Name: removal of exogenous materials from the outer portion of frozen cores to investigate the ancient biological communities harbored inside
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This work was funded through the U.S. Army Engineer Research and Development Center, Basic Research Program Office. Permission for publishing this information has been granted by the Chief of Engineers.

1. Preparação Equipamentos e Permafrost Coleção Núcleo <ol><li> Preparação de equipamento e coleta de amostra de campo e equipamentos de preservação <ol><li> Montar trado para coleta de amostras, inserindo o adaptador de unidade no topo do barril e girando a alavanca para travá-lo no lugar. Pin o tubo de adaptador para o adaptador de unidade e fixe o motor no tubo adaptador. Insira os cortadores no barril. </li><li> Usar ternos de luz dever, luvas de borracha nitrílica e máscaras para reduzir qualquer cont...

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	<li>Solomon, 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=.">.</a> Climate Change 2007: The Physical Science Basis. , (2007).</li><li>Marchenko, S., Romanovsky, V., Tipenko, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Numerical+Modeling+of+Spatial+Permafrost+Dynamics+in+Alaska.">Numerical Modeling of Spatial Permafrost Dynamics in Alaska.</a> Proc. Ninth Int. Conferen. Permafr. 29, 1125-1130 (2008).</li><li>Pachauri, R. K., Meyer, L. 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> Climate Change 2014: Synthesis Report. Contributions of Working Groups I, II, and III to the Fifth Assessment Report of the Intergovernmental Panel on Climate Change. , (2007).</li><li>Osterkamp, T. E., Romanovsky, V. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evidence+for+warming+and+thawing+of+discontinuous+permafrost+in+Alaska.">Evidence for warming and thawing of discontinuous permafrost in Alaska.</a> Permafr. Periglac. Process. 10 (1), 17-37 (1999).</li><li>Wolken, J. 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=Evidence+and+implications+of+recent+and+projected+climate+change+in+Alaska's+forest+ecosystems.">Evidence and implications of recent and projected climate change in Alaska's forest ecosystems.</a> Ecosphere. 2 (11), 1-35 (2011).</li><li>Hinzman, L. D., Kane, D. L., Gieck, R. E., Everett, K. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hydrologic+and+thermal+properties+of+the+active+layer+in+the+Alaskan+Arctic.">Hydrologic and thermal properties of the active layer in the Alaskan Arctic.</a> Cold Reg. Sci. Technol. 19 (2), 95-110 (1991).</li><li>Hinzman, L. D., Goering, D. J., Kane, D. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+distributed+thermal+model+for+calculating+temperature+profiles+and+depth+of+thaw+in+permafrost+regions.">A distributed thermal model for calculating temperature profiles and depth of thaw in permafrost regions.</a> J. Geophys. Res.: Atmos. 103 (D22), 28975-28991 (1998).</li><li>Osterkamp, T. E., Jorgenson, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Warming+of+Permafrost+in+the+Arctic+National+Wildlife+Refuge.">Warming of Permafrost in the Arctic National Wildlife Refuge.</a> Alaska. Permafr. Periglac. Process. 17, 65-69 (2006).</li><li>Petrone, K. C., Jones, J. B., Hinzman, L. D., Boone, R. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Seasonal+export+of+carbon,+nitrogen,+and+major+solutes+from+Alaskan+catchments+with+discontinuous+permafrost.">Seasonal export of carbon, nitrogen, and major solutes from Alaskan catchments with discontinuous permafrost.</a> J. Geophys. Res. 111, G02020 (2006).</li><li>Guo, L., Ping, C. -. L., Macdonald, R. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mobilization+pathways+of+organic+carbon+from+permafrost+to+arctic+rivers+in+a+changing+climate.">Mobilization pathways of organic carbon from permafrost to arctic rivers in a changing climate.</a> Geophys. Res. Lett. 34 (13), L13603 (2007).</li><li>Katayama, 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=Phylogenetic+analysis+of+bacteria+preserved+in+a+permafrost+ice+wedge+for+25,000+years.">Phylogenetic analysis of bacteria preserved in a permafrost ice wedge for 25,000 years.</a> Appl. Environ. Microbiol. 73 (7), 2360-2363 (2007).</li><li>Katayama, 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=Glaciibacter+superstes+gen.+nov.,+sp.+nov.,+a+novel+member+of+the+family+Microbacteriaceae+isolated+from+a+permafrost+ice+wedge.">Glaciibacter superstes gen. nov., sp. nov., a novel member of the family Microbacteriaceae isolated from a permafrost ice wedge.</a> Int. J. Syst. Evol. Microbiol. 59, 482-486 (2009).