该研究是由奥地利科学基金会（FWF）:P23681-B16。

1.选煤厂材料<ol><li>使用主体植物材料的任叶或整株植物（ 玉簪属或菜豆 ）。 </li></ol> 2.准备水处理解决方案含冰核活性（INA）细菌<ol><li>培养INA的细菌， 丁香假单胞菌 （菌株CIT-7）在培养皿中，在25℃对铜绿琼脂˚F制备按照生产商的方向100％的甘油的10克/升。 </li><li>后培养已经长大充分，发生在4℃直至需要的，但保持在4℃下进行，以确保冰核活性的高水平之前两天。 </li><li>刮由单一的板的细菌从琼脂表面用塑料，一次性或可重复使用的金属刮刀在使用和地点的时间在10〜15毫升去离子水在25毫升一次性杯。的浓度应为1×10 7至1×10 9的范围· </sup&gt;毫升-1。该解决方案将出现混浊。没有必要确认使用血球或分光光度计的浓度，如浓度仅需要近似的。 </li><li>涡试管最少10秒的分发细菌。 注意:得到的INA混合物的具体浓度是不重要的，描述的协议将提供超过冰核活性的适当水平。 INA细菌和水的混合物将在核试验以后使用。 </li></ol> 3.设置冷冻实验<ol><li>放置高分辨率红外摄像机（SC-660）的保护性丙烯酸盒内以便通过开口在盒前面的透镜的项目，并通过盒的后部开口将相机连接到笔记本电脑或记录装置出口处的线。固定框的盖板，将箱中的位置的环境室或冷冻室里面的那个将所有流待观察对象的植物材料。 <ol><li>由衬的腔室的壁与黑色施工纸，以防止反射的红外能量干扰提供围绕植物材料在暗背景。 </li><li>适合室LED照明时可见光波长记录图像，需要从光源尽量减少发热。只有最小的照明，如电池供电的壁橱光或其他小的LED装置中，是所必需的植物是由相机可见。 <ol><li>一旦受试者植物材料的可见图像被拍摄时，关闭LED照明。通过端口或其他室内分布开放所有外部有线连接（FireWire连接到电脑，电源线等）连接到摄像机。 </li></ol></li><li>填入口或开口的任何额外的空间用绝缘泡沫材料，以避免或减少在腔室中的温度梯度。在1°C的设置室的初始温度。 </li></ol></li><li>对齐植物或植物部分，以使植物材料是在磁场 - 视摄像机和植物材料是可见的远程观看屏幕上或所选择的软件内。 </li><li>允许植物平衡在1℃下进行30分钟至1小时，这取决于植物材料的大小之前，发起受控冷冻试验。这保证了装置的温度不会被许多度落后气温一旦冷冻实验被启动。当所述植物材料的温度在0.5℃的空气温度的平衡得以实现。 <ol><li>放置一层泡沫塑料绝缘对盆栽土壤的顶部，如果盆栽被使用。一旦植物已经平衡，开始该腔室的冷却。 注意:绝缘锅的土壤表面上的层减小了从锅到植物周围的空气继续热损耗量，并且防止了树根从FREezing，因为这将不通常在自然界中结霜事件的发生是由于存在于土壤余热大规模贮存。 </li></ol></li><li>设置所需的照相机参数（调色板，温度范围，感兴趣的特定区域等 ），如在3.4.1-3.4.4讨论。 <ol><li>选择调色板彩虹同时观看实时图像显示温度的变化。 </li><li>通过调整位于正好在软件的图像下方的温度栏设定温度跨度至5℃。 </li><li>选择线性标尺（算法），用于将所述红外数据到假彩色图像由所选择的调色板（彩虹）定义的与设定温度的范围内以5℃，并以基于在图像上自动跟踪。另外，手动调整设定的范围，同时进行了实验。 <ol><li>使用特定的点或温度的平均温度的利益由软件提供的规定区域内的是。检索所记录的视频序列或从嵌入在图像文件中的信息的所有像素的温度数据。 图3示出了一个典型的屏幕截图从ResearchIR软件内。 </li></ol></li><li>将光标置于上的植物组织，代表感兴趣的特定点的位置。定义兴趣点（大小1-3像素），框，线条，椭圆，或圆的面积。点或形状的多种组合可以位于​​在图像上。 </li></ol></li><li>记录视频序列<ol><li>将相机设定为60赫兹和录音手动停止录制。 </li><li>指示计算机或外部驱动器，其中所记录的视频文件将被放置的位置。 </li><li>开始录制。 注:录制到外部硬盘驱动器，强烈推荐，因为大的视频文件将被生成。录制的视频文件可以在以后修改为仅包含含NE的部分cessary信息。这将大大降低文件大小。 </li><li> 0.5 -1.0℃的降低增量腔室的温度。等到植物温度达到平衡与空气的温度，然后通过0.5-1.0℃下再次降低温度。取决于植物组织被观察和它的形态的质量，平衡可能需要10至15分钟。因此，给予约4℃/小时的冷却速度。 </li><li>继续以这种方式，直到工厂冻结和提出的意见完成。结束记录时在冷冻过程已经完成。 注意:植物组织已经平衡的空气温度时，植物材料和背景是相同的颜色，因为它们是在相同的温度。由于背景温度和植物组织的温度是相同的，也可能是很难看到的植物材料，直到再次降低温度并且在植物组织和一之间的温度差IR温度。 </li></ol></li></ol>

