Name: surface renewal: an advanced micrometeorological method for measuring and processing field-scale energy flux density data
Uploaded: 2013-12-12T08:45:00
Description: Watch this Scientific Journal Video about Surface Renewal: An Advanced Micrometeorological Method for Measuring and Processing Field-Scale Energy Flux Density Data at JoVE.com

Partial support for this research was provided by J. Lohr Vineyards &amp; Wines, the National Grape and Wine Institute, a NIFA Specialty Crops Research Initiative grant to AJM, and USDA-ARS CRIS funding (Research Project #5306-21220-004-00).

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	<li>Perform a wind direction and footprint analysis prior to field installation; this step is important to insure that the tower is situated properly to capture the representative flux in the study field.
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		<li>Before establishing the field station, you must evaluate the prevailing wind direction in the study plot, and determine how much fetch (upwind distance) is needed for your particular application. The distance of the fetch is dependent on the height of the surface to be measured and the height of the...

<ol>
	<li>Moratiel, R., Mart&#237;nez-Cob, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evapotranspiration+of+table+grape+trained+to+a+gable+trellis+system+under+netting+and+black+plastic+mulching.">Evapotranspiration of table grape trained to a gable trellis system under netting and black plastic mulching.</a> Irrigation Science. 30 (3), 167-178 (2011).</li><li>DWR (Department of Water Resources). <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> California Water Plan Update, Bulletin 160-05. , (2005).</li><li>Snyder, R. L., Spano, D., Duce, P., Paw, U. K. T., Qiu, J., Su, H. B., Watanabe, T., Brunet, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Surface+renewal+analysis:+a+new+method+to+obtain+scalar+fluxes+without+velocity+data.">Surface renewal analysis: a new method to obtain scalar fluxes without velocity data.</a> Agric. For. Meteorol. 74, 119-137 (1995).</li><li>Rosenberg, N. J., Blad, B. L., Verma, S. 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> Microclimate: the biological environment. , (1983).</li><li>Snyder, R. L., Spano, D., Duce, P., Paw, U. K. T., Anderson, F. E., Falk, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Surface Renewal Manual. , (2007).</li><li>Shapland, T. M., McElrone, A. J., Paw, U. T. K., Snyder, R. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+turnkey+data+logger+program+for+field-scale+energy+flux+density+measurements+using+eddy+covariance+and+surface.">A turnkey data logger program for field-scale energy flux density measurements using eddy covariance and surface.</a> Ital. J. Agrometeorol. , .</li><li>Shapland, T. M., Snyder, R. L., Paw, U. K. T., Lang, D., McElrone, A. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=New+tools+to+improve+access+to+micrometeorological+methods+for+agricultural+research+scientists.">New tools to improve access to micrometeorological methods for agricultural research scientists.</a> , (2011).</li><li>Van Atta, C. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+coherent+structures+on+structure+functions+of+temperature+in+the+atmospheric+boundary+layer.">Effect of coherent structures on structure functions of temperature in the atmospheric boundary layer.</a> Arch. of Mech. 29, 161-171 (1977).</li><li>Snyder, R. L., Spano, D., Paw, U. K. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Surface+Renewal+analysis+for+sensible+and+latent+heat+flux+density.">Surface Renewal analysis for sensible and latent heat flux density.</a> Boundary-Layer Meteorol. 77, 249-266 (1996).</li><li>Spano, D., Duce, P., Snyder, R. L., Paw, U. K. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Surface+renewal+estimates+of+evapotranspiration.+Tall+canopies.">Surface renewal estimates of evapotranspiration. Tall canopies.</a> Acta Hort. 449, 63-68 (1997).</li><li>Spano, D., Snyder, R. L., Duce, P., Paw, U. K. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Surface+renewal+analysis+for+sensible+heat+flux+density+using+structure+functions.">Surface renewal analysis for sensible heat flux density using structure functions.</a> Agric. For. Meteorol. 86, 259-271 (1997).</li><li>Shapland, T. M., McElrone, A. J., Snyder, R. L., Paw, U. K. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Structure+function+analysis+of+two-scale+scalar+ramps.+Part+I:+Theory+and+modeling.">Structure function analysis of two-scale scalar ramps. Part I: Theory and modeling.</a> Boundary Layer Meterology. , (2012).</li><li>Shapland, T. M., McElrone, A. J., Snyder, R. L., Paw, U. K. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Structure+function+analysis+of+two-scale+scalar+ramps.+Part+II:+Applications.">Structure function analysis of two-scale scalar ramps. Part II: Applications.</a> Boundary Layer Meterology. , (2012).</li></ol>

