特别鸣谢植物生长融资及EcoCore分析设备在科罗拉多州立大学和许多学生和老师谁帮助在这个项目上。这项工作是由美国国家科学基金会授予的DEB＃0918482，美国农业部国家需求奖学金，以及Cotrufo-Hoppess基金，用于土壤生态学研究提供资金。

1。硐室施工<ol><li>构造的标签室中的温室，以允许最大的自然光潜力的植物生长。确保充足的电力供应是提供给所有的电源室部件。 </li><li>通过安装3.175毫米厚的透明亚克力墙（聚碳酸酯也将是合适的）和6.35毫米与白色烤漆钢板地面上以最大化太阳能反射率的铝框厚的透明亚克力吊顶构造标记室。室的尺寸可以定制，以满足个人的研究需要。 </li><li>安装在¾室中（19毫米）胶合板上的煤渣块。 </li><li>钻洞的亚克力玻璃，铝框和钢地板和使用螺钉固定所有组件连接在一起。 </li><li>轴封采用硅填缝所有接缝，以确保密封良好。 </li><li>通过安装在长螺丝的压克力面板中的一个部分，它可以用雷莫拧下来构建一个门vable蝶形螺母。 </li><li>封门挡风雨条，防止漏气。 </li><li>选择一个区域直接相邻的腔作为控制中心安装的所有温度，湿度， 二氧化碳控制和监测设备。 </li><li>小心钻小孔中相邻的控制中心的所有电线和气体配管腔壁。使用硅填缝密封周围所有电线和管道的孔洞，防止漏气。 </li><li>通过与高水平的CO 2（ 例如 800ppm的）填充它，并让它坐通宵测试室漏气。如果CO 2浓度在腔室中被保持在其原有的水平，那么它是不透气的。如果浓度下降过夜，然后所有的接缝应检查并重新密封采用硅填缝，直到气密封的实现。 </li><li>对于一些植物适应高光条件下，添加一个连接到一个定时器到c的直接外部灯光汉伯以增加光线的穿透和植物生产力。 </li></ol> 2。温度和湿度控制器<ol><li>通过安装一个商用的分体式空调器与位于所述腔室内部的冷却（蒸发器）的线圈和位于温室中的​​热量消散以外的压缩机和冷凝器盘管调节室温度。置的空气调节，以维持所需的温度。 </li><li>使用一个小房间除湿机来控制湿度的室内。 <ol><li>通过相邻的除湿腔室的地板上钻一个排水孔。 </li><li>从除湿器除去冷凝物收集器，和从除湿通过在腔室底板上的排水孔连接引流管。 </li><li>下室，将一个开放的罐子装满水的除湿机直接排入。这将创建一个密闭的密封，但也允许压力平衡。 </li></ol></li><lI&gt;安装的控制器，如欧米茄i系列的控制器，在控制中心与所述腔室内部的湿度传感器。 <ol><li>控制器连接到除湿系统，固态继电器和设定湿度控制器具有高报警和摆动，以保持在最佳的室内生长条件。最佳的温度和湿度条件下会有所不同对于不同的植物物种。 </li></ol></li></ol> 3。 CO 2的控制<ol><li>的13 C-CO 2富集是使用两种纯的CO 2气体储罐，10原子％1实现13 C-CO 2或更高和1.1％（原子）的13 C-CO 2（天然丰度）1。 </li><li>由具有隔膜泵连续地通过一个红外气体分析仪（IRGA）吸取室空气中，然后泵送空气回到腔室中，从而保持一个封闭的系统（ 图1）监测CO 2浓度。 </li><li>硒TA低报警和死区的IRGA软件到一个期望的范围内维持CO 2浓度。在这里，我们用360 ppm的低限报警用40 ppm的死区，以保持二氧化碳 360-400 ppm的浓度之间。 </li><li>计量阀连接到每个罐，仔细调整以达到目标的13 C丰度水平。设置水箱的监管机构20磅。 </li><li>插入计量阀和各槽的调节器之间的电磁阀。加入两个计量阀的出口一起和管成腔室的中心。丝的固态继电器的IRGA输出来控制电磁阀（ 图1）。 </li></ol> 4。基于Web的远程监控系统<ol><li>监测CO 2浓度从IRGA软件将其记录到本地文件每隔30秒。 </li><li>创建一个自定义实用程序（ 如 Perl或其他编程语言）来接从本地的CO 2日志文件条目起来，伴随着目前的笔记本电脑时间戳，并将其上传到后端的Web应用程序。 </li><li>设置Web应用程序来查询温湿度传感器的数据。 </li><li>使用监控系统，以检查室，每五分钟的温度状态，以防止潜在的温度峰值会破坏植物，如果空调系统出现故障。 </li><li>监视在任何标准web浏览器中的CO 2，温度和湿度的数据，以便任何意外温度尖峰或CO 2滴可以立即参加到。 </li></ol> 5。灌溉系统<ol><li>钻一个小孔中，每罐的丙烯酸玻璃腔室的壁。 </li><li>使用冲洗管，以创建每罐1的滴灌环，并通过该室壁喂冲洗管到外部。 </li><li>密封周围的冲洗管与硅酮填缝孔以防止空气泄漏。 </li><li>在腔室的外部，连接冲洗管的管，用于蠕动泵。 </li><li>使用小管夹，以关闭所有冲洗管道，以防止浇水之间的空气泄漏。 </li></ol> 6。盆栽植物<ol><li>选择一个锅的大小适合正在生长的植物。这里，40个，15升罐的使用。 </li><li>通过混合砂，蛭石，和更新的多孔陶瓷建立一个土壤 - 自由盆栽混合土。 </li><li>测试盆栽组合称量填充锅干，用浸泡的水锅中，并允许它完全耗尽，并称重锅湿的保水能力。使用该最大持水能力，确保多余的标记肥水不从花盆浇水过程中泄漏。 </li><li>他们种植在花盆前发芽种子盆栽土壤。这确保了只有成功发芽的种子开始在标签室。 </li><li>接种吨<html>他幼苗新鲜的土壤泥浆引入有益微生物。 </li><li>一旦种子已发芽，仔细移苗到所需的数字盆。 </li><li>盆栽后，将花盆放入腔和组装每罐与个人灌溉管。 </li></ol> 7。密封室<ol><li>当第一密封的门，腔室中的大质量的外部空气充满腔室空间。擦洗此外部的CO 2通过连接一个碱石灰洗涤器，以使空气泵填充所述腔室回升至400ppm的使用13 C-CO 2罐混合物前擦洗CO 2浓度降低到至少200-250 ppm的。 </li><li>尽量保持室内封闭过生长季节的持续时间，以减少天然丰度的二氧化碳污染。 </li><li>监测植物生长可视化，并根据需求调整施肥和灌溉。 </li></ol>书名“&gt; 8。施肥和灌溉<ol><li>用肥料的溶液，如修饰的Hoagland溶液23，通过灌溉系统施肥的植物。 </li><li>用15 N-KNO 3下解的有针对性的原子％15 N级混合98原子％15 N-KNO 3与天然丰度15标记的肥料15 N，N-KNO 3（0.37原子％15 N）。 </li><li>混淆够在每次受精事件的肥料溶液在整个腔室的基础上，盆栽混合土的持水能力的。将肥料适量的一锅在一个玻璃瓶，并准备尽可能多的坛子里有盆。 </li><li>松开灌溉软管，并把他们每个人在用肥液罐，然后将它们连接到蠕动泵。 </li><li>水厂通过通过个人点滴IRRI蠕动泵抽水gation软管定期为植物需要它。 </li><li>施肥与霍格兰的解决方案应该遵循植物的需求或实验设计，以增加营养需求为工厂提高生产率，以最大限度地提高生产力。 </li><li>首先，通过灌溉管泵的Hoagland溶液中，然后通过泵进行水漂洗，以尽量减少在管藻类和细菌的生长。 </li><li>施肥和灌溉，消除室内空气泄漏后夹紧固定所有的软管。 </li></ol> 9。统一和差异化标记<ol><li>对于结构和代谢成分鉴别标签，从标签室取出的植物前1-3周的收获。植物是能够被均匀地标记可以连续保持在密封室中的标签，直到收获和灌溉用15 N-霍格兰溶液。 </li><li>保持在这段时间在温室中取出植物，使他们得到足够的光和CO 2的天然丰度13 C。 </li><li>对于差分15 N标记，继续施肥和灌溉植物像往常一样，但在霍格兰的肥液使用天然丰度15 N-KNO 3。 </li></ol> 10。收成<ol><li>停止浇水1周厂前收获这么植物开始衰老和盆栽介质变干。 </li><li>打开腔和移动花盆出来的地上生物量的直接裁剪和收获</li><li>倒出盆栽组合和根过粗筛。 </li><li>使用屏幕的根源来自盆栽组合中分离出来，动摇自由组合盆栽的根部。 </li><li>将根部2毫米筛上的和漂洗它们与水以除去任何剩余的灌封材料。使用镊子去除可能附着在根部任何蛭石。 </li><li>让根部风干，准备未来的实验。 </li></ol><ol><li>称取风干的植物材料，以确定标签腔生物量。 </li><li>磨子样本进行化学分析。 </li><li>将2.0克烘干（60℃）垃圾在125毫升酸洗烧瓶中，加入50毫升去离子水。 </li><li>将样品放置在预热（60℃）搅拌盘，并把搅拌棒的烧瓶中。设置搅拌200转，并允许样品加热30分钟。 </li><li> 30分钟后，过滤通过20微米尼龙网丝上的真空过滤系统的垃圾溶液。 </li><li>提取物转移到预先称重的酸洗管和冻结。 </li><li>将固体残余物​​垫料转移到预先称重的铝盘中，干燥在60℃下干燥，以确定热水残留量后秤盘和垃圾。 </li><li>冷冻干燥的热水提取物并称重，以确定热水提取物的质量。 </li><li>分析烘干（60℃）垃圾，冻干热水提取和烘干热水中残留的元素分析仪 - 同位素比值质谱（EA-IRMS）。 </li></ol>