<ol>
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作者有没有竞争经济利益或利益冲突。

水具有过冷到温度低于0℃，并在该水会冻结可以相当可变温度的能力。36的温度限制为纯水过冷是约-40℃，并且被定义为均相成核点。当水结冰的温度比回暖-40°C它带来的异质存在成核剂，使小冰的胚胎形成，然后作为催化剂，冰的形成和生长。37有众多的自然分子的充当非常有效的冰成核剂，因此大多数的性质的冷冻水发生在温度刚好低于0℃。调节或影响异质成核剂的活性的能...

冷冻时植物生长活跃时发生的温度可以是致命的，特别是如果植物具有很少或没有冰冻耐受性。这样的事件霜冻往往会对农业生产造成破坏性影响，也可以在植物自然种群形成群落结构发挥了重要作用，特别是在高山，亚寒带和寒带生态系统1-6。严重的春季霜冻事件在最近几年7-9对水果生产在美国和南美洲的重大影响，并加剧了发病初期温暖的天气其次是较典型的平均气温低。早期的温暖的天气诱使芽突破，激活新枝，叶和花所有这些都很少无霜容忍1,3,10-12增长。这种变化无常的天气模式已经被报道正在发生的气候变化的直接反映，并预计将在FORES一个共同的天气模式eeable未来的13。努力提供经济，有效和环境友好的管理技术或农用化学品，可以提供增加的冰冻耐受性取得了有限的成功为宿主的原因，但是这可以部分地归因于耐冻性和冰冻避免机制在植物的复杂性。 14 霜存活于植物相关联的自适应机制已经传统上被分为两类，耐冻性和冰冻回避。前一类与由一组特定的基因是允许植物容忍与存在和冰在其组织的脱水作用有关的应力的调节生化机制相关联。而在后一类通常是，但不是唯一，与植物中的植物14结构方面确定是否，何时，何地冰形成有关。尽管冻结回避的患病率作为广告aptive机制，一些研究一直致力于在近期了解的底层机制和冻结规避监管。读者可以参考最近15检讨关于这个主题的更详细。 而形成的冰在低温下可能看起来像一个简单的过程中，许多因素有助于确定在哪些冰成核在植物组织和它如何在植物内扩散的温度。如外在和内在冰的存在下成核，异质与均相成核事件，热滞后（防冻剂）的蛋白质，具体糖和其它渗透剂的存在下，和一台主机的植物的结构方面参数都可以发挥显著在植物冷冻过程中的作用。总的来说，这些参数影响在哪个工厂结冰，冰的地方开始以及如何成长的温度。它们也可以影响所得冰晶体的形态。各种方法已被用于研究在植物冷冻过程在实验室条件，包括核磁共振谱（NMR）16，磁共振成像（MRI）17，冷冻显微镜18-19，和低温扫描电子显微镜下（LTSEM ）。20冻结在实验室和现场设置整株植物，然而，主要被监测热电偶。使用热电偶来研究冷冻的是基于热解放（熔化焓）当水经历从液体到固体的相变。冷冻，然后被记录作为一个放热事件。21-23即使热电偶是选择在研究中的植物冷冻的典型方法，它们的使用有很大的局限性，限制了冷冻事件期间所获得的信息的量。例如，用热电偶就很难几乎不可能确定在何处开始在植物中的冰，它是如何传播，如果它在传播的速度，甚至，如果某些组织保持无冰。 