The surface renewal method presents a tangible opportunity to develop a stand-alone and inexpensive technique to quantify crop water use in real time. Recent advances in signal processing, data logger programming, calibration, and data management have brought this goal into clearer focus. This manuscript and our recently developed turnkey data logger program6 render advanced micrometeorological methods more accessible for agricultural researchers. The output table from the turnkey data program that contains th...

Water scarcity threatens agricultural sustainability in dry growing regions. This situation will likely worsen with changing climatic conditions and increasing competition between agricultural, municipal, industrial, and conservation entities for limited water supplies. To deal with this ongoing dilemma, growers continually search for ways to improve irrigation efficiency, and technology that better estimates crop water use (i.e. evapotranspiration, ET) in real time, and better agrometeorological methods will certainly help these efforts. Current technologies used by growers to estimate ET depend on the use of reference evapotranspiration (ET0) and an empirical crop coefficient (Kc), both of which are highly susceptible to estimation errors. Agrometeorologists also measure ET to evaluate experimental treatments for improving crop water use efficiency1 and to parameterize regional water allocation strategies2, but these methods are highly technical and expensive. Efforts are currently underway to translate existing research-based ET measurement methods into cost-effective and user-friendly technologies for growers.
Surface renewal (SR) is one agrometeorological method used to measure crop ET. SR is based on analyzing the energy budget of air parcels that reside ephemerally within the crop canopy during the turbulent exchange process3 as measured with the station shown in Figure 1 and illustrated theoretically in Figure 2 and Movie 1. The air parcels are manifested as ramp-like shapes in turbulent temperature time series data, and the amplitude and period of the ramps are used to calculate the flux density (Figures 2 and&#160;3). With the SR method, ET for a given crop surface is determined by calculating latent heat flux density (LE) as the residual of the following energy balance equation 
LE=Rn-G-H,
Here, LE is the energy flux density associated with the phase change of water from a crop surface, Rn is the net radiation, G is the soil heat flux density (i.e. energy conducted into or out of the ground), and H is the sensible heat flux density (i.e. energy flux density from the surface to the air or vice-versa that results in a temperature change). Rn is a positive number when the net flux is downward (energy added to the surface), LE and H are positive numbers when the flux is upwards (energy added to the air), and G is a positive number when the flux is downward (energy added to the soil). LE (MJ/m2sec) is then divided by the latent heat of evaporation (L=2.45 MJ/kg) to obtain the mass flux density of water vapor from the surface (i.e. ET).
Measurements of Rn and G are relatively straightforward and inexpensive. Direct measurements of H are more complex and require high frequency data acquisition. The most common method to determine H is with eddy covariance; however, the sonic anemometer required for this method is expensive, complex and, consequently, not widely used by agronomists, horticulturalists, or engineers to determine H and LE. On the other hand, H derived from the SR technique is obtained by a simpler and less expensive method, which uses fine wire thermocouples to measure high frequency air temperatures at the surface-atmosphere interface. Despite the simplicity of the SR, current measurements still require calibration against a sonic anemometer&#39;s eddy covariance estimate of H to obtain sensible heat flux density.
The successful deployment of an SR flux tower for ET measurements can be a daunting challenge, especially for many agricultural researchers without formal training in atmospheric science. SR and eddy covariance measurements require sophisticated technical skills in both programming data loggers to execute tasks with complex instrumentation and writing computer programs to post-process the raw turbulence data into meaningful fluxes. Here, we describe how to setup a field station, install sensors, and utilize our new turnkey data logger program for data collection and post-processing procedures.