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	<li>Bird, J. A., Torn, M. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fine+roots+vs.+Needles:+A+comparison+of+(13)C+and+(15)N+dynamics+in+a+ponderosa+pine+forest+soil.">Fine roots vs. Needles: A comparison of (13)C and (15)N dynamics in a ponderosa pine forest soil.</a> Biogeochemistry. 79, 361-382 (2006).</li><li>Bird, J. A., van Kessel, C., Horwath, W. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Stabilization+of+C-13-carbon+and+immobilization+of+N-15-nitrogen+from+rice+straw+in+humic+fractions.">Stabilization of C-13-carbon and immobilization of N-15-nitrogen from rice straw in humic fractions.</a> Soil Sci. Soc. Am. J. 67, 806-816 (2003).</li><li>Denef, K., Six, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Contributions+of+incorporated+residue+and+living+roots+to+aggregate-associated+and+microbial+carbon+in+two+soils+with+different+clay+mineralogy.">Contributions of incorporated residue and living roots to aggregate-associated and microbial carbon in two soils with different clay mineralogy.</a> Eur. J. Soil Sci. 57, 774-786 (2006).</li><li>Rubino, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Carbon+input+belowground+is+the+major+C+flux+contributing+to+leaf+litter+mass+loss:+Evidences+from+a+C-13+labelled-leaf+litter+experiment.">Carbon input belowground is the major C flux contributing to leaf litter mass loss: Evidences from a C-13 labelled-leaf litter experiment.</a> Soil Biol. Biochem. 42, 1009-1016 (2010).</li><li>Cotrufo, M. F., Wallenstein, M. D., Boot, C. M., Denef, K., Paul, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Microbial+Efficiency-Matrix+Stabilization+(MEMS)+framework+integrates+plant+litter+decomposition+with+soil+organic+matter+stabilization:+do+labile+plant+inputs+form+stable+soil+organic+matter.">The Microbial Efficiency-Matrix Stabilization (MEMS) framework integrates plant litter decomposition with soil organic matter stabilization: do labile plant inputs form stable soil organic matter.</a> Glob. Change Biol. 19, 988-995 (2013).</li><li>Mambelli, S., Bird, J. A., Gleixner, G., Dawson, T. E., Torn, M. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Relative+contribution+of+foliar+and+fine+root+pine+litter+to+the+molecular+composition+of+soil+organic+matter+after+in+situ+degradation.">Relative contribution of foliar and fine root pine litter to the molecular composition of soil organic matter after in situ degradation.</a> Org. Geochem. 42, 1099-1108 (2011).</li><li>Prescott, C. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Litter+decomposition:+what+controls+it+and+how+can+we+alter+it+to+sequester+more+carbon+in+forest+soils.">Litter decomposition: what controls it and how can we alter it to sequester more carbon in forest soils.</a> Biogeochemistry. 101, 133-149 (2010).</li><li>Denef, K., Roobroeck, D., Wadu, M., Lootens, P., Boeckx, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microbial+community+composition+and+rhizodeposit-carbon+assimilation+in+differently+managed+temperate+grassland+soils.">Microbial community composition and rhizodeposit-carbon assimilation in differently managed temperate grassland soils.</a> Soil Biol. Biochem. 41, 144-153 (2009).</li><li>Bird, J. A., Kleber, M., Torn, M. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=C-13+and+N-15+stabilization+dynamics+in+soil+organic+matter+fractions+during+needle+and+fine+root+decomposition.">C-13 and N-15 stabilization dynamics in soil organic matter fractions during needle and fine root decomposition.</a> Org. Geochem. 39, 465-477 (2008).</li><li>Andresen, L. C., Jonasson, S., Strom, L., Michelsen, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Uptake+of+pulse+injected+nitrogen+by+soil+microbes+and+mycorrhizal+and+non-mycorrhizal+plants+in+a+species-diverse+subarctic+heath+ecosystem.">Uptake of pulse injected nitrogen by soil microbes and mycorrhizal and non-mycorrhizal plants in a species-diverse subarctic heath ecosystem.</a> Plant Soil. 313, 283-295 (2008).</li><li>Horwath, W. R., Pregitzer, K. S., Paul, E. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=C-14+Allocation+in+tree+soil+systems.">C-14 Allocation in tree soil systems.</a> Tree Physiol. 14, 1163-1176 (1994).</li><li>Denef, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Community+shifts+and+carbon+translocation+within+metabolically-active+rhizosphere+microorganisms+in+grasslands+under+elevated+CO2.">Community shifts and carbon translocation within metabolically-active rhizosphere microorganisms in grasslands under elevated CO2.</a> Biogeosciences. 4, 769-779 (2007).</li><li>Pollierer, M. M., Langel, R., Korner, C., Maraun, M., Scheu, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+underestimated+importance+of+belowground+carbon+input+for+forest+soil+animal+food+webs.">The underestimated importance of belowground carbon input for forest soil animal food webs.</a> Ecol. Lett. 10, 729-736 (2007).</li><li>Stewart, D. P. C., Metherell, A. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Carbon+(C-13)+uptake+and+allocation+in+pasture+plants+following+field+pulse-labelling.">Carbon (C-13) uptake and allocation in pasture plants following field pulse-labelling.</a> Plant Soil. 210, 61-73 (1999).</li><li>Santos, F., Torn, M. S., Bird, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biological+degradation+of+pyrogenic+organic+matter+in+temperate+forest+soils.">Biological degradation of pyrogenic organic matter in temperate forest soils.</a> Soil Biol. Biochem. 51, 115-124 (2012).</li><li>Bromand, S., Whalen, J. K., Janzen, H. H., Schjoerring, J. K., Ellert, B. 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+pulse-labelling+method+to+generate+13C-+enriched+plant+materials.">A pulse-labelling method to generate 13C- enriched plant materials.</a> Plant Soil. 235, 253-257 (2001).</li><li>Putz, B. <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+in+situ-labelling+with+15N+and+13C+of+grassland+plant+species+by+foliar+brushing.">A simple method for in situ-labelling with 15N and 13C of grassland plant species by foliar brushing.</a> Methods Ecol. Evol. 2, 326-332 (2011).</li><li>Wichern, F., Mayer, J., Joergensen, R., M&#252;ller, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evaluation+of+the+wick+method+for+in+situ+&amp;lt;sup&amp;gt;13&amp;lt;/sup&amp;gt;C+and+&amp;lt;sup&amp;gt;15&amp;lt;/sup&amp;gt;N+labelling+of+annual+plants+using+sugar-urea+mixtures.">Evaluation of the wick method for in situ &amp;lt;sup&amp;gt;13&amp;lt;/sup&amp;gt;C and &amp;lt;sup&amp;gt;15&amp;lt;/sup&amp;gt;N labelling of annual plants using sugar-urea mixtures.</a> Plant Soil. 329, 105-115 (2010).</li><li>Fahey, T. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transport+of+Carbon+and+Nitrogen+Between+Litter+and+Soil+Organic+Matter+in+a+Northern+Hardwood+Forest.">Transport of Carbon and Nitrogen Between Litter and Soil Organic Matter in a Northern Hardwood Forest.</a> Ecosystems. 14, 326-340 (2011).</li><li>Stewart, C. E., Paustian, K., Conant, R. T., Plante, A. F., Six, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Soil+carbon+saturation:+Evaluation+and+corroboration+by+long-term+incubations.">Soil carbon saturation: Evaluation and corroboration by long-term incubations.</a> Soil Biol. Biochem. 40, 1741-1750 (2008).</li><li>Fry, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Stable Isotope Ecology. , (2006).</li><li>Nippert, J. B., Fay, P. A., Carlisle, J. D., Knapp, A. K., Smith, M. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ecophysiological+responses+of+two+dominant+grasses+to+altered+temperature+and+precipitation+regimes.">Ecophysiological responses of two dominant grasses to altered temperature and precipitation regimes.</a> Acta Oecol. Int. J. Ecol. 35, 400-408 (2009).</li><li>Hoagland, D. R. a. A., I, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> The Water-Culture Method for Growing Plants without Soil. , (1950).</li><li>Lett, M. S., Knapp, A. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Consequences+of+shrub+expansion+in+mesic+grassland:+Resource+alterations+and+graminoid+responses.">Consequences of shrub expansion in mesic grassland: Resource alterations and graminoid responses.</a> J. Veg. Sci. 14, 487-496 (2003).</li><li>Knapp, A. K., Seastedt, T. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Detritus+Aaccumulation+limits+productivity+of+tallgrass+prairie.">Detritus Aaccumulation limits productivity of tallgrass prairie.</a> Bioscience. 36, 662-668 (1986).</li><li>Ting, K. C., Giacomelli, G. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Solar+photosynthetically+active+radiation+transmission+through+greenhouse+glazings.">Solar photosynthetically active radiation transmission through greenhouse glazings.</a> Energy Agric. 6, 121-132 (1987).</li><li>McDermitt, D. K., Welles, J. M., Eckles, 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=Effects+of+Temperature,+Pressure+and+Water+Vapor+on+Gas+Phase+Infrared+Absorption+by+CO2.">Effects of Temperature, Pressure and Water Vapor on Gas Phase Infrared Absorption by CO2.</a> , .</li></ol>

