Name: establishment of microbial eukaryotic enrichment cultures from a chemically stratified antarctic lake and assessment of carbon fixation potential
Uploaded: 2012-04-20T13:00:00
Description: Watch this Scientific Journal Video about Establishment of Microbial Eukaryotic Enrichment Cultures from a Chemically Stratified Antarctic Lake and Assessment of Carbon Fixation Potential at JoVE.com

作者感谢J. Priscu，A. Chiuchiolo和麦克默多LTER网络湖沼队在南极样品的收集和保全的协助。我们感谢Ratheon极地后勤支援服务和PHI的直升机。光显微镜在迈阿密的先进的显微镜和影像中心为中心的产生。这项工作是由国家科学基金会极地项目办公室资助0631659和1056396的支持。

1。样品采集<ol><li>选择和准备采样点有一天前采样水柱。这将使水柱分层层扰动后，由于钻井和冰孔熔化改革。识别GPS演练现场的位置。 </li><li>要访问的水柱，首先通过一个瞬间冰到了4英寸的jiffy的飞行延伸和钻头的螺旋冰钻了一个洞。为了防止冻结孔钻，尽量避免停止钻探约4-6英寸以上水柱顶部钻成液态水。 </li><li>钻孔将需要从15至60厘米，直径扩大采样前。这是通过使用一个洞熔炉（Hotsy?...