<table border="1">
	<tbody>
		<tr>
			<td>Equipment/Material</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td></td>
		</tr>
		<tr>
			<td>Datalogger</td>
			<td>Campbell Scientific, Inc.</td>
			<td>CR1000</td>
			<td></td>
		</tr>
		<tr>
			<td>Datalogger Enclosure</td>
			<td>Campbell Scientific, Inc.</td>
			<td>ENC12/14-SC</td>
			<td></td>
		</tr>
		<tr>
			<td>Tower (6 ft) with grounding &#38; lightning rods</td>
			<td>Campbell Scientific, Inc.</td>
			<td>CM6</td>
			<td></td>
		</tr>
		<tr>
			<td>Cross arm (4 ft)</td>
			<td>Campbell Scientific, Inc.</td>
			<td>CM204</td>
			<td></td>
		</tr>
		<tr>
			<td>Nu-rail crossover fitting</td>
			<td>Campbell Scientific, Inc.</td>
			<td>17953</td>
			<td></td>
		</tr>
		<tr>
			<td>Power supply (12 V) with regulator &#38; battery</td>
			<td>Campbell Scientific, Inc.</td>
			<td>PS100</td>
			<td></td>
		</tr>
		<tr>
			<td>Charger Regulator (12 V)</td>
			<td>Campbell Scientific, Inc.</td>
			<td>CH100</td>
			<td></td>
		</tr>
		<tr>
			<td>Battery Extension Cable</td>
			<td>Campbell Scientific, Inc.</td>
			<td>6186</td>
			<td></td>
		</tr>
		<tr>
			<td>Thermocouple Extension Cable</td>
			<td>Campbell Scientific, Inc.</td>
			<td>FWC-20</td>
			<td></td>
		</tr>
		<tr>
			<td>Thermocouples</td>
			<td>Campbell Scientific, Inc.</td>
			<td>FW3</td>
			<td></td>
		</tr>
		<tr>
			<td>3D sonic anemometer with long neck</td>
			<td>RM Young Company</td>
			<td>81000 RE</td>
			<td></td>
		</tr>
		<tr>
			<td>8 conductor cable for anemometer</td>
			<td>RM Young Company</td>
			<td>18660</td>
			<td></td>
		</tr>
		<tr>
			<td>Soil heat flux plates</td>
			<td>REBS, Inc.</td>
			<td>HFT3.1</td>
			<td></td>
		</tr>
		<tr>
			<td>Soil thermocouples</td>
			<td>Campbell Scientific, Inc.</td>
			<td>TCAV</td>
			<td></td>
		</tr>
		<tr>
			<td>Net radiometer with cable</td>
			<td>Kipp and Zonen, Inc.</td>
			<td>NR Lite 2</td>
			<td></td>
		</tr>
		<tr>
			<td>Heavy duty pole mount for radiometer</td>
			<td>Kipp and Zonen, Inc.</td>
			<td>L-CMB1</td>
			<td></td>
		</tr>
	</tbody>
</table>

surface renewal: an advanced micrometeorological method for measuring and processing field-scale energy flux density data

Advanced micrometeorological methods have become increasingly important in soil, crop, and environmental sciences. For many scientists without formal training in atmospheric science, these techniques are relatively inaccessible. Surface renewal and other flux measurement methods require an understanding of boundary layer meteorology and extensive training in instrumentation and multiple data management programs. To improve accessibility of these techniques, we describe the underlying theory of surface renewal measurements, demonstrate how to set up a field station for surface renewal with eddy covariance calibration, and utilize our open-source turnkey data logger program to perform flux data acquisition and processing. The new turnkey program returns to the user a simple data table with the corrected fluxes and quality control parameters, and eliminates the need for researchers to shuttle between multiple processing programs to obtain the final flux data. An example of data generated from these measurements demonstrates how crop water use is measured with this technique. The output information is useful to growers for making irrigation decisions in a variety of agricultural ecosystems. These stations are currently deployed in numerous field experiments by researchers in our group and the California Department of Water Resources in the following crops: rice, wine and raisin grape vineyards, alfalfa, almond, walnut, peach, lemon, avocado, and corn.

Acquiring the Rn and G components of the energy balance equation are relatively straightforward and inexpensive. Measurements of H are more complex and require high frequency temperature traces measured with a fine wire thermocouple for surface renewal or a sonic anemometer for eddy covariance. An example temperature trace measured with a fine wire thermocouple is represented in Figure 9, and it shows the need for mathematical analysis to extract the signal from this...