</li><li>Waldrop, M. P., White, R., Douglas, T. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isolation+and+identification+of+cold-adapted+fungi+in+the+Fox+Permafrost+Tunnel,+Alaska.">Isolation and identification of cold-adapted fungi in the Fox Permafrost Tunnel, Alaska.</a> Proc. Ninth Int. Conferen. Permafr. , 1887-1891 (2008).</li><li>Douglas, T. 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=Biogeochemical+and+geocryological+characteristics+of+wedge+and+thermokarst-cave+ice+in+the+CRREL+Permafrost+Tunnel.">Biogeochemical and geocryological characteristics of wedge and thermokarst-cave ice in the CRREL Permafrost Tunnel.</a> Alaska Permafr. Periglac. Process. 21 (2), 120-128 (2011).</li><li>Willerslev, 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=Diverse+plant+and+animal+genetic+records+from+Holocene+and+Pleistocene+sediments.">Diverse plant and animal genetic records from Holocene and Pleistocene sediments.</a> Science. 300 (5620), 791-795 (2003).</li><li>Bellemain, 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=Fungal+palaeodiversity+revealed+using+high-throughput+metabarcoding+of+ancient+DNA+from+Arctic+permafrost.">Fungal palaeodiversity revealed using high-throughput metabarcoding of ancient DNA from Arctic permafrost.</a> Environ. Microbiol. 15 (4), 1176-1189 (2013).</li><li>Steven, B., Pollard, W. H., Greer, C. W., Whyte, L. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microbial+diversity+and+activity+through+a+permafrost/ground+ice+core+profile+from+the+Canadian+high+Arctic.">Microbial diversity and activity through a permafrost/ground ice core profile from the Canadian high Arctic.</a> Environ. Microbiol. 10 (12), 3388-3403 (2008).</li><li>Lorenzen, E. D., 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=Species-specific+responses+of+Late+Quaternary+megafauna+to+climate+and+humans.">Species-specific responses of Late Quaternary megafauna to climate and humans.</a> Nature. 479 (7373), 359-364 (2011).</li><li>Wilhelm, R. C., Radtke, K., Mykytczuk, N. C. S., Greer, C. W., Whyte, L. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Life+at+the+wedge:+The+activity+and+diversity+of+Arctic+ice+wedge+microbial+communities.">Life at the wedge: The activity and diversity of Arctic ice wedge microbial communities.</a> Astrobiol. 12 (4), 347-360 (2012).</li><li>Sheriden, P. P., Miteva, V. I., Brenchley, J. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Phylogenetic+analysis+of+anaerobic+psychrophilic+enrichment+cultures+obtained+from+a+Greenland+glacier+ice+core.">Phylogenetic analysis of anaerobic psychrophilic enrichment cultures obtained from a Greenland glacier ice core.</a> Appl. Environ. Microbiol. 69 (4), 2153-2160 (2003).</li><li>Rivkina, 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=Biogeochemistry+of+methane+and+methanogenic+archaea+in+permafrost.">Biogeochemistry of methane and methanogenic archaea in permafrost.</a> FEMS Microbiol. Ecol. 61 (1), 1-15 (2007).</li><li>Juck, D. F., 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=Utilization+of+fluorescent+microspheres+and+a+green+fluorescent+protein-marked+strain+for+assessment+of+microbiological+contamination+of+permafrost+and+ground+ice+core+samples+from+the+Canadian+High+Arctic.">Utilization of fluorescent microspheres and a green fluorescent protein-marked strain for assessment of microbiological contamination of permafrost and ground ice core samples from the Canadian High Arctic.</a> Appl. Environ. Microbiol. 71 (2), 1035-1041 (2005).</li><li>Griffiths, R. I., Whiteley, A. S., O'Donnell, A. G., Bailey, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rapid+method+for+coextraction+of+DNA+and+RNA+from+natural+environments+for+analysis+of+ribosomal+DNA-+and+rRNA-based+microbial+community+composition.">Rapid method for coextraction of DNA and RNA from natural environments for analysis of ribosomal DNA- and rRNA-based microbial community composition.</a> Appl. Environ. Microbiol. 66 (12), 5488-5491 (2000).</li><li>T&#246;we, 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=Improved+protocol+for+the+simultaneous+extraction+and+column-based+separation+of+DNA+and+RNA+from+different+soils.">Improved protocol for the simultaneous extraction and column-based separation of DNA and RNA from different soils.</a> J. Microbiol. Methods. 84 (3), 406-412 (2011).</li><li>Nadkarni, M. A., Martin, F. E., Jacques, N. A., Hunter, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Determination+of+bacterial+load+by+real-time+PCR+using+a+broad+range+(universal)+probe+and+primers+set.">Determination of bacterial load by real-time PCR using a broad range (universal) probe and primers set.