作者什么都没有透露。

这种设计为一个连续的同位素标记腔室被用来生产均匀且差异13 C和15 N标记为A。 gerardii随后的现场和实验室试验。在三个生长操作的季节，室成功维持温度，湿度，和设定参数范围内CO 2浓度（ 图2）。温度控制系统的可靠性时，高太阳辐射可以引起气密室过热夏季的高峰期是至关重要的。消除由蒸腾过量湿度从生长植物可确保植物气孔保持开启的光合作用吸收22和冷凝水不抑制光渗透或损坏腔室的结构。 不久的连续监测CO 2浓度由IRGA软件保持了持续植物的13 C标记，当他们在室内越来越大。由于A的高光合活性gerardii在这个房间成长，CO 2是在白天高峰生长期时，光合活性画了CO 2浓度降至360 ppm时，约每15-20分钟频频注入到系统中。测光富集并通过统一的植物组织标识的生长季节允许受控4.4原子％的13 C大气自然丰度13 C-CO 2罐13 C-CO 2的产生也可以通过混合13 C-钠来实现碳酸氢盐或碳酸盐13 C-钠和盐酸，但这种类型的系统比较复杂，并且需要更多的监测和维护，因此，建议使用13 C-CO 2气体。用于监测CO 2 conce一个重要的考虑使用IRGA ntrations是红外分析仪失去三分之二的灵敏度测量13 C-CO 2时。约2.9％PPM为4.4％，13 C-CO 2的混合物，这种低估是我们非常关注的不是，但是当在更高的13 C水平27标记可能会成为一个更显著的问题。 A. gerardii是一个温暖的季节常年高草草原禾草种。该室的设计A.进行了优化gerardii生产（ 图1）。所述腔室的尺寸和高度被选择考虑了A的最大高度生产率gerardii领域，以及为所希望的植物的生物量产生对将来的实验。A. gerardii是已知的光限制在该领域24,25。温室内的PAR可以通过30-47％，而外部电平26被削弱。由于我们的工厂非常明显的生长在温室内的亚克力玻璃室，光合有效辐射的限制是一个问题。当打开时，荧光灯的PAR增加了室内的9.5％，这可能有助于提高生产力这个角度敏感的物种。当使用这个腔体设计，以种植其他种类的植物特有的生理需要，如大小，灯光，营养要求，对温度的敏感性和土壤水分应精心定制，以维持植物最佳生长条件。 当使用昂贵的同位素标记的化合物，如10％（原子）的13 C-CO 2和98％（原子）15 N-KNO 3，贴标签的工作效率是一个重要的考虑因素。这个腔体设计使所有的努力，以密封腔和空气泄漏减少进入和离开该室的优化的13 C标记。如果该室在生长季节从未打开过，那么，没有的13 C标记的版的CO 2从该腔室泄漏到大气中。在CO 2夜间呼吸过程中建立不出现损坏植物生长，并迅速溶于日出（ 图2）之后。在差异化标记，该室被短暂打开，取出差异化标记盆但这并未冲淡连续标记植物（ 表1）目标4.4原子％的13 C标记。的13 C标记还通过优化洗涤出来的在密封初始大气被困在室内。在没有大气CO 2的初始擦洗室初步测试，植物进行测定，以在第一叶比后叶生产产生了稀释13 C水平。大气中的CO 2室后关闭的初始擦洗出现，允许连续的13 C标记的消除这个问题幼苗到成熟。保持砂，蛭石，和粘土的土壤-自由灌封混合物也消除了天然丰度从土壤呼吸CO 2的污染。土壤的消除来自系统确实需要小心受精和接种的考虑，这可能是唯一的，以不同的植物物种。通过在6.7％（原子）15 N（ 表产生高度标记的植物材料中的靶向7原子％15 N Hoegland的溶液15 N标记1）。从目标15 N标签轻微的稀释可能会被一些自然丰度N的盆栽组合或从乡土接种引起的。 在化合物的生物合成，13℃（或15 N）的天然歧视的发生是由于动力分馏和合成的化合物非统计同位素分布的结果。因而，在C的情况下，次要产物（ 例如脂质，酚化合物）相比，初级产品（碳水化合物），一般在耗尽的13 C。这个天然的13 C歧视似乎也坚持当植物生长中的富集的13 C的气氛，可以在热水提取物和均匀标记的植物（ 表1中的热水残基的原子％的13 C的细微差别可以看出）。相比富集这种自然动力分馏是非常小的，不妥协的标签的均匀性。 的结构和代谢植物组织中差异化标记是一种新的技术有潜力为凋落物分解，微生物生态学和土壤有机质的形成深造。在13 C和15 N的热水提取物和热水残留的差异表明一个显著稀释13℃，在浸出15 N，低从结构材料厂（热水残基）分子量化合物（热水提取物）后，7，14，22天差异化标记（P &lt;0.005）的。这个差的植物组织的标记可用于通过一个生态系统分别跟踪的结构和代谢成分的命运。 13 C微分标签比15无差异化标记更极端。从13 C-CO 2标记的去除大气中的植物时，这可能是由于中的13 C稀释的即时性，同时在15 N标记仍停留在盆栽混合一段时间，并且发生15 N稀释更慢。对于15 N更激烈的差异化标记，可以考虑在最后几周增长的腔外冲洗水之前，自然丰度受精盆。 这种连续13的设计和操作&lt;均匀或差分，代谢和结构，植物组织贴标/ SUP&gt; C和15 N标记系统提供了一种新的方法用于生产同位素标记的植物材料为先进的研究。该室的设计和操作细节已被选定为4.4原子％，13℃和7％（原子）A的15 N标记gerardii，但可以根据其他植物种类和同位素标记的水平。这里所描述的生长条件应该定制，以满足特定的植物物种的利益的大小，温度，湿度，光，水和营养需求。标签带18 O或2小时，也可以通过标记在灌溉系统中使用的水来实现。这里所描述的系统解决了许多均匀的挑战和差分植物材料的13 C标记。这个基本的腔室设计可用于其他研究小组，以产生高度标记的植物材料为副词anced生态系统生物地球化学研究。