Watch this Scientific Journal Video about Surface Renewal: An Advanced Micrometeorological Method for Measuring and Processing Field-Scale Energy Flux Density Data at JoVE.com

Surface Renewal: An Advanced Micrometeorological Method for Measuring and Processing Field-Scale Energy Flux Density Data

Surface renewal is a micrometeorological method that is being used increasingly to determine energy fluxes, but its technical complexity makes it inaccessible to a broad audience. We describe the steps needed to set up and calibrate a surface renewal field station, to acquire and process data, and to correctly interpret results.

Advanced micrometeorological methods have become  increasingly important in soil, crop, and environmental sciences.  For many scientists without formal ...

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Measuring Carbon-based Contaminant Mineralization Using Combined CO2 Flux and Radiocarbon Analyses

A method is described which uses the absence of radiocarbon in industrial chemicals and fuels made from petroleum feedstocks which frequently contaminate the environment. This radiocarbon signal &#8212; or rather the absence of signal &#8212; is evenly distributed throughout a contaminant source pool (unlike an added tracer) and is not impacted by biological, chemical or physical processes (e.g., the 14C radioactive decay rate is immutable). If the fossil-derived contaminant is fully degraded to CO2, a harmless end-product, that CO2 will contain no radiocarbon. CO2 derived from natural organic matter (NOM) degradation will reflect the NOM radiocarbon content (usually &#60;30,000 years old). Given a known radiocarbon content for NOM (a site background), a two end-member mixing model can be used to determine the CO2 derived from a fossil source in a given soil gas or groundwater sample. Coupling the percent CO2 derived from the contaminant with the CO2 respiration rate provides an estimate for the total amount of contaminant degraded per unit time. Finally, determining a zone of influence (ZOI) representing the volume from which site CO2 is collected allows determining the contaminant degradation per unit time and volume. Along with estimates for total contaminant mass, this can ultimately be used to calculate time-to-remediate or otherwise used by site managers for decision-making.

Measuring Carbon-based Contaminant Mineralization Using Combined CO2 Flux and Radiocarbon Analyses

A protocol is described wherein CO2 mineralized from organic contaminant (derived from petroleum feedstocks) biodegradation is trapped, quantified, and analyzed for 14C content. A model is developed to determine CO2 capture zone's spatial extent. Spatial and temporal measurements allow integrating contaminant mineralization rates for predicting remediation extent and time.

A method is described which uses the absence of radiocarbon in industrial chemicals and fuels made from petroleum feedstocks which frequently contaminate ...

Extracting Metrics for Three-dimensional Root Systems: Volume and Surface Analysis from In-soil X-ray Computed Tomography Data

Plant roots play a critical role in plant-soil-microbe interactions that occur in the rhizosphere, as well as processes with important implications to climate change and crop management. Quantitative size information on roots in their native environment is invaluable for studying root growth and environmental processes involving plants. X-ray computed tomography (XCT) has been demonstrated to be an effective tool for in situ root scanning and analysis. We aimed to develop a costless and efficient tool that approximates the surface and volume of the root regardless of its shape from three-dimensional (3D) tomography data. The root structure of a Prairie dropseed (Sporobolus heterolepis) specimen was imaged using XCT. The root was reconstructed, and the primary root structure was extracted from the data using a combination of licensed and open-source software. An isosurface polygonal mesh was then created for ease of analysis. We have developed the standalone application imeshJ, generated in MATLAB1, to calculate root volume and surface area from the mesh. The outputs of imeshJ are surface area (in mm2) and the volume (in mm3). The process, utilizing a unique combination of tools from imaging to quantitative root analysis, is described. A combination of XCT and open-source software proved to be a powerful combination to noninvasively image plant root samples, segment root data, and extract quantitative information from the 3D data. This methodology of processing 3D data should be applicable to other material/sample systems where there is connectivity between components of similar X-ray attenuation and difficulties arise with segmentation.

A methodology for obtaining visual and quantitative root structure information from X-ray computed tomography data acquired in-soil is presented.

Plant roots play a critical role in plant-soil-microbe interactions that occur in the rhizosphere, as well as processes with important implications to ...