高分辨率红外热（HRIT）24-27进展，然而，有显著增加，以获取有关整个植物冷冻处理信息的能力，在差分成像模式下使用时尤其如此。28-33在本报告中，我们描述了使用该技术的研究冷冻过程的各个方面，该影响在哪里和以什么样的温度冰在植物中被启动的各种参数。的协议将被呈现，将展示冰核活性（INA）细菌作为一个非本征成核启动在高温，零下温度在一种草本植物冷冻能力， 丁香假单胞菌 （CIT-7）。 高分辨率红外摄像机该协议和实例本报告中采用红外线高分辨率视频辐射计。所述辐射计（ 图1）提供的红外和可见光谱图像和温度数据的组合。照相机的光谱响应是在7.5至13.5微米的范围内，并提供640×480像素的分辨率。所产生的可见光谱图像的内置相机可以与红外图像进行实时，这有利于复杂，热图像的解释是稠合的。透镜用于摄像机A范围可以被用来制造特写和显微镜观察。该相机可以在一个独立的模式下使用，或接口和控制使用的专有软件的笔记本电脑。该软件可以被用来获得各种嵌入在录制的视频热数据。要注意的是各种各样的红外线辐射计是市售的是重要的。因此，至关重要的是，研究者讨论一个知识渊博的产品工程师其预期应用和研究人员测试任何specifi的能力ç辐射计，以提供所需的信息。在所描述的协议中使用的成象辐射计被放置在丙烯酸类盒（ 图2）绝缘的聚苯乙烯泡沫塑料I N为了在升温和冷却的协议，以防止暴露于冷凝。无需对所有摄像机或应用这种保护。

<table><tbody><tr><td>Infrared Camera</td><td>FLIR</td><td>SC-660</td><td>Many models available depending on application</td></tr><tr><td>Infrared Analytical Software</td><td>FLIR</td><td>ResearchIR 4.10.2.5</td><td>$3,500</td></tr><tr><td>Pseudomonas syringae (strain Cit-7)</td><td></td><td></td><td>&lt;a href=&quot;mailto:icelab@berkeley.edu&quot;&gt;Kindly provided by Dr. Steven Lindow, University of California&amp;nbsp; Berkeley icelab@berkeley.edu&lt;/a&gt;</td></tr><tr><td>Pseudomonas Agar F</td><td>Fisher Scientific</td><td>DF0448-17-1</td><td></td></tr></tbody></table>

the use of high-resolution infrared thermography (hrit) for the study of ice nucleation and ice propagation in plants