</a> Microbiol. 148, 257-266 (2002).</li><li>Takai, K., Horikoshi, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rapid+detection+and+quantification+of+members+of+the+archaeal+community+by+quantitative+PCR+using+fluorogenic+probes.">Rapid detection and quantification of members of the archaeal community by quantitative PCR using fluorogenic probes.</a> Appl. Environ. Microbiol. 66 (11), 5066-5072 (2000).</li><li>Fogel, G. B., Collins, C. R., Brunk, C. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prokaryotic+genome+size+and+SSU+rDNA+copy+number:+Estimation+of+microbial+relative+abundance+from+a+mixed+population.">Prokaryotic genome size and SSU rDNA copy number: Estimation of microbial relative abundance from a mixed population.</a> Microb. Ecol. 38, 93-113 (1999).</li><li>Bodilis, J., Nsigue-Meilo, S., Besaury, L., Quillet, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Variable+copy+number,+intra-genomic+heterogeneities+and+later+transfers+of+the+16S+rRNA+gene+in+Pseudomonas.">Variable copy number, intra-genomic heterogeneities and later transfers of the 16S rRNA gene in Pseudomonas.</a> PLOS One. 7, e35647 (2012).</li><li>Caporaso, J. 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=QIIME+allows+analysis+of+high-throughput+community+sequencing+data.">QIIME allows analysis of high-throughput community sequencing data.</a> Nature Methods. 7, 335-336 (2010).</li><li>Sellmann, P. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Geology of the USA CRREL permafrost tunnel, Fairbanks, Alaska. US Army Cold Reg. Res. Eng. Lab. Technical Rep. 199. , (1967).</li><li>Sellmann, P. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Additional information on the geology and properties of materials exposed in the USA CRREL permafrost tunnel. US Army CRREL Special Rep. , (1972).</li><li>Christner, B. C., Mikucki, J. A., Foreman, C. M., Denson, J., Priscu, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Glacial+ice+cores:+A+model+system+for+developing+extraterrestrial+decontamination+protocols.">Glacial ice cores: A model system for developing extraterrestrial decontamination protocols.</a> Icarus. 174 (2), 572-584 (2005).</li><li>Mackelprang, 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=Metagenomic+analysis+of+a+permafrost+microbial+community+reveals+a+rapid+response+to+thaw.">Metagenomic analysis of a permafrost microbial community reveals a rapid response to thaw.</a> Nature. 480 (7377), 368-371 (2011).</li><li>Champlot, 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=An+efficient+multistrategy+DNA+decontamination+of+PCR+reagents+for+hyper+sensitive+PCR+applications.">An efficient multistrategy DNA decontamination of PCR reagents for hyper sensitive PCR applications.</a> PLoS One. 5 (9), e13042 (2010).</li><li>Yergeau, E., Hogues, H., Whyte, L. G., Greer, C. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+functional+potential+of+high+Arctic+permafrost+revealed+by+metagenomic+sequencing,+qPCR,+and+microarray+analyses.">The functional potential of high Arctic permafrost revealed by metagenomic sequencing, qPCR, and microarray analyses.</a> The ISME J. 4 (9), 1206-1214 (2010).</li><li>Welzl, G., Schloter, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bacterial+community+structure+in+soils+of+the+Tibetan+Plateau+affected+by+discontinuous+permafrost+or+seasonal+freezing.">Bacterial community structure in soils of the Tibetan Plateau affected by discontinuous permafrost or seasonal freezing.</a> Biol. Fertil. Soils. 50 (3), 555-559 (2014).</li><li>Vishnivetskaya, T. 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=Commercial+DNA+extraction+kits+impact+observed+microbial+community+composition+in+permafrost+samples.">Commercial DNA extraction kits impact observed microbial community composition in permafrost samples.</a> FEMS Microbiol. Ecol. 87 (1), 217-230 (2014).</li><li>Wagner, D., Kobabe, S., Liebner, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bacterial+community+structure+and+carbon+turnover+in+permafrost-affected+soils+of+the+Lena+Delta,+northeastern+Siberia.">Bacterial community structure and carbon turnover in permafrost-affected soils of the Lena Delta, northeastern Siberia.</a> Can. J. Microbiol. 55 (1), 73-83 (2009).</li><li>Jiang, N., 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=Characteristic+microbial+communities+in+the+continuous+permafrost+beside+the+bitumen+in+Qinghai-Tibetan+Plateau.">Characteristic microbial communities in the continuous permafrost beside the bitumen in Qinghai-Tibetan Plateau.</a> Environ. Earth Sci. 74, 1343-1352 (2015).</li></ol>