Protocol for Measuring the Thermal Properties of a Supercooled Synthetic Sand-water-gas-methane Hydrate Sample

Methane hydrates (MHs) are present in large amounts in the ocean floor and permafrost regions. Methane and hydrogen hydrates are being studied as future energy resources and energy storage media. To develop a method for gas production from natural MH-bearing sediments and hydrate-based technologies, it is imperative to understand the thermal properties of gas hydrates.
The thermal properties' measurements of samples comprising sand, water, methane, and MH are difficult because the melting heat of MH may affect the measurements. To solve this problem, we performed thermal properties' measurements at supercooled conditions during MH formation. The measurement protocol, calculation method of the saturation change, and tips for thermal constants' analysis of the sample using transient plane source techniques are described here.
The effect of the formation heat of MH on measurement is very small because the gas hydrate formation rate is very slow. This measurement method can be applied to the thermal properties of the gas hydrate-water-guest gas system, which contains hydrogen, CO2, and ozone hydrates, because the characteristic low formation rate of gas hydrate is not unique to MH. The key point of this method is the low rate of phase transition of the target material. Hence, this method may be applied to other materials having low phase-transition rates.

We present a protocol for measuring the thermal properties of synthetic hydrate-bearing sediment samples comprising sand, water, methane, and methane hydrate.

Methane hydrates (MHs) are present in large amounts in the ocean floor and permafrost regions. Methane and hydrogen hydrates are being studied as future ...

Method to Produce Durable Pellets at Lower Energy Consumption Using High Moisture Corn Stover and a Corn Starch Binder in a Flat Die Pellet Mill

A major challenge in the production of pellets is the high cost associated with drying biomass from 30 to 10% (w.b.) moisture content. At Idaho National Laboratory, a high-moisture pelleting process was developed to reduce the drying cost. In this process the biomass pellets are produced at higher feedstock moisture contents than conventional methods, and the high moisture pellets produced are further dried in energy efficient dryers. This process helps to reduce the feedstock moisture content by about 5-10% during pelleting, which is mainly due to frictional heat developed in the die. The objective of this research was to explore how binder addition influences the pellet quality and energy consumption of the high-moisture pelleting process in a flat die pellet mill. In the present study, raw corn stover was pelleted at moistures of 33, 36, and 39% (w.b.) by addition of 0, 2, and 4% pure corn starch. The partially dried pellets produced were further dried in a laboratory oven at 70 &#176;C for 3-4 hr to lower the pellet moisture to less than 9% (w.b.). The high moisture and dried pellets were evaluated for their physical properties, such as bulk density and durability. The results indicated that increasing the binder percentage to 4% improved pellet durability and reduced the specific energy consumption by 20-40% compared to pellets with no binder. At higher binder addition (4%), the reduction in feedstock moisture during pelleting was &#60;4%, whereas the reduction was about 7-8% without the binder. With 4% binder and 33% (w.b.) feedstock moisture content, the bulk density and durability values observed of the dried pellets were &#62;510 kg/m3 and &#62;98%, respectively, and the percent fine particles generated was&#160;reduced to &#60;3%.

In this study, a protocol was developed to produce good quality pellets using a flat die pellet mill at reduced specific energy consumption testing high-moisture corn stover and a starch based binder. The results indicated that adding a corn starch binder improved the pellet durability, reduced percent fines and decreased specific energy consumption.

A major challenge in the production of pellets is the high cost associated with drying biomass from 30 to 10% (w.b.) moisture content. At Idaho National ...

Protocol for Microplastics Sampling on the Sea Surface and Sample Analysis

Microplastic pollution in the marine environment is a scientific topic that has received increasing attention over the last decade. The majority of scientific publications address microplastic pollution of the sea surface. The protocol below describes the methodology for sampling, sample preparation, separation and chemical identification of microplastic particles. A manta net fixed on an &#187;A frame&#171; attached to the side of the vessel was used for sampling. Microplastic particles caught in the cod end of the net were separated from samples by visual identification and use of stereomicroscopes. Particles were analyzed for their size using an image analysis program and for their chemical structure using ATR-FTIR and micro FTIR spectroscopy. The described protocol is in line with recommendations for microplastics monitoring published by the Marine Strategy Framework Directive (MSFD) Technical Subgroup on Marine Litter. This written protocol with video guide will support the work of researchers that deal with microplastics monitoring all over the world.

The protocol below describes the methodology for: microplastics sampling on the sea surface, separation of microplastic and chemical identification of particles. This protocol is in line with the recommendations for microplastics monitoring published by the MSFD Technical Subgroup on Marine Litter.