了解植物 - 土壤 - 大气过程的动力学是至关重要的准确预测全球碳（C）和氮（N）当前和未来的环境条件下循环功能如何。稳定同位素在植物 - 土壤 - 大气碳，氮循环的定量研究的强大工具。从植物材料通过生态系统追踪罕见的稳定同位素提供了在生物地球化学循环研究的敏感信息，由CO 2通量与土壤有机质形成小规模的稳定同位素生物标志物的探测，例如 4,8,9。组合的13 C标记与15 N标记，或其它稳定同位素，如2 H或植物组织中18 O提供了高的检测，可追溯的，但复杂基质中植物和土壤生物化学耦合研究使用。均匀或差分标签结构和代谢的植物材料广告的能力DS还能力，以解决关于C和N通过生态系统循环的复杂问题。利用同位素标记的植物材料中碳，氮会计定量研究的好处，但是，依赖于生产13 C和15 N标记物，要么是一致或不同标记的能力。 同位素标记已被用于研究解决植物碳，氮同化10，分配11和根际沉积12。均匀13 C和15 N标记的植物材料为凋落物分解1,4研究一个复杂的标记底物，土壤有机质形成2,6，土壤CO 2排放量4，土壤食物网研究13，和土壤碳的研究停留时间2,14。利用13 C标记的生物炭的标记植物材料的研究也开始揭示以前被忽视的新信息土炭池15。而15 N，2 H和18 O标记都比较容易实现，通过水肥处理，仍然面临挑战，通过13 C-CO 2固定生产均匀13 C标记的植物材料。 在密封腔连续同位素标记从幼苗到成熟生产全厂统一的同位素标记。其它方法，如重复脉冲标记16和叶面施用或排汗17,18不产生均匀同位素标记的植物材料，也不具体的C-的化合物的明确差异化标记（ 例如，代谢与结构）19。在同位素标记的一个重要考虑是标记效率，由于在标签中使用稀有同位素富集的化合物的成本高。虽然连续的13 C标记在过去2-4,20被使用，也没有给我们的知识连续标签腔具有高效率标签和金额以及同位素标记的均匀性精确控制证据公布详细的技术说明。 对凋落物分解及土壤的最前沿有机质形成的研究是代谢的植物材料（ 即浸出的，不稳定的，低分子量化合物）和结构材料厂（ 即 ，木质素，纤维素，半纤维素）是不同的微生物方面处理的概念利用效率，土壤有机质的形成和长期土壤碳贮存5-7。即差异标记在其结构和代谢成分的植物材料，因此，在推进凋落物分解及土壤有机质的形成研究的有用工具。具有双同位素差异化标记分别允许对结构和代谢成分的追踪通过生态系统使用多池同位素techniquE 21。 与13 C等同位素在密封腔连续同位素标记需要认真注意植物的生理条件，最大限度地提高工厂的生产效率和同位素标记效率。白天温度尖峰必须加以控制，以防止植物损伤中的气密室生长时。湿度和温度的最佳范围须维持开放的植物气孔和CO 2吸收22。高水平的湿度原因雾化室壁，从而最大限度地减少光的可用性，并可能损坏腔室结构。仔细考虑消除自然丰度同位素从腔（ 例如，从与土壤有机质灌封未来），并防止暴露于外部空气的同位素标记效率与昂贵的重型同位素标记的化合物，工作时是很重要的。 在这里，我们提出了一个方法，用于构建和运行一个连续双13 C和15 N同位素标记室为植物材料的生产要么是统一标示或标示有在不同的层次的结构和代谢成分。的13 C标记被控制在室内的水平，而受精和15 N标记是在个人锅电平控制。代表性的结果示，说明这种方法来控制温度，湿度和CO 2浓度在整个生长季节的能力。结果从生长须芒草gerardii，卡奥也证明了该方法的产生均匀的或不同标记植物材料的能力。所描述的特定腔室的设计和操作方案可以被修改，以生长不同的植物物种，以及容纳2 H或18 O标记。