A criosfera oferece acesso a organismos preservados que persistiram sob condições ambientais do passado. Embora a taxa recuperado pode não representar a comunidade histórica completa, aqueles recuperados a partir de análise de amostras de gelo e permafrost glacial pode render valiosas informações históricas sobre seleccionar períodos de tempo 15-16. Por exemplo, a informação biológica significativa foi elaborado a partir de estudos de gelo que investigam atividade anaeróbica no gelo da Groenlând...

A criosfera (por exemplo, solos de permafrost, características gelo, neve glacial, firn e gelo) oferece um vislumbre do que tipos de organismos persistiu em condições ambientais do passado. Uma vez que estes substratos pode ser dezenas a centenas de milhares de anos de idade, suas comunidades microbianas, quando preservado congeladas desde a deposição, refletem as condições ambientais antigos. Para analisar adequadamente estes ecossistemas e extrair informação biológica significativa de solos congelados e gelo, coleta adequada e processamento das amostras congeladas é necessário. Isto é de extrema importância como projeções climáticas para o século 21 indicam o potencial para um aquecimento acentuado em regiões árticas e sub-árticas 1. Especificamente, Interior do Alasca e Groenlândia são esperados para aquecer cerca de 5 ° C e 7 ° C, respectivamente em 2100 2,3. Isto é esperado para afetar significativamente o solo e comunidades microbianas aquáticos, e, portanto, relacionadaprocessos biogeoquímicos. As temperaturas mais quentes e regime de precipitação alterados são esperados para iniciar a degradação do permafrost em muitas áreas 2-5 potencialmente levando a uma mais grossa, sazonalmente descongelado (ativo) camada 6,7, o degelo dos solos congelados, ea fusão de corpos de gelo enormes, tais como gelo no solo, cunhas de gelo, e segregação de gelo 8. Isto mudaria drasticamente os atributos biogeoquímicos além da biodiversidade de plantas e animais nestes ecossistemas. gelo glacial e características de sedimentos permafrost e gelo singenéticos ter aprisionado química e evidência biológica de um ambiente representando o que viveu lá no momento as características formado. Por exemplo, no Interior do Alasca, tanto Illinoisan e Wisconsin permafrost com idades estão presentes e este permafrost em particular fornece os locais únicos que datam de moderno a 150.000 anos antes do presente (YBP) que contêm evidência biológica e geoquímica do impact das mudanças climáticas passadas sobre a biodiversidade. Como resultado, estes sedimentos fornecem um registro da biogeoquímica e da biodiversidade ao longo de muitos milhares de anos. Uma vez que a área tem taxas de sedimentação baixos e nunca foi glaciar, as amostras não perturbadas são acessíveis para recolha e análise, tanto de perfuração verticalmente no perfil do solo ou de perfuração horizontal em túneis. Mais importante ainda, existem extensos registros que, especialmente destacar as características únicas de biogeoquímicos permafrost nessa região 9-14. Especificamente, a aplicação da análise de DNA para estimar presença e extensão da biodiversidade em ambas as amostras de gelo e permafrost existentes e antigas permite a exploração da ligação das condições ambientais antigas e habitat para a ocupação por organismos específicos. Estudos anteriores identificaram impactos climáticos sobre mamíferos, plantas e microorganismos de amostras datando de 50k YBP 11, 15-19, embora cada estudo utilizou uma differemetodologia nt para recolher e descontaminar as permafrost ou gelo núcleos. Em alguns casos, os núcleos de perfuração foram esterilizados 16, 20-21, embora a metodologia específica não esclarecer se os ácidos nucleicos estranhos, foram igualmente eliminadas a partir das amostras. Em outros estudos, 15 isolados bacterianos (por exemplo, Serratia marcescens), bem como as microesferas fluorescentes 22 foram utilizadas para medir a eficácia de processos de descontaminação. Este experimento foi parte de um estudo maior investigando comunidades microbianas a partir de amostras de permafrost que datam YBP cerca de 40k. O objectivo específico desta parte do estudo foi para descontaminar com sucesso de gelo e permafrost núcleos. Para nosso conhecimento, nenhuma metodologia tem integrado o uso de soluções concebidas para eliminar os ácidos nucleicos estranhos e nucleases associados a partir da porção exterior dos núcleos congelados. Isto apesar do fato de que estas soluções são commonly usado para descontaminar equipamentos de laboratório para experimentos moleculares. Uma vez que os núcleos foram descontaminadas, o DNA genómico foi extraído utilizando os protocolos desenvolvidos por Griffiths et al. 23 e TOWE et al. 24, quantificado utilizando um espectrofotómetro, e diluiu-se com água estéril, isenta de ADN para atingir 20 ng por reacção. Bacterianas genes 16S rRNA foram amplificadas com primers 331F e 797R e sonda BacTaq 25 e 16S archaeal genes rRNA foram amplificadas com primers Arch 349F e Arch 806R e a sonda TM Arch 516F 26 sob as seguintes condições: 95 ° C durante 600 segundos seguido por 45 ciclos de 95 ° C durante 30 seg, 57 ° C durante 60 segundos, e 72 ° C durante 25 seg, com extensão final a 40 ° C durante 30 seg. Todas as reacções foram conduzidas qPCR em duplicado. Os volumes de reacção de 20 ul incluídos 20 ng de ADN, 10 pM de iniciadores, 5 uM de sonda, e 10 ul da mistura de reacção qPCR. normas for qPCR bacteriana e archaea foram preparadas usando ADN genómico de Pseudomonas fluorescens e Halobacterium salinarum, respectivamente. Ambos foram cresceu até à fase log. As contagens foram realizadas e o ADN foi isolado a partir das culturas. O ADN genómico foi quantificada com um espectrofotómetro com o pressuposto de um e seis cópias do gene 16S rRNA por genoma de H. salinarum e P. fluorescens, respectivamente 27-28. número de cópias dos genes de bactérias e archaea foram calculados com base na curva padrão, log-transformada em conta as especificidades desiguais entre tratamentos, e avaliados por análise de variância. Composição da comunidade foi determinada por sequenciamento do gene 16S rRNA usando células de fluxo e tecnologias de amplificação ponte e analisar as comunidades com insights quantitativos em ecologia microbiana &quot;(QIIME) 29. Para a frente e reverso lê foram unidas e depois sequências foram filtrados, indexado,e representantes de alta qualidade foram selecionados para unidades taxonômicas operacionais (OTU) cessão de novo através de um alinhamento de sequência com uma base de dados de referência. sequências alinhadas foram comparados com um banco de dados de referência separado para atribuição taxonômica. Uma tabela OTU nível filo foi criado para determinar a composição da comunidade em geral.