Freezing events that occur when plants are actively growing can be a lethal event, particularly if the plant has no freezing tolerance. Such frost events often have devastating effects on agricultural production and can also play an important role in shaping community structure in natural populations of plants, especially in alpine, sub-arctic, and arctic ecosystems. Therefore, a better understanding of the freezing process in plants can play an important role in the development of methods of frost protection and understanding mechanisms of freeze avoidance. Here, we describe a protocol to visualize the freezing process in plants using high-resolution infrared thermography (HRIT). The use of this technology allows one to determine the primary sites of ice formation in plants, how ice propagates, and the presence of ice barriers. Furthermore, it allows one to examine the role of extrinsic and intrinsic nucleators in determining the temperature at which plants freeze and evaluate the ability of various compounds to either affect the freezing process or increase freezing tolerance. The use of HRIT allows one to visualize the many adaptations that have evolved in plants, which directly or indirectly impact the freezing process and ultimately enables plants to survive frost events.

冰+细菌的冰核活动， 丁香假单胞菌 （株CIT-7） 水的10μl降和10微升水含有P.丁香 （CIT-7）被放置于一玉簪叶的背面（ 玉簪属）（ 图4）。如图所示，对含有冰核细菌水滴冻结第一和负​​责诱导叶冻结而水在叶子表面上的压降保持解冻。 冻结和冰繁殖的木本植物 图5示出冰都开始和冰繁殖...

Watch this Scientific Journal Video about The Use of High-resolution Infrared Thermography HRIT for the Study of Ice Nucleation and Ice Propagation in Plants at JoVE.com

The Use of High-resolution Infrared Thermography HRIT for the Study of Ice Nucleation and Ice Propagation in Plants

Here we present a protocol that allows one to visualize sites of ice formation and avenues of ice propagation in plants utilizing high resolution infrared thermography (HRIT).

高分辨率红外热成像（HRIT）的冰核和冰传播的植物研究中的应用

Freezing events that occur when plants are actively growing can be a lethal event, particularly if the plant has no freezing tolerance. Such frost events ...

the-use-high-resolution-infrared-thermography-hrit-for-study-ice

Universidad de Cartagena UDEC

Research

JoVE Journal

Environment

The Use of High-resolution Infrared Thermography (HRIT) for the Study of Ice Nucleation and Ice Propagation in Plants

9.5K 次观看.  Agricultural Research Service (USDA-ARS), Kearneysville, WV. Here we present a protocol that allows one to visualize sites of ice formation and avenues of ice propagation in plants utilizing high resolution infrared thermography (HRIT).

视频: The Use of High-resolution Infrared Thermography HRIT for the Study of Ice Nucleation and Ice Propagation in Plants

A Rapid Laser Probing Method Facilitates the Non-invasive and Contact-free Determination of Leaf Thermal Properties

Plants can produce valuable substances such as secondary metabolites and recombinant proteins. The purification of the latter from plant biomass can be streamlined by heat treatment (blanching). A blanching apparatus can be designed more precisely if the thermal properties of the leaves are known in detail, i.e., the specific heat capacity and thermal conductivity. The measurement of these properties is time consuming and labor intensive, and usually requires invasive methods that contact the sample directly. This can reduce the product yield and may be incompatible with containment requirements, e.g., in the context of good manufacturing practice. To address these issues, a non-invasive, contact-free method was developed that determines the specific heat capacity and thermal conductivity of an intact plant leaf in about one minute. The method involves the application of a short laser pulse of defined length and intensity to a small area of the leaf sample, causing a temperature increase that is measured using a near infrared sensor. The temperature increase is combined with known leaf properties (thickness and density) to determine the specific heat capacity. The thermal conductivity is then calculated based on the profile of the subsequent temperature decline, taking thermal radiation and convective heat transfer into account. The associated calculations and critical aspects of sample handling are discussed.

A method was developed to determine the specific heat capacity and thermal conductivity of leaf tissue by non-invasive, contact-free near infrared laser probing, which requires less than 1 min per sample.

一种快速探测激光方法有利于叶热性能的非侵入性和无接触测定

Plants can produce valuable substances such as secondary metabolites and recombinant proteins. The purification of the latter from plant biomass can be ...

