td>
			<td>FisherBrand</td>
			<td>02-591-44</td>
			<td></td>
		</tr>
		<tr>
			<td>27.25x70mm glass vials</td>
			<td>FisherBrand</td>
			<td>03-339-22K</td>
			<td></td>
		</tr>
		<tr>
			<td>Breathe Easier plate membranes</td>
			<td>Midwest Scientific</td>
			<td>BERM-2000</td>
			<td></td>
		</tr>
		<tr>
			<td>Alcohol wipes</td>
			<td>Diversified Biotech</td>
			<td>BPWP-1000</td>
			<td></td>
		</tr>
		<tr>
			<td>Heater shaker incubator</td>
			<td>Benchmark, Incu-Shaker Mini</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Analog rotisserie tube rotator</td>
			<td>SoCal BioMed, LLC</td>
			<td>82422001</td>
			<td></td>
		</tr>
		<tr>
			<td>Filter-Aided-Sample-Prep kit</td>
			<td>FASP; Expedeon</td>
			<td>44250</td>
			<td></td>
		</tr>
		<tr>
			<td>Microplate reader</td>
			<td>Biotek, EPOCH</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>-20 Degree Celsius Freezer</td>
			<td>Fisher</td>
			<td>13986149</td>
			<td></td>
		</tr>
		<tr>
			<td>-80 Degree Celsius Freezer</td>
			<td>Stirling Ultracold</td>
			<td>SU78OUE</td>
			<td></td>
		</tr>
		<tr>
			<td>Q-Exactive ion trap mass spectrometer</td>
			<td>Thermo Scientific</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Agilent 7890A gas chromatograph coupled with a single quadrupole 5975C mass spectrometer</td>
			<td>Agilent Technologies, Inc.</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>LTQ-Orbitrap Velo</td>
			<td>Thermo Scientific</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Waters NanoEquityTM UPLC system</td>
			<td>Millford, MA</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>250mL media bottle</td>
			<td>FisherBrand</td>
			<td>1395-250</td>
			<td></td>
		</tr>
		<tr>
			<td>Waters vial</td>
			<td>Waters</td>
			<td>186002805</td>
			<td></td>
		</tr>
		<tr>
			<td>Glass MS sample vial and inserts</td>
			<td>MicroSolv</td>
			<td>9502S-WCV, 9502S-02ND</td>
			<td></td>
		</tr>
		<tr>
			<td>Glass HPLC vial and snap caps</td>
			<td>MicroSolv</td>
			<td>9512C-0DCV, 9502C-10C-B</td>
			<td></td>
		</tr>
		<tr>
			<td>HPLC 96-well plate</td>
			<td>Agilent</td>
			<td>5042-6454</td>
			<td></td>
		</tr>
		<tr>
			<td>Large glass vial 27.25x70mm</td>
			<td>FisherBrand</td>
			<td>03-339-22K</td>
			<td></td>
		</tr>
		<tr>
			<td>Lyophilizer</td>
			<td>Labconco</td>
			<td>7934021</td>
			<td></td>
		</tr>
		<tr>
			<td>Polished stainless steel flat head spatula</td>
			<td>Spoonula; FisherBrand</td>
			<td>14-375-10</td>
			<td></td>
		</tr>
		<tr>
			<td>Kim wipes</td>
			<td>Kimberly-Clark</td>
			<td>34721</td>
			<td></td>
		</tr>
		<tr>
			<td>XBridge C18, 250x4.6 mm, 5 &#956;M with 4.6x20 mm guard column</td>
			<td>Waters</td>
			<td>186003117, 186003064</td>
			<td></td>
		</tr>
		<tr>
			<td>Agilent 1100 series HPLC system</td>
			<td>Agilent Technologies</td>
			<td>G1380-90000</td>
			<td></td>
		</tr>
		<tr>
			<td>1.7mL centrifuge tube</td>
			<td>Sorenson</td>
			<td>11700</td>
			<td></td>
		</tr>
		<tr>
			<td>Hamilton Glass Syringes, 5mL, 50&#181;L and 250&#181;L</td>
			<td>Hamilton</td>
			<td>81517, 80975, 81175</td>
			<td></td>
		</tr>
		<tr>
			<td>Pasteur Pipettes</td>
			<td>FisherBrand</td>
			<td>13-678-20A</td>
			<td></td>
		</tr>
		<tr>
			<td>Pasteur Pipette Bulbs</td>
			<td>Sigma-Aldrich</td>
			<td>Z111597</td>
			<td></td>
		</tr>
		<tr>
			<td>Bath Sonicator</td>
			<td>Branson 1800 Ultrasonic Cleaner</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Vacuum Centrifuge</td>
			<td colspan="2">Labconco Centrivap Acid-Resistant Concentrator System</td>
			<td></td>
		</tr>
		<tr>
			<td>MicroSpin Columns, C18 Silica</td>
			<td>The Nest Group</td>
			<td>SEM SS18V</td>
			<td></td>
		</tr>
	</tbody>
</table>