<table border="0" cellpadding="0" cellspacing="0" width="1084">
	<tbody>
		<tr height="42">
			<td height="42" style="height:42px;width:439px;">Auger</td>
			<td style="width:300px;">Snow, Ice, and Permafrost Research Establishment (SIPRE), Fairbanks, AK</td>
			<td style="width:161px;">N/A</td>
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		<tr height="21">
			<td height="21" style="height:21px;">70% Isopropanol</td>
			<td style="width:300px;">Walmart</td>
			<td>551116880</td>
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			<td height="21" style="height:21px;">95% Ethyl Alcohol (denatured)&#160;</td>
			<td style="width:300px;">Fisher Scientific, Pittsburgh, PA</td>
			<td style="width:161px;">A407-4</td>
			<td></td>
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			<td height="21" style="height:21px;">DNA decontamination solution, DNA Away</td>
			<td>Molecular Bio-Products, Inc., San Diego, CA</td>
			<td style="width:161px;">7010</td>
			<td></td>
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			<td height="21" style="height:21px;">RNase decontamination solution, RNase Away</td>
			<td>Molecular Bio-Products, Inc., San Diego, CA&#160;</td>
			<td style="width:161px;">7002</td>
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			<td style="width:300px;">Kimberly-Clark Professional, Roswell, GA</td>
			<td style="width:161px;">10606</td>
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			<td height="21" style="height:21px;">Nitrile Gloves</td>
			<td style="width:300px;">Fisher Scientific, Pittsburgh, PA</td>
			<td style="width:161px;">FFS-700</td>
			<td></td>
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			<td height="21" style="height:21px;">Antiviral Masks</td>
			<td style="width:300px;">Curad, Walgreens</td>
			<td style="width:161px;">CUR3845</td>
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			<td height="21" style="height:21px;">Sterile Sample Bags&#160;</td>
			<td>Nasco, Fort Atkinson, WI</td>
			<td style="width:161px;">B01445</td>
			<td></td>
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			<td height="21" style="height:21px;">Steel Microtome Blade&#160;</td>
			<td style="width:300px;">B-Sharp Microknife, Wake Forest, NC</td>
			<td style="width:161px;">N/A</td>
			<td></td>
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			<td height="21" style="height:21px;">Metal Rack</td>
			<td style="width:300px;">Fabricated at CRREL, Hanover, NH</td>
			<td style="width:161px;">N/A</td>
			<td></td>
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			<td height="21" style="height:21px;width:439px;">Tray</td>
			<td style="width:300px;">Handy Paint Products, Chanhassen, MN</td>
			<td style="width:161px;">7500-CC</td>
			<td></td>
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			<td height="21" style="height:21px;width:439px;">Aluminum Foil</td>
			<td style="width:300px;">Western Plastics, Temecula, CA</td>
			<td style="width:161px;">N/A</td>
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			<td height="21" style="height:21px;width:439px;">500 ml Bottle with 0.22 &#956;m Filter</td>
			<td style="width:300px;">Corning, Corning, NY</td>
			<td style="width:161px;">430513</td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;">Serratia marcescens&#160;</td>
			<td style="width:300px;">ATCC, Manassas, VA</td>
			<td style="width:161px;">17991</td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Biosafety Hood</td>
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			<td style="width:161px;">NU-425-400</td>
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			<td height="21" style="height:21px;width:439px;">Petri Dish</td>
			<td style="width:300px;">Fisher Scientific, Pittsburgh, PA</td>
			<td style="width:161px;">FB0875712</td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;">ATCC Agar 181- Tryptone</td>
			<td style="width:300px;">Acros Organics, NJ</td>
			<td style="width:161px;">61184-5000</td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;">ATCC Agar 181- Glucose</td>
			<td style="width:300px;">Fisher Scientific, Pittsburgh, PA</td>