Microplastic pollution in the marine environment is a scientific topic that has received increasing attention over the last decade. The majority of ...

Data Processing Methods for 3D Seismic Imaging of Subsurface Volcanoes: Applications to the Tarim Flood Basalt

The morphology and structure of plumbing systems can provide key information on the eruption rate and style of basalt lava fields. The most powerful way to study subsurface geo-bodies is to use industrial 3D&#160;reflection seismological imaging. However, strategies to image subsurface volcanoes are very different from that of oil and gas reservoirs. In this study, we process seismic data cubes from the Northern Tarim Basin, China, to illustrate how to visualize sills through opacity rendering techniques and how to image the conduits by time-slicing. In the first case, we isolated probes by the seismic horizons marking the contacts between sills and encasing strata, applying opacity rendering techniques to extract sills from the seismic cube. The resulting detailed sill morphology shows that the flow direction is from the dome center to the rim. In the second seismic cube, we use time-slices to image the conduits, which corresponds to marked discontinuities within the encasing rocks. A set of time-slices obtained at different depths show that the Tarim flood basalts erupted from central volcanoes, fed by separate pipe-like conduits.

Three-dimensional (3D) reflection seismology is a powerful method for imaging subsurface volcanoes. By using industrial 3D seismological data from the Tarim Basin, we illustrate how to extract the sills and the conduits of subsurface volcanoes from seismic data cubes.

The morphology and structure of plumbing systems can provide key information on the eruption rate and style of basalt lava fields. The most powerful way ...

Combining Eye-tracking Data with an Analysis of Video Content from Free-viewing a Video of a Walk in an Urban Park Environment

As individuals increasingly live in cities, methods to study their everyday movements and the data that can be collected becomes important and valuable. Eye-tracking informatics are known to connect to a range of feelings, health conditions, mental states and actions. But because vision is the result of constant eye-movements, teasing out what is important from what is noise is complex and data intensive. Furthermore, a significant challenge is controlling for what people look at compared to what is presented to them.
The following presents a methodology for combining and analyzing eye-tracking on a video of a natural and complex scene with a machine learning technique for analyzing the content of the video. In the protocol we focus on analyzing data from filmed videos, how a video can be best used to record participants&#39; eye-tracking data, and importantly how the content of the video can be analyzed and combined with the eye-tracking data. We present a brief summary of the results and a discussion of the potential of the method for further studies in complex environments.

The objective of the protocol is to detail how to collect video data for use in the laboratory; how to record eye-tracking data of participants looking at the data and how to efficiently analyze the content of the videos that they were looking at using a machine learning technique.

As individuals increasingly live in cities, methods to study their everyday movements and the data that can be collected becomes important and valuable. ...

Detection of Tilapia Lake Virus Using Conventional RT-PCR and SYBR Green RT-qPCR

The aim of this method is to facilitate the fast, sensitive and specific detection of Tilapia Lake Virus (TiLV) in tilapia tissues. This protocol can be used as part of surveillance programs, biosecurity measures and in TiLV basic research laboratories. The gold standard of virus diagnostics typically involves virus isolation followed by complementary techniques such as reverse-transcription polymerase chain reaction (RT-PCR) for further verification. This can be cumbersome, time-consuming and typically requires tissue samples heavily infected with virus. The use of RT-quantitative (q)PCR in the detection of viruses is advantageous because of its quantitative nature, high sensitivity, specificity, scalability and its rapid time to result. Here, the entire method of PCR based approaches for TiLV detection is described, from tilapia organ sectioning, total ribonucleic acid (RNA) extraction using a guanidium thiocyanate-phenol-chloroform solution, RNA quantification, followed by a two-step PCR protocol entailing, complementary deoxyribonucleic acid (cDNA) synthesis and detection of TiLV by either conventional PCR or quantitative identification via qPCR using SYBR green I dye. Conventional PCR requires post-PCR steps and will simply inform about the presence of the virus. The latter approach will allow for absolute quantification of TiLV down to as little as 2 copies and thus is exceptionally useful for TiLV diagnosis in sub-clinical cases. A detailed description of the two PCR approaches, representative results from two laboratories and a thorough discussion of the critical parameters of both have been included to ensure that researchers and diagnosticians find their most suitable and applicable method of TiLV detection.