the mplex protocol for multi-omic analyses of soil samples

질량 분석 (MS)-기반된 통합된 metaproteomic, metabolomic, 및 lipidomic (멀티-omic) 연구 우리의 능력을 이해 하 고 환경 및 생물 학적 시스템에서 미생물 지역 사회 특성을 변형 시키고 있다. 이러한 측정도 가장 복잡 한 토양 미생물 커뮤니티의 향상 된 분석 사용 하 고 있습니다 현재까지 알려진 복잡 한 미생물 시스템. 그러나 멀티-omic 분석,, 있다 샘플 준비 과제, 별도 기사는 일반적으로 매우 준비 시간과 필요한 샘플의 양을 증폭 각 omic 연구를 위해 필요 하기 때문. 이 한계를 해결 하기 위해 3-에서-1 방법 같은 토양 샘플에서 대사, 단백질, 지질 (MPLEx)의 동시 추출 용 매 기반 접근을 적응 하 여 만들었습니다. 이 MPLEx 프로토콜은 모두 간단 하 고 많은 샘플 유형에 대 한 강력한 복잡 한 토양 샘플의 한정 된 수량에 대 한 활용 하는 경우에 입증 되었습니다. MPLEx 메서드는 또한 크게 생물학과 환경 물결 따라 변화를 평가 하는 동안 각 미생물 커뮤니티의 구성원의 더 나은 이해를 얻기 위하여 필요한 급속 한 멀티-omic 측정을 사용할 수 있습니다.

MPLEx 프로토콜은 캔자스 네이티브 대초원 토양 (Mollisol 토양)에서 분자를 추출 하는 데 사용 됩니다, triplicate 분석 3376 펩 티 드, 105 지질 및 102 극 대사 (모든 고유 식별)에 대 한 결과 제공. 반면 MPLEx 프로토콜 지질과 대사 산물12,13,14,15,,16<sup cl...

Watch this Scientific Journal Video about The MPLEx Protocol for Multi-omic Analyses of Soil Samples at JoVE.com

The MPLEx Protocol for Multi-omic Analyses of Soil Samples

프로토콜은 동시에 하나의 토양 샘플에서 대사 산물, 단백질과 지질을 추출, 감소 샘플 준비 시간 허용 및 한정 된 수량으로 샘플의 멀티-omic 질량 분석 분석을 활성화에 대 한 제공 됩니다.

멀티-omic 토양 샘플의 분석을 위한 MPLEx 프로토콜

Mass spectrometry (MS)-based integrated metaproteomic, metabolomic, and lipidomic (multi-omic) studies are transforming our ability to understand and ...

Research

JoVE Journal

Chemistry

Transport of Surface-modified Carbon Nanotubes through a Soil Column

토양 칼럼을 통해 표면 개질 된 카본 나노 튜브의 운송

Carbon nanotubes (CNTs) are widely manufactured nanoparticles, which are being utilized in a number of consumer products, such as sporting goods, ...