科研

生物化学

Identification of Plant Ice-binding Proteins Through Assessment of Ice-recrystallization Inhibition and Isolation Using Ice-affinity Purification

Ice-binding proteins (IBPs) belong to a family of stress-induced proteins that are synthesized by certain organisms exposed to subzero temperatures. In plants, freeze damage occurs when extracellular ice crystals grow, resulting in the rupture of plasma membranes and possible cell death. Adsorption of IBPs to ice crystals restricts further growth by a process known as ice-recrystallization inhibition (IRI), thereby reducing cellular damage. IBPs also demonstrate the ability to depress the freezing point of a solution below the equilibrium melting point, a property known as thermal hysteresis (TH) activity. These protective properties have raised interest in the identification of novel IBPs due to their potential use in industrial, medical and agricultural applications. This paper describes the identification of plant IBPs through 1) the induction and extraction of IBPs in plant tissue, 2) the screening of extracts for IRI activity, and 3) the isolation and purification of IBPs. Following the induction of IBPs by low temperature exposure, extracts are tested for IRI activity using a &#39;splat assay&#39;, which allows the observation of ice crystal growth using a standard light microscope. This assay requires a low protein concentration and generates results that are quickly obtained and easily interpreted, providing an initial screen for ice binding activity. IBPs can then be isolated from contaminating proteins by utilizing the property of IBPs to adsorb to ice, through a technique called &#39;ice-affinity purification&#39;. Using cell lysates collected from plant extracts, an ice hemisphere can be slowly grown on a brass probe. This incorporates IBPs into the crystalline structure of the polycrystalline ice. Requiring no a priori biochemical or structural knowledge of the IBP, this method allows for recovery of active protein. Ice-purified protein fractions can be used for downstream applications including the identification of peptide sequences by mass spectrometry and the biochemical analysis of native proteins.

This paper outlines the identification of ice-binding proteins from freeze-tolerant plants through the assessment of ice-recrystallization inhibition activity and subsequent isolation of native IBPs using ice-affinity purification.

植物冰结合蛋白通过冰再结晶抑制和隔离的评估使用冰亲和力纯化鉴定

Ice-binding proteins (IBPs) belong to a family of stress-induced proteins that are synthesized by certain organisms exposed to subzero temperatures. In ...

RGB and Spectral Root Imaging for Plant Phenotyping and Physiological Research: Experimental Setup and Imaging Protocols

Better understanding of plant root dynamics is essential to improve resource use efficiency of agricultural systems and increase the resistance of crop cultivars against environmental stresses. An experimental protocol is presented for RGB and hyperspectral imaging of root systems. The approach uses rhizoboxes where plants grow in natural soil over a longer time to observe fully developed root systems. Experimental settings are exemplified for assessing rhizobox plants under water stress and studying the role of roots. An RGB imaging setup is described for cheap and quick quantification of root development over time. Hyperspectral imaging improves root segmentation from the soil background compared to RGB color based thresholding. The particular strength of hyperspectral imaging is the acquisition of chemometric information on the root-soil system for functional understanding. This is demonstrated with high resolution water content mapping. Spectral imaging however is more complex in image acquisition, processing and analysis compared to the RGB approach. A combination of both methods can optimize a comprehensive assessment of the root system. Application examples integrating root and aboveground traits are given for the context of plant phenotyping and plant physiological research. Further improvement of root imaging can be obtained by optimizing RGB image quality with better illumination using different light sources and by extension of image analysis methods to infer on root zone properties from spectral data.

An experimental protocol is presented for assessment of soil grown plant root systems with RGB and hyperspectral imaging. Combination of RGB image time series with chemometric information from hyperspectral scans optimizes insights into plant root dynamics.

RGB 和光谱根成像植物表型和生理研究: 实验装置和成像协议

Better understanding of plant root dynamics is essential to improve resource use efficiency of agricultural systems and increase the resistance of crop ...