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		<tr height="42">
			<td height="42" style="height:42px;width:216px;">40 in Remote Tower Fan</td>
			<td style="width:81px;">Living Accents</td>
			<td style="width:119px;">FS10-S3R-A</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">42 in Electric Tower Fan with remote control</td>
			<td style="width:81px;">LASKO</td>
			<td style="width:119px;">2559</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Mr. Slim Split-type air conditioner</td>
			<td style="width:81px;">Mitsubishi Electric</td>
			<td style="width:119px;">R410A</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Electronic 24 hr time switches</td>
			<td style="width:81px;">Intermatic</td>
			<td style="width:119px;">ET100C</td>
			<td>Operates Fluorescent Light System</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">iSeries Humidity and Temp Controller</td>
			<td style="width:81px;">Omega</td>
			<td style="width:119px;">CHiTH-i8DV33-5-El</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Solid State Relay</td>
			<td style="width:81px;">Omega</td>
			<td style="width:119px;">SSRL240</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">CKR Series Solid State Relay</td>
			<td style="width:81px;">Crydon</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Solenoid Coils</td>
			<td style="width:81px;">Dayton</td>
			<td style="width:119px;">6X543</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">GasHound CO2 Analyzer</td>
			<td style="width:81px;">LI-COR</td>
			<td style="width:119px;">LI-800</td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Dehumidifier</td>
			<td style="width:81px;">Dayton</td>
			<td style="width:119px;">1DGX4</td>
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			<td height="21" style="height:21px;width:216px;">Specialty gas regulator</td>
			<td style="width:81px;">Airgas</td>
			<td style="width:119px;">CGA 580</td>
			<td></td>
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		<tr height="42">
			<td height="42" style="height:42px;width:216px;">T5 Hight Output Commercial Fluorescent Light System</td>
			<td style="width:81px;">Hydro Farm</td>
			<td style="width:119px;">FLT48</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Air pump</td>
			<td style="width:81px;">Thermo Scientific</td>
			<td style="width:119px;">420-1901</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Masterflex Cartridge Pump HeadSystem</td>
			<td style="width:81px;">Cole Parmer</td>
			<td style="width:119px;">7553-70</td>
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			<td height="21" style="height:21px;width:216px;">1900 Series Network Switch</td>
			<td style="width:81px;">Catalyst</td>
			<td style="width:119px;"></td>
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		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Profile Porus Ceramic Top Dressing</td>
			<td style="width:81px;">Greens Grade</td>
			<td style="width:119px;"></td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Industrial Quartz 40 mesh</td>
			<td style="width:81px;">Unimin</td>
			<td style="width:119px;">4095</td>
			<td></td>
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			<td height="42" style="height:42px;width:216px;">Mycorrhizae Professional Growing Medium</td>
			<td style="width:81px;">ProMix</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Vermiculite and Perlite</td>
			<td style="width:81px;">Thermo-Ran</td>
			<td style="width:119px;">C633</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Potassium Nitrate- 15N 98 atom %</td>
			<td style="width:81px;">Sigma-Aldrich</td>
			<td style="width:119px;">335134</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Carbon-13C Dioxide 10 atom %</td>
			<td style="width:81px;">Sigma-Aldrich</td>
			<td style="width:119px;">600180</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Medical grade CO2</td>
			<td style="width:81px;">Airgas</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Regulator</td>
			<td style="width:81px;">Airgas</td>
			<td style="width:119px;">&#160;CGA 350</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">4 in box fans</td>
			<td style="width:81px;">Grainger</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Masterflex PharMed BPT Tubing</td>
			<td style="width:81px;">Cole Parmer</td>
			<td style="width:119px;">HV-06508-24</td>
			<td></td>
		</tr>
	</tbody>
</table>

design and operation of a continuous 13c and 15n labeling chamber for uniform or differential, metabolic and structural, plant isotope labeling

从植物材料通过生态系统追踪罕见的稳定同位素提供了有关生态系统过程中最敏感的信息，由CO 2通量与土壤有机质形成小规模的稳定同位素生物标志物的探测。耦合多个稳定同位素，如13下用15 N，18 O 2或H有透露有关生物地球化学过程的复杂变换的化学计量关系更加信息的潜力。同位素标记的植物材料已用于凋落物分解及土壤有机质形成1-4的各项研究。从这些和其他的研究，然而，它已经很明显，植物材料的结构部件不同的表现比在微生物利用与长期碳储存5-7而言代谢成分（ 即 ，可浸出的低分子量化合物）。研究结构和代谢成分的能力分别提供了推进生态系统的生物地球化学研究的前沿一个强大的新工具。在这里，我们描述了用于生产13 C和15 N标记的植物材料要么是统一整个工厂标记或不同标记的结构和代谢植物成分的方法。 在这里，我们提出了一个连续13 C和15 N标记室，可以进行修改，以满足不同的研究需求的建设和运营。均匀标记的植物材料是由从幼苗到收获连续标记产生的，而差异化标记被从腔室周之前收获除去生长的植物获得。从生长须芒草代表性结果gerardii卡奥验证系统的能力，以有效地标记的植物材料在目标水平。通过这种方法，我们已经生产的植物材料用4.4原子％，13℃和6.7％（原子）15 &lt;/ SUP&gt; N均布厂的标签，或材料的差异达1.29原子％，13℃和0.56％（原子）标记15 N在其代谢和结构部件（热水提取和热水残留成分，分别）。挑战在于保持适当的温度，湿度，CO 2浓度，和光水平在一个密闭的13 C-CO 2气氛中成功的植物的生产。该室的描述是一项有用的研究工具，以有效地产生均匀或差分多同位素标记的植物材料在生态系统生物地球化学循环实验。

我们的标签腔为1.2米×2.4米×3.6米在大小，并持有40,15 L罐（ 图1）。计算机化IRGA控制系统维护的CO 2本组360和400 ppm的值之间的浓度在一天中的光合有效期间内（ 图2a）。在IRGA的低CO 2报警功能触发电磁阀，使二氧化碳从的13 C丰富和天然丰度的坦克进入室内，当浓度低于最小阈值（ 如 360 ppm的）。死区功能停止了流动，当浓度达到上设定值（ 如 400 ppm的）。在iSeries温度，并连接到整个生长季节设定的参数范围内举行的气候条件下，空调器和除湿机湿度监测系统（ 图2b）。我们使用了一吨重（3.5千瓦），空调机组，以柯EP室内凉爽。 远程监控系统所允许的记录数据被随时通过一个标准的Web浏览器中查看。在CO 2浓度，温度和湿度值下采样的Web应用程序来显示图形在过去的24-240小时，在24小时的增量。这创造了一个快速的视觉来确认每天的波动都在预期范围。查看Web界面也显示当前箱内的状态，以及提供警示潜在的问题，如没有收到新的数据。在任何时候，完整的数据集也可以从Web界面下载。 我们测量光合有效辐射（PAR）在立即室内和室的外部的四个点有和没有在使用量子传感器的夏天和一天中的中间的灯。在腔室中的PAR比外部低31.5％时，该腔室灯瓦特埃雷关闭，比当灯是在外观上降低22％。因此，腔室灯帮助在腔室中通过9.5％（P &lt;0.05），以增加显著PAR渗透。 我们连续标签制度能够产生2759克A的gerardii生物量，其中37％为地上生物量和其中63％为地下生物量。我们在统一的植物材料通过设置电磁阀上的两个二氧化碳储罐相应的（ 图1，表1）实现了4.4％（原子）的13 C全株标签。我们在统一的植物材料通过的修改霍格兰的解决方案23 KNO 3下解（ 表1）将98原子％15 N-KNO 3与0.37原子％15 N-KNO 3实现了6.7原子％15 N全株标签。我们浇水A. gerardii每周用750ml总流体（水加霍格兰的解决方案）在整个生长海儿子。我们受精与200-500毫升15 N取决于工厂的生产率标记霍格兰的每周解决方案。 我们利用热水提取法，以确定是否有统一和差异化标记的植物材料之间的同位素差异。对于差异化标记的植物，收获后，我们删除了完全死亡和另案处理这些，因为他们可能没有差异标记任何树叶。当看着13 C的含量，所有四个掺入天数分别为显著彼此不同的全草和热水提取物，但是对于热水残天14和22分别不显著彼此不同（ 表1）。当比较每一天内的植物组织的馏分中，热水提取物和残余物分别为显著彼此不同的所有四天，由第22天的整个植物，提取物，和残余物都显彼此ficantly不同（ 表1）。对于15 N掺入植物成分，有注册及植物组织馏分天之间的差异。对于热水提取所有四个纳入天都是显著互不相同为15 N和全厂和热水秸秆还田的短天数分别比成立（ 表1）的长天显著不同。植物组织的馏分中的均匀植株不显著彼此不同的15个 N，但对于差异化标记的垫料的热水提取物和残余物分别为显著彼此不同的15 N。 所有同位素值所使用的原子百分比（原子％）表示（ 等式1），这比％至高水平的重使用更精确的符号报道同位素富集21。例如： <img alt="" height="54" src="/files/ftp_upload/51117/51117eq2.jpg" width="202" /> （1） 在这项研究中，我们跑了统计分析采用SAS 9.2版。我们使用配对t检验来检验腔室内部和外部的光水平之间的差异。我们测试了在13℃，在PROC ANOVA采用单向方差分析（ANOVA）的热水提取物和热水残留15 N标记的差异。我们使用Duncan的多范围检验进行多重比较分析。意义被接受为0.05的P的水平。我们使用Wilcoxon秩和检验，以测试数据满足分析的假设。 <img alt="图1" fo:content-width="5in" fo:src="/files/ftp_upload/51117/51117fig1highres.jpg" src="/files/ftp_upload/51117/51117fig1.jpg" /> 图1原理图从空中鸟瞰的40锅容量连续多同位素标记室的IC图。虚线表示的电线，而实线表示气体或水管道。 <a href="https://www.jove.com/files/ftp_upload/51117/51117fig1highres.jpg" target="_blank">点击这里查看大图。</a> <img alt="图2" fo:content-width="5in" fo:src="/files/ftp_upload/51117/51117fig2highres.jpg" src="/files/ftp_upload/51117/51117fig2.jpg" /> 图2：A）平均CO 2浓度（ppm）（+ / - SE）在二十四小时内为整个生长季节B）的平均温度（℃），空心圆，湿度（％），实心圆（+ / - SE）在二十四小时内为整个生长季节<a href="https://www.jove.com/files/ftp_upload/51117/51117fig2highres.jpg" target="_blank">点此查看大IM</a>年龄。 <table border="0" cellpadding="0" cellspacing="0" width="559"><colgroup><col /><col /><col /><col /><col span="2" /></colgroup><tbody><tr height="17"><td height="17" style="height:17px;width:77px;"></td><td style="width:85px;"></td><td style="width:92px;">制服（0） </td><td style="width:97px;">差（7） </td><td style="width:104px;">微分（14） </td><td style="width:104px;">微分（22） </td></tr><tr height="19"><td height="38" rowspan="2" style="height:38px;width:77px;">整个产仔</td><td> 13碳原子％ </td><td> 4.46±0.02古物咨询委员会</td><td> 3.93±0.05巴</td><td> 3.64±0.03钙</td><td> 3.35±0.06分贝</td></tr><tr height="19"><td height="19" style="height:19px;"> 15 N原子％ </td><td> 6.69±0.07机管局</td><td> 6.72±0.01机管局</td><td> 6.33±0.06巴</td><td> 6.41±0.07巴</td></tr><tr height="19"><td height="38" rows泛=“2”的风格=“高度：38px;宽度：77px;”&gt;热水提取物</td><td> 13碳原子％ </td><td> 4.59±0.04机管局</td><td> 3.35±0.06降B </td><td> 2.79±0.06 CB </td><td> 2.37±0.03直流</td></tr><tr height="19"><td height="19" style="height:19px;"> 15 N原子％ </td><td> 6.69±0.03机管局</td><td> 6.43±0.01降B </td><td> 5.89±0.07分贝</td><td> 6.16±0.05 CB </td></tr><tr height="19"><td height="38" rowspan="2" style="height:38px;width:77px;">热水残渣</td><td> 13碳原子％ </td><td> 4.37±0.06抗体</td><td> 4.1±0.03巴</td><td> 3.79±0.10钙</td><td> 3.66±0.05钙</td></tr><tr height="19"><td height="19" style="height:19px;"> 15 N原子％ </td><td> 6.57±0.04巴</td><td> 6.71±0.02机管局</td><td> 6.45±0.02钙</td><td> 6.44±0.03钙</td></tr></tbody></table> 表1中。 <strong&gt;同位素组成和化学垃圾的统一和差异标记垃圾。腔以外的差异化标记的天数是在括号中。注册成立日之间的比较是大写字母（跨行）与垃圾组分之间是小写字母（下栏）为每个变量。