연구

화학

A Direct, Early Stage Guanidinylation Protocol for the Synthesis of Complex Aminoguanidine-containing Natural Products

복잡한 아미노 구아니딘 함유 천연 제품의 합성에 대한 직접, 초기 단계 Guanidinylation 프로토콜

The guanidine functional group, displayed most prominently in the amino acid arginine, one of the fundamental building blocks of life, is an important ...

Protocol for the Synthesis of Ortho-trifluoromethoxylated Aniline Derivatives

Protocol for the Synthesis of Ortho-trifluoromethoxylated Aniline Derivatives

의 합성을위한 프로토콜<em&gt; 직교</em&gt; -trifluoromethoxylated 아닐린 파생 상품

Molecules bearing trifluoromethoxy (OCF3) group often show desired pharmacological and biological properties. However, facile synthesis of ...

A Protocol for the Production of Gliadin-cyanoacrylate Nanoparticles for Hydrophilic Coating

친수성 코팅 글 리아 딘 - 시아 노 아크릴 레이트 나노 입자의 생산을위한 프로토콜

This article presents a protocol for the production of protein-based nanoparticles that changes the hydrophobic surface to hydrophilic by a simple spray ...

A Simple and Efficient Protocol for the Catalytic Insertion Polymerization of Functional Norbornenes

기능 노보 넨의 촉매 삽입 중합을위한 간단하고 효율적인 프로토콜

Norbornene can be polymerized by a variety of mechanisms, including insertion polymerization whereby the double bond is polymerized and the bicyclic ...

A Protocol for Safe Lithiation Reactions Using Organolithium Reagents

유기 리튬 시약을 사용하여 안전 리튬 화의 반응을위한 프로토콜

Organolithium reagents are powerful tools in the synthetic chemist&#39;s toolbox. However, the extreme pyrophoric nature of the most reactive reagents ...

An Optimized Protocol for the Efficient Radiolabeling of Gold Nanoparticles by Using a 125I-labeled Azide Prosthetic Group

An Optimized Protocol for the Efficient Radiolabeling of Gold Nanoparticles by Using a 125I-labeled Azide Prosthetic Group

를 사용하여 금 나노 입자의 효율적인 방사성 표지를위한 최적화 된 프로토콜<sup&gt; (125)</sup&gt; I-표시 아 지드 보철 그룹

Here, we demonstrate a detailed protocol for the radiosynthesis of a 125I-labeled azide prosthetic group and its application to the efficient ...

Facile Protocol for the Synthesis of Self-assembling Polyamine-based Peptide Amphiphiles (PPAs) and Related Biomaterials

Facile Protocol for the Synthesis of Self-assembling Polyamine-based Peptide Amphiphiles PPAs and Related Biomaterials

Polyamine 기반 펩타이드 Amphiphiles (PPAs) 및 관련된 생체 자기 조립의 합성에 대 한 손쉬운 프로토콜

Polyamine-based Peptide Amphiphiles (PPAs) are a new class of self-assembling amphiphilic biomaterials-related to the peptide amphiphiles (PAs). ...

Automation of a Positron-emission Tomography (PET) Radiotracer Synthesis Protocol for Clinical Production

Automation of a Positron-emission Tomography PET Radiotracer Synthesis Protocol for Clinical Production

양전자 방출 단층 촬영 (PET) Radiotracer 합성 프로토콜의 임상 생산을 위한 자동화

The development of new positron-emission tomography (PET) tracers is enabling researchers and clinicians to image an increasingly wide array of biological ...

Biological Samples Preparation for Speciation at Cryogenic Temperature using High-Resolution  X-Ray Absorption Spectroscopy

고분해능 X선 흡수 분광법을 이용한 극저온에서의 종분화를 위한 생물학적 시료 준비

The study of elements with X-ray absorption spectroscopy (XAS) is of particular interest when studying the role of metals in biological systems. Sample ...