环境科学

Protocol for Producing Three-Dimensional Infrared Video of Freezing in Plants

Freezing in plants can be monitored using infrared (IR) thermography, because when water freezes, it gives off heat. However, problems with color contrast make 2-dimensions (2D) infrared images somewhat difficult to interpret. Viewing an IR image or the video of plants freezing in 3 dimensions (3D) would allow a more accurate identification of sites for ice nucleation as well as the progression of freezing. In this paper, we demonstrate a relatively simple means to produce a 3D infrared video of a strawberry plant freezing. Strawberry is an economically important crop that is subjected to unexpected spring freeze events in many areas of the world. An accurate understanding of the freezing in strawberry will provide both breeders and growers with more economical ways to prevent any damage to plants during freezing conditions.
The technique involves a positioning of two IR cameras at slightly different angles to film the strawberry freezing. The two video streams will be precisely synchronized using a screen capture software that records both cameras simultaneously. The recordings will then be imported into the imaging software and processed using an anaglyph technique. Using red-blue glasses, the 3D video will make it easier to determine the precise site of ice nucleation on leaf surfaces.

Here, we present a protocol to image a strawberry plant freezing in 3 dimensions. Two infrared cameras positioned at slightly different angles are used to produce a red-blue anaglyph video to observe the freezing of the plant in 3 dimensions.

植物冷冻三维红外视频制作协议

Freezing in plants can be monitored using infrared (IR) thermography, because when water freezes, it gives off heat. However, problems with color contrast ...

Simulating Impacts of Ice Storms on Forest Ecosystems

Ice storms can have profound and lasting effects on the structure and function of forest ecosystems in regions that experience freezing conditions. Current models suggest that the frequency and intensity of ice storms could increase over the coming decades in response to changes in climate, heightening interest in understanding their impacts. Because of the stochastic nature of ice storms and difficulties in predicting when and where they will occur, most past investigations of the ecological effects of ice storms have been based on case studies following major storms. Since intense ice storms are exceedingly rare events it is impractical to study them by waiting for their natural occurrence. Here we present a novel alternative experimental approach, involving the simulation of glaze ice events on forest plots under field conditions. With this method, water is pumped from a stream or lake and sprayed above the forest canopy when air temperatures are below freezing. The water rains down and freezes upon contact with cold surfaces. As the ice accumulates on trees, the boles and branches bend and break; damage that can be quantified through comparisons with untreated reference stands. The experimental approach described is advantageous because it enables control over the timing and amount of ice applied. Creating ice storms of different frequency and intensity makes it possible to identify critical ecological thresholds necessary for predicting and preparing for ice storm impacts.

Ice storms are important weather events that are challenging to study because of difficulties in predicting their occurrence. Here, we describe a novel method for simulating ice storms that involves spraying water over a forest canopy during sub-freezing conditions.

模拟冰风暴对森林生态系统的影响

Ice storms can have profound and lasting effects on the structure and function of forest ecosystems in regions that experience freezing conditions. ...

Near-Infrared Temperature Measurement Technique for Water Surrounding an Induction-heated Small Magnetic Sphere

A technique to measure the temperature of water and non-turbid aqueous media surrounding an induction-heated small magnetic sphere is presented. This technique utilizes wavelengths of 1150 and 1412 nm, at which the absorption coefficient of water is dependent on temperature. Water or a non-turbid aqueous gel containing a 2.0-mm- or 0.5-mm-diameter magnetic sphere is irradiated with 1150 nm or 1412 nm incident light, as selected using a narrow bandpass filter; additionally, two-dimensional absorbance images, which are the transverse projections of the absorption coefficient, are acquired via a near-infrared camera. When the three-dimensional distributions of temperature can be assumed to be spherically symmetric, they are estimated by applying inverse Abel transforms to the absorbance profiles. The temperatures were observed to consistently change according to time and the induction heating power.