Watch this Scientific Journal Video about Design and Operation of a Continuous 13C and 15N Labeling Chamber for Uniform or Differential, Metabolic and Structural, Plant Isotope Labeling at JoVE.com

Design and Operation of a Continuous 13C and 15N Labeling Chamber for Uniform or Differential, Metabolic and Structural, Plant Isotope Labeling

这个方法介绍了如何建立和运营一个连续13 C和15 N同位素标记室统一或差分植物组织标签。从代谢和须芒草gerardii结构标记代表结果进行了讨论。

设计一个连续和操作<sup&gt; 13</sup&gt; C和<sup&gt; 15</sup&gt;Ñ贴标商会统一或差分，代谢和结构，植物同位素标记

Tracing rare stable isotopes from plant material through the ecosystem provides the most sensitive information about ecosystem processes; from CO2 fluxes ...

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Research

JoVE Journal

Environment

Design and Operation of a Continuous 13C and 15N Labeling Chamber for Uniform or Differential, Metabolic and Structural, Plant Isotope Labeling

21.5K 次观看.  Colorado State University. 这个方法介绍了如何建立和运营一个连续13 C和15 N同位素标记室统一或差分植物组织标签。从代谢和须芒草gerardii结构标记代表结果进行了讨论。 

视频: Design and Operation of a Continuous 13C and 15N Labeling Chamber for Uniform or Differential, Metabolic and Structural, Plant Isotope Labeling

A Rapid and Efficient Method for Assessing Pathogenicity of Ustilago maydis on Maize and Teosinte Lines

Maize is a major cereal crop worldwide. However, susceptibility to biotrophic pathogens is the primary constraint to increasing productivity. U. maydis is a biotrophic fungal pathogen and the causal agent of corn smut on maize. This disease is responsible for significant yield losses of approximately $1.0 billion annually in the U.S.1 Several methods including crop rotation, fungicide application and seed treatments are currently used to control corn smut2. However, host resistance is the only practical method for managing corn smut. Identification of crop plants including maize, wheat, and rice that are resistant to various biotrophic pathogens has significantly decreased yield losses annually3-5. Therefore, the use of a pathogen inoculation method that efficiently and reproducibly delivers the pathogen in between the plant leaves, would facilitate the rapid identification of maize lines that are resistant to U. maydis. As, a first step toward indentifying maize lines that are resistant to U. maydis, a needle injection inoculation method and a resistance reaction screening method was utilized to inoculate maize, teosinte, and maize x teosinte introgression lines with a U. maydis strain and to select resistant plants.
Maize, teosinte and maize x teosinte introgression lines, consisting of about 700 plants, were planted, inoculated with a strain of U. maydis, and screened for resistance. The inoculation and screening methods successfully identified three teosinte lines resistant to U. maydis. Here a detailed needle injection inoculation and resistance reaction screening protocol for maize, teosinte, and maize x teosinte introgression lines is presented. This study demonstrates that needle injection inoculation is an invaluable tool in agriculture that can efficiently deliver U. maydis in between the plant leaves and has provided plant lines that are resistant to U. maydis that can now be combined and tested in breeding programs for improved disease resistance.

A Rapid and Efficient Method for Assessing Pathogenicity of Ustilago maydis on Maize and Teosinte Lines

The use of a needle injection method to inoculate maize and teosinte plants with the biotrophic pathogen Ustilago maydis is described. The needle injection inoculation method facilitates the controlled delivery of the fungal pathogen in between the plant leaves where the pathogen enters the plant through the formation of appresoria. This method is highly efficient, enabling reproducible inoculations with U. maydis.

评估 乌西拉戈 病原性的快速高效方法，可在玉米和特奥辛特线上使用

Maize is a major cereal crop worldwide. However, susceptibility to biotrophic pathogens is the primary constraint to increasing productivity. U. maydis is ...