<ol>
	<li>Morgan-Kiss, R. M., Priscu, J. P., Pocock, T., Gudynaite-Savitch, L., H&#252;ner, N. 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=Adaptation+and+acclimation+of+photosynthetic+microorganisms+to+permanently+cold+environments.">Adaptation and acclimation of photosynthetic microorganisms to permanently cold environments.</a> Microbiol Mol. Biol. Rev. 70, 222-252 (2006).</li><li>Priscu, J. C., Wolf, C. F., Takacs, C. D., Fritsen, C. H., Laybourn-Parry, J., Roberts, J. K. M., Berry-Lyons, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Carbon+transformations+in+the+water+column+of+a+perennially+ice-covered+Antarctic+Lake.">Carbon transformations in the water column of a perennially ice-covered Antarctic Lake.</a> Biosci. 49, 997-1008 (1999).</li><li>Caron, D. A., Worden, A. Z., Countway, P. D., Demir, E., Heidelberg, K. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Protists+are+microbes+too:+a+perspective.">Protists are microbes too: a perspective.</a> ISME J. 3, 4-12 (2009).</li><li>Bielewicz, S., Bell, E. M., Kong, W., Friedberg, I., Priscu, J. C., Morgan-Kiss, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Protist+diversity+in+a+permanently+ice-covered+Antarctic+lake+during+the+polar+night+transition.">Protist diversity in a permanently ice-covered Antarctic lake during the polar night transition.</a> ISME J. 5, 1559-1564 (2011).</li><li>Laybourn-Parry, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=No+place+too+cold.">No place too cold.</a> Science. 324, 1521-1522 (2009).</li><li>Roberts, E. C., Laybourn-Parry, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mixotrophic+cryptophytes+and+their+predators+in+the+Dry+Valley+lakes+of+Antarctica.">Mixotrophic cryptophytes and their predators in the Dry Valley lakes of Antarctica.</a> Freshwat. Biol. 41, 737-746 (1999).</li><li>Roberts, E. C., Priscu, J. C., Laybourn-Parry, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microplankton+dynamics+in+a+perennially+ice-covered+Antarctic+lake-Lake+Hoare.">Microplankton dynamics in a perennially ice-covered Antarctic lake-Lake Hoare.</a> Freshwat Biol. 27, 238-249 (2004).</li><li>Bell, E. M., Laybourn-Parry, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mixotrophy+in+the+Antarctic+phytoflagellate+Pyramimonas+gelidicola.">Mixotrophy in the Antarctic phytoflagellate Pyramimonas gelidicola.</a> J. Phycol. 39, 644-649 (2003).</li><li>De Wever, A., Leliaert, F., Verleyen, E., Vanormelingen, P., Van der Gucht, K., Hodgson, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hidden+levels+of+phylodiversity+in+Antarctic+green+algae:+further+evidence+for+the+existence+of+glacial+refugia.">Hidden levels of phylodiversity in Antarctic green algae: further evidence for the existence of glacial refugia.</a> Proc. Biol. Sci. , 276-3591 (2009).</li><li>Laybourn-Parry, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Survival+mechanisms+in+Antarctic+lakes.">Survival mechanisms in Antarctic lakes.</a> Philos. Trans. R. Soc. Lond. B. Biol. Sci. 357, 863-869 (2002).</li><li>Lizotte, M. P., Priscu, J. C., 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=Distribution,+succession+and+fate+of+phytoplankton+in+the+dry+valley+lakes+of+Antarctica,+based+on+pigment+analysis.">Distribution, succession and fate of phytoplankton in the dry valley lakes of Antarctica, based on pigment analysis.</a> Ecosystem Dynamics in a Polar Desert: The McMurdo Dry Valleys. , 229-240 (1998).</li><li>Ellis, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Most+abundance+protein+in+the+world.">Most abundance protein in the world.</a> Tren. Biochem. Sci. 4, 241-244 (1979).</li><li>Tortell, P. D., Martin, C. L., Corkum, M. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Inorganic+carbon+uptake+and+intracellular+assimilation+by+subarctic+Pacific+phytoplankton+assemblages.">Inorganic carbon uptake and intracellular assimilation by subarctic Pacific phytoplankton assemblages.</a> Limnol. Oceanogr. 51, 2102-2110 (2006).</li><li>Jeffrey, S. W., Humphrey, 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=New+spectrophotometric+equations+for+determining+chlorophyll+a,+b,+c1,+c2+in+higher+plants,+algae+and+natural+phytoplankton.">New spectrophotometric equations for determining chlorophyll a, b, c1, c2 in higher plants, algae and natural phytoplankton.</a> Biochem. Physiol. Pflanz. 167, 191-194 (1975).</li><li>Morgan, R. M., Ivanov, A. G., Priscu, J. C., Maxwell, D. P., H&#252;ner, N. 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=Structure+and+composition+of+the+photochemical+apparatus+of+the+Antarctic+green+alga,+Chlamydomonas+subcaudata.">Structure and composition of the photochemical apparatus of the Antarctic green alga, Chlamydomonas subcaudata.</a> Photosyn. Res. 56, 303-314 (1998).</li><li>Guillard, R. R. L., Smith, W. L., Chanley, M. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Culture+of+phytoplankton+for+feeding+marine+invertebrates.">Culture of phytoplankton for feeding marine invertebrates.</a> Culture of Marine Invertebrate Animals. , 29-60 (1975).</li><li>Johnson, M. D., Tengs, T., Oldach, D., Stoecker, D. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sequestration,+performance,+and+functional+control+of+cryptophyte+plastids+in+the+ciliate+Myrionecta+rubra+(Ciliophora.">Sequestration, performance, and functional control of cryptophyte plastids in the ciliate Myrionecta rubra (Ciliophora.</a> J. Phycol. 42, 1235-1246 (2006).</li><li>Rippka, R., Deruelles, J., Waterbury, J., Herdman, M., Stanier, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Generic+assignments,+strain+histories+and+properties+of+pure+cultures+of+cyanobacteria.">Generic assignments, strain histories and properties of pure cultures of cyanobacteria.</a> J. Gen. Microbiol. 111, 1-61 (1979).</li><li>Not, F., del Campo, J., Balague, V., De Vargas, C., Massana, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=New+insights+into+the+diversity+of+marine+picoeukaryotes.">New insights into the diversity of marine picoeukaryotes.</a> PLoS ONE. 4, e7143 (2009).</li><li>Sherr, B. F., Sherr, E. B., Caron, D. A., Vaulot, D., Worden, A. Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Oceanic+Protists.">Oceanic Protists.</a> Oceanography. 20, 130-135 (2007).</li><li>Finlay, B. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Protist+taxonomy:+an+ecological+perspective.">Protist taxonomy: an ecological perspective.</a> Philos. Trans. R. Soc. Lond. B. Biol. Sci. 359, 599-610 (2004).</li><li>Foissner, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Protist+diversity:+estimates+of+the+near-imponderable.">Protist diversity: estimates of the near-imponderable.</a> Protist. 150, 363-368 (1999).</li><li>Gast, R. J., Moran, D. M., Dennett, M. R., Caron, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Kleptoplasty+in+an+Antarctic+dinoflagellate:+caught+in+evolutionary+transition.">Kleptoplasty in an Antarctic dinoflagellate: caught in evolutionary transition.</a> Environmental Microbiology. 9, 39-45 (2007).</li><li>Gast, R. J., Moran, M. A., Beaudoin, D. J., Blythe, J. N., Dennett, M. R., Caron, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High+abundance+of+a+novel+dinoflagellate+phylotype+in+the+Ross+Sea.">High abundance of a novel dinoflagellate phylotype in the Ross Sea.</a> Antarctica. J. Phycol. 42, 233-242 (2006).</li><li>Moran, D. M., Anderson, O. R., Dennett, M. R., Caron, D. A., Gast, R. 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+Description+of+Seven+Antarctic+Marine+Gymnamoebae+Including+a+New+Subspecies,+Two+New+Species+and+a+New+Genus:+Neoparamoeba+aestuarina+antarctica+n.+subsp.,+Platyamoeba+oblongata+n.+sp.,+Platyamoeba+contorta+n.+sp.+and+Vermistella+antarctica+n.+gen.+n.+sp.">A Description of Seven Antarctic Marine Gymnamoebae Including a New Subspecies, Two New Species and a New Genus: Neoparamoeba aestuarina antarctica n. subsp., Platyamoeba oblongata n. sp., Platyamoeba contorta n. sp. and Vermistella antarctica n. gen. n. sp.</a> Journal of Eukaryotic Microbiology. 54, 169-183 (2007).</li><li>Rose, J. M., Vora, N. M., Countway, P. D., Gast, R. J., Caron, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+temperature+on+growth+rate+and+gross+growth+efficiency+of+an+Antarctic+bacterivorous+protist.">Effects of temperature on growth rate and gross growth efficiency of an Antarctic bacterivorous protist.</a> The ISME journal. 3, 252-260 (2009).</li></ol>