A technique utilizing wavelengths of 1150 and 1412 nm to measure the temperature of water surrounding an induction-heated small magnetic sphere is presented.

感应加热小磁球周围水近红外测温技术

A technique to measure the temperature of water and non-turbid aqueous media surrounding an induction-heated small magnetic sphere is presented. This ...

工程学

Ice-Cap: A Method for Growing Arabidopsis and Tomato Plants in 96-well Plates for High-Throughput Genotyping

It is becoming common for plant scientists to develop projects that require the genotyping of large numbers of plants. The first step in any genotyping project is to collect a tissue sample from each individual plant. The traditional approach to this task is to sample plants one-at-a-time. If one wishes to genotype hundreds or thousands of individuals, however, using this strategy results in a significant bottleneck in the genotyping pipeline. The Ice-Cap method that we describe here provides a high-throughput solution to this challenge by allowing one scientist to collect tissue from several thousand seedlings in a single day 1,2. This level of throughput is made possible by the fact that tissue is harvested from plants 96-at-a-time, rather than one-at-a-time.
The Ice-Cap method provides an integrated platform for performing seedling growth, tissue harvest, and DNA extraction. The basis for Ice-Cap is the growth of seedlings in a stacked pair of 96-well plates. The wells of the upper plate contain plugs of agar growth media on which individual seedlings germinate. The roots grow down through the agar media, exit the upper plate through a hole, and pass into a lower plate containing water. To harvest tissue for DNA extraction, the water in the lower plate containing root tissue is rapidly frozen while the seedlings in the upper plate remain at room temperature. The upper plate is then peeled away from the lower plate, yielding one plate with 96 root tissue samples frozen in ice and one plate with 96 viable seedlings. The technique is named "Ice-Cap" because it uses ice to capture the root tissue. The 96-well plate containing the seedlings can then wrapped in foil and transferred to low temperature. This process suspends further growth of the seedlings, but does not affect their viability. Once genotype analysis has been completed, seedlings with the desired genotype can be transferred from the 96-well plate to soil for further propagation. We have demonstrated the utility of the Ice-Cap method using Arabidopsis thaliana, tomato, and rice seedlings. We expect that the method should also be applicable to other species of plants with seeds small enough to fit into the wells of 96-well plates.

Ice-Cap: A Method for Growing Arabidopsis and Tomato Plants in 96-well Plates for High-Throughput Genotyping

The Ice-Cap method allows one to grow plants in 96-well plates and non-destructively harvest root tissue from each seedling. DNA extracted from this root tissue can be used for genotyping reactions. We have found that Ice-Cap works well for Arabidopsis thaliana, tomato, and rice seedlings.

冰帽：为发展方法拟南芥和番茄植株在96孔板高通量的基因分型

It is becoming common for plant scientists to develop projects that require the genotyping of large numbers of plants. The first step in any genotyping ...

生物学

Using Flatbed Scanners to Collect High-resolution Time-lapsed Images of the Arabidopsis Root Gravitropic Response

Research efforts in biology increasingly require use of methodologies that enable high-volume collection of high-resolution data. A challenge laboratories can face is the development and attainment of these methods. Observation of phenotypes in a process of interest is a typical objective of research labs studying gene function and this is often achieved through image capture. A particular process that is amenable to observation using imaging approaches is the corrective growth of a seedling root that has been displaced from alignment with the gravity vector. Imaging platforms used to measure the root gravitropic response can be expensive, relatively low in throughput, and/or labor intensive. These issues have been addressed by developing a high-throughput image capture method using inexpensive, yet high-resolution, flatbed scanners. Using this method, images can be captured every few minutes at 4,800 dpi. The current setup enables collection of 216 individual responses per day. The image data collected is of ample quality for image analysis applications.

This protocol describes a process for rapid collection of images of Arabidopsis seedlings responding to a gravity stimulus using commercially-available flatbed scanners. The method allows for inexpensive, high-volume capture of high-resolution images amenable for downstream analysis algorithms.