科研

环境科学

Transcript and Metabolite Profiling for the Evaluation of Tobacco Tree and Poplar as Feedstock for the Bio-based Industry

The global demand for food, feed, energy, and water poses extraordinary challenges for future generations. It is evident that robust platforms for the exploration of renewable resources are necessary to overcome these challenges. Within the multinational framework MultiBioPro we are developing biorefinery pipelines to maximize the use of plant biomass. More specifically, we use poplar and tobacco tree (Nicotiana glauca) as target crop species for improving saccharification, isoprenoid, long chain hydrocarbon contents, fiber quality, and suberin and lignin contents. The methods used to obtain these outputs include GC-MS, LC-MS and RNA sequencing platforms. The metabolite pipelines are well established tools to generate these types of data, but also have the limitations in that only well characterized metabolites can be used. The deep sequencing will allow us to include all transcripts present during the developmental stages of the tobacco tree leaf, but has to be mapped back to the sequence of Nicotiana tabacum. With these set-ups, we aim at a basic understanding for underlying processes and at establishing an industrial framework to exploit the outcomes. In a more long term perspective, we believe that data generated here will provide means for a sustainable biorefinery process using poplar and tobacco tree as raw material. To date the basal level of metabolites in the samples have been analyzed and the protocols utilized are provided in this article.

Plant biomass offers a renewable resource for multiple products, including fuel, feed, food, and a variety of materials. In this paper we investigate the properties of tobacco tree (Nicotiana glauca) and poplar as suitable sources for a biorefinery pipeline.

成绩单及代谢物谱烟草树和杨树的评价作为原料的生物基工业

The global demand for food, feed, energy, and water poses extraordinary challenges for future generations. It is evident that robust platforms for the ...

Design and Construction of an Urban Runoff Research Facility

As the urban population increases, so does the area of irrigated urban landscape. Summer water use in urban areas can be 2-3x winter base line water use due to increased demand for landscape irrigation. Improper irrigation practices and large rainfall events can result in runoff from urban landscapes which has potential to carry nutrients and sediments into local streams and lakes where they may contribute to eutrophication. A 1,000 m2 facility was constructed which consists of 24 individual 33.6 m2 field plots, each equipped for measuring total runoff volumes with time and collection of runoff subsamples at selected intervals for quantification of chemical constituents in the runoff water from simulated urban landscapes. Runoff volumes from the first and second trials had coefficient of variability (CV) values of 38.2 and 28.7%, respectively. CV values for runoff pH, EC, and Na concentration for both trials were all under 10%. Concentrations of DOC, TDN, DON, PO4-P, K+, Mg2+, and Ca2+ had CV values less than 50% in both trials. Overall, the results of testing performed after sod installation at the facility indicated good uniformity between plots for runoff volumes and chemical constituents. The large plot size is sufficient to include much of the natural variability and therefore provides better simulation of urban landscape ecosystems.

This paper describes the design, construction, and function of a 1,000 m2 facility containing 24 individual 33.6 m2 field plots equipped for measuring total runoff volumes with time and collection of runoff subsamples at selected intervals for quantification of chemical constituents in the runoff water from simulated home lawns.

设计一个城市径流研究设施的建设

As the urban population increases, so does the area of irrigated urban landscape. Summer water use in urban areas can be 2-3x winter base line water use ...

Operation of a 25 KWth Calcium Looping Pilot-plant with High Oxygen Concentrations in the Calciner

Calcium looping (CaL) is a post-combustion CO2 capture technology that is suitable for retrofitting existing power plants. The CaL process uses limestone as a cheap and readily available CO2 sorbent. While the technology has been widely studied, there are a few available options that could be applied to make it more economically viable. One of these is to increase the oxygen concentration in the calciner to reduce or eliminate the amount of recycled gas (CO2, H2O and impurities); therefore, decreasing or removing the energy necessary to heat the recycled gas stream. Moreover, there is a resulting increase in the energy input due to the change in the combustion intensity; this energy is used to enable the endothermic calcination reaction to occur in the absence of recycled flue gases. This paper presents the operation and first results of a CaL pilot plant with 100% oxygen combustion of natural gas in the calciner. The gas coming into the carbonator was a simulated flue gas from a coal-fired power plant or cement industry. Several limestone particle size distributions are also tested to further explore the effect of this parameter on the overall performance of this operating mode. The configuration of the reactor system, the operating procedures, and the results are described in detail in this paper. The reactor showed good hydrodynamic stability and stable CO2 capture, with capture efficiencies of up to 70% with a gas mixture simulating the flue gas of a coal-fired power plant.

Operation of a 25 KWth Calcium Looping Pilot-plant with High Oxygen Concentrations in the Calciner

This manuscript describes a procedure for operating a calcium looping pilot-plant for post-combustion carbon capture with high oxygen concentrations in the calciner in order to reduce or eliminate the flue gas recycle.

在煅烧炉中使用高氧浓度的 25 KWth钙循环试验装置的操作

Calcium looping (CaL) is a post-combustion CO2 capture technology that is suitable for retrofitting existing power plants. The CaL process uses limestone ...

Continuous Hydrologic and Water Quality Monitoring of Vernal Ponds

Vernal ponds, also referred to as vernal pools, provide critical ecosystem services and habitat for a variety of threatened and endangered species. However, they are vulnerable parts of the landscapes that are often poorly understood and understudied. Land use and management practices, as well as climate change are thought to be a contribution to the global amphibian decline. However, more research is needed to understand the extent of these impacts. Here, we present methodology for characterizing a vernal pond's morphology and detail a monitoring station that can be used to collect water quantity and quality data over the duration of a vernal pond's hydroperiod. We provide methodology for how to conduct field surveys to characterize the morphology and develop stage-storage curves for a vernal pond. Additionally, we provide methodology for monitoring the water level, temperature, pH, oxidation-reduction potential, dissolved oxygen, and electrical conductivity of water in a vernal pond, as well as monitoring rainfall data. This information can be used to better quantify the ecosystem services that vernal ponds provide and the impacts of anthropogenic activities on their ability to provide these services.

Understanding the ecosystem services and processes provided by vernal ponds and the impacts of anthropogenic activities on their ability to provide these services requires intensive hydrologic monitoring. This sampling protocol using in-situ monitoring equipment was developed to understand the impact of anthropogenic activities on water levels and quality.

春季池塘连续水文水质监测

Vernal ponds, also referred to as vernal pools, provide critical ecosystem services and habitat for a variety of threatened and endangered species. ...

The Plant Infection Test: Spray and Wound-Mediated Inoculation with the Plant Pathogen Magnaporthe Grisea

Plants possess a powerful system to defend themselves against potential threats by pathogenic fungi. For agriculturally important plants, however, current measures to combat such pathogens have proved too conservative and, thus, not sufficiently effective, and they can potentially pose environmental risks. Therefore, it is extremely necessary to identify host-resistance factors to assist in controlling plant diseases naturally through the identification of resistant germplasm, the isolation and characterization of resistance genes, and the molecular breeding of resistant cultivars. In this regard, there is need to establish an accurate, rapid, and large-scale inoculation method to breed and develop plant resistance genes. The rice blast fungal pathogen Magnaporthe grisea causes severe disease symptoms and yield losses. Recently, M. grisea has emerged as a model organism for studying the mechanisms of plant-fungal pathogen interactions. Hence, we report the development of a plant virulence test method that is specific for M. grisea. This method provides for both spray inoculation with a conidial suspension and wounding inoculation with mycelium cubes or droplets of conidial suspension. The key step of the wounding inoculation method for detached rice leaves is to make wounds on plant leaves, which avoids any interference caused by host penetration resistance. This spray/wounding protocol contributes to the rapid, accurate, and large-scale screening of the pathotypes of M. grisea isolates. This integrated and systematic plant infection method will serve as an excellent starting point for gaining a broad perspective of issues in plant pathology.

The Plant Infection Test: Spray and Wound-Mediated Inoculation with the Plant Pathogen Magnaporthe Grisea

Here, we present a protocol to test plant virulence with the plant pathogen Magnaporthe grisea. This report will contribute to the large-scale screening of the pathotypes of fungi isolates and serve as an excellent starting point for understanding the resistant mechanisms of plants during molecular breeding.