			<td style="width:161px;">BP381-500</td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;">ATCC Agar 181- Yeast Extract</td>
			<td style="width:300px;">Fisher Scientific, Pittsburgh, PA</td>
			<td style="width:161px;">BP1422-500</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">ATCC Agar 181- Dipotassium Phosphate</td>
			<td style="width:300px;">JT Baker, Phillipsburg, NJ</td>
			<td style="width:161px;">3252-01</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">ATCC Agar 181- Agar</td>
			<td style="width:300px;">Difco, Sparks, MD</td>
			<td style="width:161px;">214530</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">NanoDrop 2000 UV Vis Spectrophotometer</td>
			<td style="width:300px;">Thermo Fisher Scientific, Wilmington, DE</td>
			<td style="width:161px;"></td>
			<td></td>
		</tr>
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			<td height="42" style="height:42px;width:439px;">Lightcycler 480 System</td>
			<td style="width:300px;">Roche Molecular Systems, Inc., Indianapolis, IN</td>
			<td style="width:161px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:439px;">Halobacterium salinarum</td>
			<td style="width:300px;">American Type Culture Collection (ATCC), Manassas, VA</td>
			<td style="width:161px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:439px;">Pseudomonas fluorescens&#160;</td>
			<td style="width:300px;">American Type Culture Collection (ATCC), Manassas, VA</td>
			<td style="width:161px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Microbial DNA Isolation Kit</td>
			<td style="width:300px;">MoBio Laboratories, Carlsbad, CA</td>
			<td style="width:161px;">12224-50</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Ear Protection</td>
			<td style="width:300px;">Elvex</td>
			<td style="width:161px;">EP-201</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Hard Hat</td>
			<td style="width:300px;">N/A</td>
			<td style="width:161px;">N/A</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Kimwipes</td>
			<td style="width:300px;">Kimberly-Clark Professional, Roswell, GA</td>
			<td style="width:161px;">34705</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Glass Wool</td>
			<td style="width:300px;">Pyrex</td>
			<td style="width:161px;">430330</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Ruler</td>
			<td style="width:300px;">N/A</td>
			<td style="width:161px;">N/A</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Weighing Tin&#160;</td>
			<td style="width:300px;">Fisher Scientific, Pittsburgh, PA</td>
			<td style="width:161px;">08-732-100</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Sodium chloride</td>
			<td style="width:300px;">Sigma Aldrich, St Louis, MO</td>
			<td style="width:161px;">S-9625</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Potassium chloride</td>
			<td style="width:300px;">JT Baker, Phillipsburg, NJ</td>
			<td style="width:161px;">3040-04</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Potassium phosphate, monobasic</td>
			<td style="width:300px;">JT Baker, Phillipsburg, NJ</td>
			<td style="width:161px;">&#160;3246-01</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Potassium phosphate, dibasic</td>
			<td style="width:300px;">JT Baker, Phillipsburg, NJ</td>
			<td>3252-01</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Sodium phosphate dibasic, anhydrous</td>
			<td style="width:300px;">Fisher Scientific, Pittsburgh, PA</td>
			<td style="width:161px;">BP332-500</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">50 ml Centrifuge Tubes</td>
			<td style="width:300px;">Corning, Corning, NY</td>
			<td style="width:161px;">4558</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">2 ml Microcentrifuge Tubes</td>
			<td style="width:300px;">MoBio Laboratories, Carlsbad, CA</td>
			<td style="width:161px;">1200-250-T</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">2 ml Ceramic Bead Tubes (1.4 mm)</td>
			<td style="width:300px;">MoBio Laboratories, Carlsbad, CA</td>
			<td style="width:161px;">13113-50</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Scoopula</td>