最近的分子研究报告通过一系列环境3，19，20高的单细胞真核生物的多样性;然而，由于缺乏整个完整的原生生物栖息地范围的分离，这些个别物种食物链的职能作用在很大程度上是未知之数。在这项研究中，我们所描述的方法，以丰富的真核生物的参展环境，从一个相对欠永久冰雪覆盖的南极湖泊的代谢多功能的微生物物种。从不同采样深度湖波尼丰富的文化表现差的固碳率​​，生长?...

<table border="1"><tbody><tr><td> 试剂名称 </td><td> 公司 </td><td> 目录编号 </td><td> 评论 </td></tr><tr><td> BBM协会</td><td>西格玛</td><td> B5282 </td><td></td></tr><tr><td> BG11 </td><td>西格玛</td><td> C3061 </td><td></td></tr><tr><td>的F / 2 </td><td>西格玛</td><td> G9903 </td><td></td></tr><tr><td> GF /过滤器，25毫米</td><td> Fisher Scientific则</td><td> 09-874-64 </td><td></td></tr><tr><td> GF / F滤波器，47毫米</td><td> Fisher Scientific则</td><td> 09-874-71 </td><td></td></tr><tr><td>聚醚砜过滤，0.45μm孔径，47毫米; </td><td>颇尔生命科学</td><td> 61854 </td><td></td></tr><tr><td>无菌细胞培养瓶，25厘米2 </td><td>康宁</td><td> 430639 </td><td></td></tr><tr><td> diurna升增长室</td><td>厂商VWR </td><td> 35960-076 </td><td></td></tr><tr><td>氧化锆/二氧化硅珠，0.1毫米diamter， </td><td> BioSpec产品</td><td> 11079101z </td><td></td></tr><tr><td>迷你珠打浆机</td><td> BioSpec产品</td><td> 3110BX </td><td></td></tr><tr><td>螺旋盖离心管（1.5微升） </td><td>美国科学</td><td> 1415-8700 </td><td></td></tr><tr><td>的NaH 14 CO 3的 </td><td> ViTrax </td><td> VC的194 </td><td>在400μL等份保持在-20°C </td></tr><tr><td> RuBP </td><td>西格玛</td><td> R0878-100毫克</td><td>溶解在10毫米的Tris-丙酸（pH值6.5） </td></tr></tbody></table>

establishment of microbial eukaryotic enrichment cultures from a chemically stratified antarctic lake and assessment of carbon fixation potential

波尼湖是众多永久位于麦克默多干谷，南极洲冰层覆盖的湖泊之一。常年冰盖保持化学分层的水柱，不同于其他内陆水体，主要是防止外部输入从河流的碳和营养。生物群暴露在众多的环境压力，包括全年的养分严重缺乏，气温低，极端的阴凉处，矿化，24小时黑暗，在冬季1。这些极端的环境条件限制在波尼湖生物群几乎完全微生物2。 单细胞微生物的真核生物（称为“原生生物”）是在全球生物地球化学循环的重要球员，在干热河谷湖泊的碳循环中发挥重要的生态作用，占小学和三级水生食物网中的角色。在干热河谷水产食品网站，修正我的原生生物organotrophic生物体的有机碳4，2 norganic碳（自养）的主要生产者。 phagotrophic或能异养细菌和较小的原生生物摄入原生生物作为食物网中的顶级掠食者。最后，未知的原生动物人口的比例是能够结合的混合营养代谢6，7。在原生生物mixotrophy涉及的能力结合猎物微生物phagotrophic摄入的光合能力。这种形式的mixotrophy不同于细菌混合营养代谢，一般涉及吸收溶解的碳分子。目前有极少数原生动物分离永久冰覆盖的极地湖泊，在这种极端环境中的原生生物多样性和生态的研究已经有限，8，4，9，10，5。的原生生物代谢的多功能性，将有助于更好地理解简单的干热河谷湖的食物网中的发展模式，为rOLE在全球碳循环中的原生生物。 我们采用隔离波尼湖潜在的光合和混合营养的原生生物富集培养方法。水柱采样深度被选为基于初级生产最大值和原生动物进化多样性4，第11的位置，以及影响原生动物的营养模式的主要非生物因素的变化：浅层取样深度为主要营养成分的限制，而更深层次的采样深度是有限的可用光。此外，湖水样品，辅以多种类型的生长介质，促进多种光合生物体的生长。 Rubisco活化酶催化的速率限制在CBB）卡尔文森Bassham（周期，主要途径，其中自养有机体修复无机碳和有机碳含量较高的营养水平，在水生和陆生食物链12步。在这项研究中，Wé应用放射性同位素法修改过滤样品13监察固碳潜力，并在湖的波尼富集文化的代谢多功能的代理最大的羧化酶活性。