植物感染试验: 以植物病原菌为载体的喷雾和伤口介导接种

Plants possess a powerful system to defend themselves against potential threats by pathogenic fungi. For agriculturally important plants, however, current ...

Quantifying Plant Soluble Protein and Digestible Carbohydrate Content, Using Corn (Zea mays) As an Exemplar

Elemental data are commonly used to infer plant quality as a resource to herbivores. However, the ubiquity of carbon in biomolecules, the presence of nitrogen-containing plant defensive compounds, and variation in species-specific correlations between nitrogen and plant protein content all limit the accuracy of these inferences. Additionally, research focused on plant and/or herbivore physiology require a level of accuracy that is not achieved using generalized correlations. The methods presented here offer researchers a clear and rapid protocol for directly measuring plant soluble proteins and digestible carbohydrates, the two plant macronutrients most closely tied to animal physiological performance. The protocols combine well characterized colorimetric assays with optimized plant-specific digestion steps to provide precise and reproducible results. Our analyses of different sweet corn tissues show that these assays have the sensitivity to detect variation in plant soluble protein and digestible carbohydrate content across multiple spatial scales. These include between-plant differences across growing regions and plant species or varieties, as well as within-plant differences in tissue type and even positional differences within the same tissue. Combining soluble protein and digestible carbohydrate content with elemental data also has the potential to provide new opportunities in plant biology to connect plant mineral nutrition with plant physiological processes. These analyses also help generate the soluble protein and digestible carbohydrate data needed to study nutritional ecology, plant-herbivore interactions and food-web dynamics, which will in turn enhance physiology and ecological research.

Quantifying Plant Soluble Protein and Digestible Carbohydrate Content, Using Corn Zea mays As an Exemplar

The protocols described herein provide a clear and approachable methodology for measuring soluble protein and digestible (non-structural) carbohydrate content in plant tissues. The ability to quantify these two plant macronutrients has significant implications for advancing the fields of plant physiology, nutritional ecology, plant-herbivore interactions and food-web ecology.

以玉米 (玉蜀黍) 为例量化植物可溶性蛋白和可消化碳水化合物含量

Elemental data are commonly used to infer plant quality as a resource to herbivores. However, the ubiquity of carbon in biomolecules, the presence of ...

Colletotrichum fioriniae Development in Water and Chloroform-based Blueberry and Cranberry Floral Extracts

To accurately monitor the phenology of the bloom period and the temporal dynamics of floral chemical cues on fungal fruit rotting pathogens, floral extraction methods and coverslip bioassays were developed utilizing Colletotrichum fioriniae. In blueberry and cranberry, this pathogen is optimally controlled by applying fungicides during the bloom period because of the role flowers play in the initial stages of infection. The protocol detailed here describes how floral extracts (FE) were obtained using water-, chloroform-, and field rainwater-based methods for later use in corresponding glass coverslip bioassays. Each FE served to provide a different set of information: response of C. fioriniae to mobilized floral chemical cues in water (water-based), pathogen response to flower and fruit surface waxes (chloroform-based), and field-based monitoring of collected floral rainwater, moving in vitro observations to an agricultural setting. The FE is broadly described as either water- or chloroform-based, with an appropriate bioassay described to compensate for the inherent differences between these two materials. Rainwater that had run off flowers was collected in unique devices for each crop, alluding to the flexibility and application of this approach for other crop systems. The bioassays are quick, inexpensive, simple, and provide the ability to generate spatiotemporal and site-specific information about the presence of stimulatory floral compounds from various sources. This information will ultimately better inform disease management strategies, as FE decrease the time needed for infection to occur, thus providing insight into changing risks for pathogen infection over the growing season.

Colletotrichum fioriniae Development in Water and Chloroform-based Blueberry and Cranberry Floral Extracts

Here, bioassays designed to monitor the development of a fungal pathogen, Colletotrichum fioriniae, in the presence of blueberry or cranberry floral extracts on glass coverslips are described. Water-, chloroform-, and field rainwater- based floral extraction techniques are detailed as well as insight into how this information can be applied.

Fioriniae水和氯蓝莓和越橘花提取物的研究进展

To accurately monitor the phenology of the bloom period and the temporal dynamics of floral chemical cues on fungal fruit rotting pathogens, floral ...

A Uniaxial Compression Experiment with CO2-Bearing Coal Using a Visualized and Constant-Volume Gas-Solid Coupling Test System

Injecting carbon dioxide (CO2) into a deep coal seam is of great significance for reducing the concentration of greenhouse gases in the atmosphere and increasing the recovery of coalbed methane. A visualized and constant-volume gas-solid coupling system is introduced here to investigate the influence of CO2 sorption on the physical and mechanical properties of coal. Being able to keep a constant volume and monitor the sample using a camera, this system offers the potential to improve instrument accuracy and analyze fracture evolution with a fractal geometry method. This paper provides all steps to perform a uniaxial compression experiment with a briquette sample in different CO2 pressures with the gas-solid coupling test system. A briquette, cold-pressed by raw coal and sodium humate cement, is loaded in high-pressure CO2, and its surface is monitored in real-time using a camera. However, the similarity between the briquette and the raw coal still needs improvement, and a flammable gas such as methane (CH4) cannot be injected for the test. The results show that CO2 sorption leads to peak strength and elastic modulus reduction of the briquette, and the fracture evolution of the briquette in a failure state indicates fractal characteristics. The strength, elastic modulus, and fractal dimension are all correlated with CO2 pressure but not with a linear correlation. The visualized and constant-volume gas-solid coupling test system can serve as a platform for experimental research about rock mechanics considering the multifield coupling effect.

A Uniaxial Compression Experiment with CO2-Bearing Coal Using a Visualized and Constant-Volume Gas-Solid Coupling Test System

This protocol demonstrates how to prepare a briquette sample and conduct a uniaxial compression experiment with a briquette in different CO2 pressures using a visualized and constant-volume gas-solid coupling test system. It also aims to investigate changes in terms of coal&#8217;s physical and mechanical properties induced by CO2 adsorption.

使用可视化和恒定体积气体-固体耦合测试系统使用 CO2轴承煤的单轴压缩实验

Injecting carbon dioxide (CO2) into a deep coal seam is of great significance for reducing the concentration of greenhouse gases in the atmosphere and ...

PARbars: Cheap, Easy to Build Ceptometers for Continuous Measurement of Light Interception in Plant Canopies

Ceptometry is a technique used to measure the transmittance of photosynthetically active radiation through a plant canopy using multiple light sensors connected in parallel on a long bar. Ceptometry is often used to infer properties of canopy structure and light interception, notably leaf area index (LAI) and effective plant area index (PAIeff). Due to the high cost of commercially available ceptometers, the number of measurements that can be taken is often limited in space and time. This limits the usefulness of ceptometry for studying genetic variability in light interception, and precludes thorough analysis of, and correction for, biases that can skew measurements depending on the time of day. We developed continuously logging ceptometers (called PARbars) that can be produced for USD $75 each and yield high quality data comparable to commercially available alternatives. Here we provide detailed instruction on how to build and calibrate PARbars, how to deploy them in the field and how to estimate PAI from collected transmittance data. We provide representative results from wheat canopies and discuss further considerations that should be made when using PARbars.

Here, we present detailed instructions on how to build and calibrate research quality ceptometers (light sensors that integrate light intensity across many sensors arrayed linearly along a horizontal bar).

Parbar: 便宜、易于制造的用于连续测量植物顶篷中的光拦截的证书台

Ceptometry is a technique used to measure the transmittance of photosynthetically active radiation through a plant canopy using multiple light sensors ...