使用平板扫描仪，以收集拟南芥根向地反应的高分辨率时间失效图片

Research efforts in biology increasingly require use of methodologies that enable high-volume collection of high-resolution data. A challenge laboratories ...

Identification of Novel Regulators of Plant Transpiration by Large-Scale Thermal Imaging Screening in Helianthus Annuus

Plant adaptation to biotic and abiotic stresses is governed by a variety of factors, among which the regulation of stomatal aperture in response to water deficit or pathogens plays a crucial role. Identifying small molecules that regulate stomatal movement can therefore contribute to understanding the physiological basis by which plants adapt to their environment. Large-scale screening approaches that have been used to identify regulators of stomatal movement have potential limitations: some rely heavily on the abscisic acid (ABA) hormone signaling pathway, therefore excluding ABA-independent mechanisms, while others rely on the observation of indirect, long-term physiological effects such as plant growth and development. The screening method presented here allows the large-scale treatment of plants with a library of chemicals coupled with a direct quantification of their transpiration by thermal imaging. Since evaporation of water through transpiration results in leaf surface cooling, thermal imaging provides a non-invasive approach to investigate changes in stomatal conductance over time. In this protocol, Helianthus annuus seedlings are grown hydroponically and then treated by root feeding, in which the primary root is cut and dipped into the chemical being tested. Thermal imaging followed by statistical analysis of cotyledonary temperature changes over time allows for the identification of bioactive molecules modulating stomatal aperture. Our proof-of-concept experiments demonstrate that a chemical can be carried from the cut root to the cotyledon of the sunflower seedling within 10 minutes. In addition, when plants are treated with ABA as a positive control, an increase in leaf surface temperature can be detected within minutes. Our method thus allows the efficient and rapid identification of novel molecules regulating stomatal aperture.

Identification of Novel Regulators of Plant Transpiration by Large-Scale Thermal Imaging Screening in Helianthus Annuus

We provide a method for identifying modulators of foliar transpiration by large-scale screening of a compound library.

赫连图斯安努斯大型热成像筛选装置新稳压器的鉴定

Plant adaptation to biotic and abiotic stresses is governed by a variety of factors, among which the regulation of stomatal aperture in response to water ...

拟南芥幼苗中叶绿素生物合成的高分辨率成像

Non-invasive Assay for Chlorophyll Biosynthesis Kinetics Determination during Early Stages of Arabidopsis De-etiolation

Chlorophyll biosynthesis is a hallmark of de-etiolation, one of the most dramatic stages in the plant life cycle. The tightly controlled and highly dynamic process of chlorophyll biosynthesis is triggered during the shift from the dark to the light in flowering plants. At the moment when etiolated seedlings are exposed to the first traces of sunlight, rapid (in order of seconds) conversion of protochlorophyllide into chlorophyllide is mediated by unique light-accepting protein complexes, leading via subsequent metabolic steps to the production of fully functional chlorophyll. Standard techniques for chlorophyll content analysis include pigment extraction from detached plant tissues, which does not apply to studying such fast processes. To investigate chlorophyll kinetics in vivo with high accuracy and spatiotemporal resolution in the first hours after light-induced de-etiolation, an instrument and protocol were developed. Here, we present a detailed procedure designed for statistically robust quantification of chlorophyll in the early stages of Arabidopsis de-etiolation.

作者聚焦:去硫醇化过程中叶绿素合成的无创高分辨率测量

Here, we describe an advanced tool designed for chlorophyll biosynthesis monitoring during the early stages of Arabidopsis seedling de-etiolation. The novel methodology provides non-invasive real-time chlorophyll fluorescence imaging at high spatial and temporal resolution.

拟南芥脱硫早期叶绿素生物合成动力学测定的非侵入性测定

Chlorophyll biosynthesis is a hallmark of de-etiolation, one of the most dramatic stages in the plant life cycle. The tightly controlled and highly ...