Watch this Scientific Journal Video about Establishment of Microbial Eukaryotic Enrichment Cultures from a Chemically Stratified Antarctic Lake and Assessment of Carbon Fixation Potential at JoVE.com

Establishment of Microbial Eukaryotic Enrichment Cultures from a Chemically Stratified Antarctic Lake and Assessment of Carbon Fixation Potential

微生物的真核生物都在永久性冰雪覆盖的南极湖泊源的光合衍生的碳和顶级掠食性物种。本报告介绍了富集培养的方法分离代谢多功能微生物真核生物，从南极湖，湖波尼，使用核酮糖1,5 - 二磷酸羧化酶加氧酶（Rubisco活化酶）活性放射性同位素检测和评估无机碳固定潜力。

建立真核微生物的富集培养从化学分层南极湖和固碳潜力评估

Lake Bonney is one of numerous permanently ice-covered lakes located in the McMurdo Dry Valleys, Antarctica. The perennial ice cover maintains a ...

Research

JoVE Journal

Biology

Preparation of Neuronal Cultures from Midgastrula Stage Drosophila Embryos

从Midgastrula舞台果蝇胚胎神经文化的制备

This video illustrates the procedure for making primary neuronal cultures from midgastrula stage Drosophila embryos. The methods for collecting embryos ...

科研

生物学

Enrichment of NK Cells from Human Blood with the RosetteSep Kit from StemCell Technologies

NK细胞与干细胞技术的RosetteSep套件的人体血液的富集

Natural killer (NK) cells are large granular cytotoxic lymphocytes that belong to the innate immune system and play major roles in fighting against cancer ...

Extraction of High Molecular Weight DNA from Microbial Mats

超高分子量DNA提取微生物席

Successful and accurate analysis and interpretation of metagenomic data is dependent upon the efficient extraction  of high-quality, high molecular weight ...

Collection, Isolation and Enrichment of Naturally Occurring Magnetotactic Bacteria from the Environment

从环境中自然产生的趋磁细菌的采集，分离和富集

Magnetotactic bacteria (MTB) are aquatic microorganisms that were first notably described in 19751 from sediment samples collected in salt marshes of ...

Pyrosequencing for Microbial Identification and Characterization

焦磷酸测序的微生物鉴定和表征

Pyrosequencing is a versatile technique that facilitates microbial genome sequencing that can be used to identify bacterial species, discriminate ...

A Hybrid DNA Extraction Method for the Qualitative and Quantitative Assessment of Bacterial Communities from Poultry Production Samples

一种混合DNA提取方法细菌群落的定性和定量评估，从家禽生产样品

The efficacy of DNA extraction protocols can be highly dependent upon both the type of sample being investigated and the types of downstream analyses ...

Genome-wide Purification of Extrachromosomal Circular DNA from Eukaryotic Cells

从真核细胞染色体外的环状DNA基因组范围内的净化

Extrachromosomal circular DNAs (eccDNAs) are common genetic elements in Saccharomyces cerevisiae and are reported in other eukaryotes as well. EccDNAs ...

A Protocol for Using Gene Set Enrichment Analysis to Identify the Appropriate Animal Model for Translational Research

一个协议来使用基因集富集分析以确定合适的动物模型转化型研究

Recent studies that compared transcriptomic datasets of human diseases with datasets from mouse models using traditional gene-to-gene comparison ...

Establishment of a Clonal Culture of Unicellular Conjugating Algae

单细胞共轭藻类无性系培养的建立

It is essential to establish clonal cultures of microalgae for use in studies of various topics, such as physiology, genetics, taxonomy, and microbiology. ...

Establishment and Maintenance of Gnotobiotic American Cockroaches (Periplaneta americana)

Establishment and Maintenance of Gnotobiotic American Cockroaches Periplaneta americana

建立和维护侏罗育美国蟑螂 （佩里普拉内塔美洲）

Gnotobiotic animals are a powerful tool for the study of controls on microbiome structure and function. Presented here is a protocol for the establishment ...