This protocol diagnoses Tilapia Lake Virus (TiLV) in tilapia tissues using RT-PCR methodologies. The entire method is described from tissue dissection to total RNA extraction, followed by cDNA synthesis and detection of TiLV by either conventional PCR or quantitative PCR using dsDNA binding a fluorescent binding dye.

The aim of this method is to facilitate the fast, sensitive and specific detection of Tilapia Lake Virus (TiLV) in tilapia tissues. This protocol can be ...

An Induction System for Clustered Stomata by Sugar Solution Immersion Treatment in Arabidopsis thaliana Seedlings

Stomatal movement mediates plant gas exchange, which is essential for photosynthesis and transpiration. Stomatal opening and closing are accomplished by a significant increase and decrease in guard cell volume, respectively. Because shuttle transport of ions and water occurs between guard cells and larger neighboring epidermal cells during stomatal movement, the spaced distribution of plant stomata is considered an optimal distribution for stomatal movement. Experimental systems for perturbing the spaced pattern of stomata are useful to examine the spacing pattern's significance. Several key genes associated with the spaced stomatal distribution have been identified, and clustered stomata can be experimentally induced by altering these genes. Alternatively, clustered stomata can be also induced by exogenous treatments without genetic modification. In this article, we describe a simple induction system for clustered stomata in Arabidopsis thaliana seedlings by immersion treatment with a sucrose-containing medium solution. Our method is easy and directly applicable to transgenic or mutant lines. Larger chloroplasts are presented as a cell biological hallmark of sucrose-induced clustered guard cells. In addition, a representative confocal microscopic image of cortical microtubules is shown as an example of intracellular observation of clustered guard cells. The radial orientation of cortical microtubules is maintained in clustered guard cells as in spaced guard cells in control conditions.

An Induction System for Clustered Stomata by Sugar Solution Immersion Treatment in Arabidopsis thaliana Seedlings

The goal of this protocol is to demonstrate how to induce clustered stomata in cotyledons of Arabidopsis thaliana seedlings by immersion treatment with a sugar-containing medium solution and how to observe intracellular structures such as chloroplasts and microtubules in the clustered guard cells using confocal laser microscopy.

Stomatal movement mediates plant gas exchange, which is essential for photosynthesis and transpiration. Stomatal opening and closing are accomplished by a ...

Processing Embryo, Eggshell, and Fungal Culture for Scanning Electron Microscopy

Although scanning electron microscopy (SEM) is being widely used for the ultra-structural analysis of various biological and non-biological samples, methods involved in processing different biological samples involve unique practices. All conventional practices described in the literature for processing samples still find useful applications, but subtle changes in the sample preparation can alter image quality, as well as, introduce artifacts. Hence, using a unique sample preparation technique specific to the type of tissue analyzed is required to obtain a good quality image with ultrastructural resolution. The focus of this study is to provide the optimal sample preparation protocols for imaging embryos, rigid eggshells, and fungal cultures using SEM. The following optimizations were recommended to yield good results for the three different delicate biological samples studied. Use of milder fixatives like 4% paraformaldehyde or 3% glutaraldehyde followed by dehydration with ethanol series is mandatory. Fungal mycelium on agar blocks obtained by slide cultures yields a better ultrastructural integrity compared to cultures taken directly from agar plates. Chemical drying of embryos with HMDS provides drying without introducing surface tension artifacts compared to critical point drying. HMDS prevents cracking caused by shrinkage as samples are less brittle during drying. However, for fungal culture, critical point drying provides acceptable image quality compared to chemical drying. Eggshells can be imaged with no special preparation steps except for thorough washing and air drying prior to mounting. Preparation methodologies were standardized based on acceptable image quality obtained with each trial.&#160;

Here, we present detailed processing protocols for imaging delicate tissue samples using scanning electron microscopy (SEM). Three different processing methods, namely, hexamethyl disilazana (HMDS) chemical drying, simple air drying, and critical point drying are described for preparing rigid eggshells, embryos at early developmental stages, and fungal cultures respectively.

Although scanning electron microscopy (SEM) is being widely used for the ultra-structural analysis of various biological and non-biological samples, ...