			<td style="width:300px;">Thermo Fisher Scientific, Wilmington, DE</td>
			<td style="width:161px;">1437520</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Balance</td>
			<td style="width:300px;">Ohaus, Parsippany, NJ</td>
			<td style="width:161px;">E12130</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Diethylpyrocarbonate (DEPC)</td>
			<td style="width:300px;">Sigma Aldrich, St Louis, MO</td>
			<td style="width:161px;">D5758</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Hexadecyltrimethylammoniabromide (CTAB)&#160;</td>
			<td style="width:300px;">Acros Organics, NJ</td>
			<td style="width:161px;">22716-5000</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Polyethylene glycol 8000&#160;</td>
			<td style="width:300px;">Sigma Aldrich, St Louis, MO</td>
			<td style="width:161px;">P5413-1KG</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Phenol-chloroform-isoamyl alcohol (25:24:1) (pH 8)&#160;</td>
			<td style="width:300px;">Fisher Scientific, Pittsburgh, PA</td>
			<td style="width:161px;">BP1752-400</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Centrifuge</td>
			<td style="width:300px;">Eppendorf, Hauppauge, NY</td>
			<td style="width:161px;">5417R</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Chloroform-isoamyl alcohol (24:1)</td>
			<td style="width:300px;">Sigma Aldrich, St Louis, MO</td>
			<td style="width:161px;">C0549-1PT</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">TE Buffer</td>
			<td style="width:300px;">Ambion (Thermo Fisher), Wilmington, DE</td>
			<td style="width:161px;">AM9860</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:439px;">Pipets</td>
			<td style="width:300px;">Rainin, Woburn, MA</td>
			<td style="width:161px;">Pipet Lite XLS, 2, 10, 200, 1nd 1000ul pipets</td>
			<td></td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:439px;">Pipet tips</td>
			<td style="width:300px;">Rainin, Woburn, MA</td>
			<td style="width:161px;">Rainin LTS presterilized, low retention, filtered tips, 10, 20, 200, 1000ul</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Vortexor</td>
			<td style="width:300px;">Scientific Industries, Bohemia, NY</td>
			<td style="width:161px;">G-560</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Vortex Adaptor</td>
			<td style="width:300px;">MoBio Laboratories, Carlsbad, CA</td>
			<td style="width:161px;">13000-V1</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Clear Bottle</td>
			<td style="width:300px;">Corning, Corning, NY</td>
			<td style="width:161px;">C1395500</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Amber Bottle</td>
			<td style="width:300px;">Corning, Corning, NY</td>
			<td style="width:161px;">C5135250</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Bottle Top Filters, 0.22um</td>
			<td style="width:300px;">Corning, Corning, NY</td>
			<td style="width:161px;">430513</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:439px;">60 mL Syringe</td>
			<td style="width:300px;">Becton, Dickenson and Company, Franklin Lakes, NJ</td>
			<td style="width:161px;">BD 309653</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Millex Syringe filters, 0.22 &#956;m</td>
			<td style="width:300px;">EMD Millipore, Billerica, MA</td>
			<td style="width:161px;">SLGV033RB</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">70% Ethanol</td>
			<td style="width:300px;">Fisher Scientific, Pittsburgh, PA</td>
			<td style="width:161px;">BP2818-500</td>
			<td>diluted &#38; filter sterilized</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Isotemp 100 L Oven</td>
			<td style="width:300px;">Fisher Scientific, Pittsburgh, PA</td>
			<td style="width:161px;">151030511</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Cell Spreader</td>
			<td style="width:300px;">Fisher Scientific, Pittsburgh, PA</td>
			<td style="width:161px;">08-100-10</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:439px;">Disposable Inoculating Loops</td>
			<td style="width:300px;">Fisher Scientific, Pittsburgh, PA</td>
			<td style="width:161px;">22-363-602</td>
			<td></td>
		</tr>
	</tbody>
</table>

removal of exogenous materials from the outer portion of frozen cores to investigate the ancient biological communities harbored inside

The cryosphere offers access to preserved organisms that persisted under past environmental conditions. In fact, these frozen materials could reflect conditions over vast time periods and investigation of biological materials harbored inside could provide insight of ancient environments. To appropriately analyze these ecosystems and extract meaningful biological information from frozen soils and ice, proper collection and processing of the frozen samples is necessary. This is especially critical for microbial and DNA analyses since the communities present may be so uniquely different from modern ones. Here, a protocol is presented to successfully collect and decontaminate frozen cores. Both the absence of the colonies used to dope the outer surface and exogenous DNA suggest that we successfully decontaminated the frozen cores and that the microorganisms detected were from the material, rather than contamination from drilling or processing the cores.

O método apresentado poderia ser utilizado para descontaminar amostras congeladas recolhidas de vários ambientes cryosphere, a partir de gelo glacial para permafrost. Aqui, apresentamos os dados especificamente recolhidos a partir de amostras de gelo e permafrost coletados a partir da Pesquisa de Engenharia e Centro de Desenvolvimento - Cold Regiões Laboratório de Pesquisa e Engenharia (ERDC-CRREL) Permafrost túnel localizado na Fox, AK (Figura 1A e 1B).</st...

Watch this Scientific Journal Video about Removal of Exogenous Materials from the Outer Portion of Frozen Cores to Investigate the Ancient Biological Communities Harbored Inside at JoVE.com

Removal of Exogenous Materials from the Outer Portion of Frozen Cores to Investigate the Ancient Biological Communities Harbored Inside

A criosfera oferece acesso a organismos preservados que persistiram sob condições ambientais do passado. Um protocolo é apresentada para recolher e descontaminar núcleos permafrost de solos e gelo. A ausência de colónias exógenas e DNA sugerem que os microrganismos detectados representam o material, em vez de contaminação de perfuração ou de processamento.

Remoção de materiais exógenos da parte exterior dos núcleos congelados para investigar a antiga comunidades biológicas abrigava Dentro

The cryosphere offers access to preserved organisms that persisted under past environmental conditions. In fact, these frozen materials could reflect ...

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Medindo nitrito e nitrato, metabolitos na Óxido Nítrico Pathway, em materiais biológicos utilizando o método de quimioluminescência

Nitric oxide (NO) is one of the main regulator molecules in vascular homeostasis and also a neurotransmitter. Enzymatically produced NO is oxidized into ...

Volume Segmentation and Analysis of Biological Materials Using SuRVoS (Super-region Volume Segmentation) Workbench

Volume Segmentation and Analysis of Biological Materials Using SuRVoS Super-region Volume Segmentation Workbench

Volume de segmentação e análise de materiais biológicos, usando a bancada de trabalho de SuRVoS (região super Volume segmentação)

Segmentation is the process of isolating specific regions or objects within an imaged volume, so that further study can be undertaken on these areas of ...

Experimental Protocol for Using Drosophila As an Invertebrate Model System for Toxicity Testing in the Laboratory

Experimental Protocol for Using Drosophila As an Invertebrate Model System for Toxicity Testing in the Laboratory

Protocolo experimental para o uso de drosófila como um sistema modelo invertebrados para testes de toxicidade em laboratório

Emergent properties and external factors (population-level and ecosystem-level interactions, in particular) play important roles in mediating ...