Acclimation Period and Effect on Temperature and Light Response in Populus grandidentata and Betula alleghaniensis Grown in Three Temperature Regimes. CHRISTINA CAMPION (Rhodes College Memphis, TN 38112) CARLA GUNDERSON (Oak Ridge National Laboratory, Oak Ridge, TN, 37831)

Rising concentrations of atmospheric carbon dioxide are projected to increase global air temperatures by 1 to 4.5^oC in the next century. If modern ecosystems are to remain unchanged through such rapid climatic warming, it is necessary that plants be able to physiologically acclimate to said increases in growth temperature. To investigate whether species differences in temperature response or adjustment are important to modeling photosynthetic and other responses to warming climates, four deciduous tree species (Quercus alba, Betula alleghaniensis, Liquidambar styraciflua, and (Populus grandidentata), have been grown for four years in replicated open-top chambers with three separate temperature treatments: ambient air temperature (A), 2^oC above ambient (E2), and 4^oC above ambient (E4). These species were selected in order to study the responses of deciduous trees to atmospheric warming as most acclimation research has focused on coniferous or other evergreen trees. Photosynthetic temperature response curves were collected from P. grandidentata and B. alleghaniensis. Optimal temperatures (T_opt) were then determined and compared statistically with air and soil temperature history for each chamber. Experimental T_opt were then compared with meteorological data collected by chamber. In P. grandidentata, evidence from 2004 and 2005 indicated an acclimation to daytime average temperature of the preceding 2.5 days. Data from B. alleghaniensis in 2003 and 2004 indicated a faster response of 1.5 days, indicating higher fitness as well as complete acclimation. Light response curves in P. grandidentata and B. alleghaniensis, taken in the A and E4 treatments, showed no significant difference in light compensation point, quantum yield, or rate at saturating light accompanying the temperature acclimation. Knowledge of the acclimation of photosynthesis to temperature allows predictions to be made concerning the future composition of forests and the chances that, specifically, these two species will survive given a moderate rise in temperature. Key words: Photosynthesis, temperature, irradiance, Populus grandidentata, Betula alleghaniensis, acclimation, photosynthetic plasticity

Cataloging and Indexing the Data Holdings of the Carbon Dioxide Information Analysis Center using the ORNL Metadata Editor and Mercury. KRISTEN REEVES (Southeastern Louisiana University Hammond, LA 70402) THOMAS A. BODEN (Oak Ridge National Laboratory, Oak Ridge, TN, 37831)

The Carbon Dioxide Information Analysis Center (CDIAC) serves as the primary data repository for The United States Department of Energy's (DOE) Global Change Research Program (GCRP). This program facilitates data acquisition and data management activities necessary for basic research on global change, promotes the enhancement of modeling designed to improve representation of Earth system interactions, and develops advanced analytic methods to facilitate fundamental research. CDIAC maintains hundreds of data sets related to climate change. These data have been submitted to CDIAC by researchers around the world. One of CDIAC's primary goals is to make well-documented data sets visible and available to users worldwide via the Web. To reach this goal computing resources have been invested to improve Web-based access, visualization, and retrieval of CDIAC data holdings. Specifically, CDIAC has developed and implemented the CDIAC OME (ORNL Metadata Editor) and the CDIAC version of Mercury. OME is a Web-based tool used to fully document databases by collecting the appropriate metadata (i.e. definitional data that provides information about, or documentation of, other data managed within an application or environment). For this project, the OME was used to generate new metadata for many of CDIAC's data holdings. This metadata consisted of author information, thematic area, beginning and end dates of measurements, variables, data location, etc. Existing OME entries were revised, embellished, and consolidated, where appropriate, to lend better organization and ease navigation. Once the metadata had been collected from the various sources an XML (eXtensible Markup Language) file was created on the development server. These XML files were then harvested by Mercury each night. Mercury is a Web-based search engine used to search for metadata and retrieve associated data. CDIAC staff helped review the metadata collected, and made the appropriate edits needed to assure the most accurate description of data. The OME was then used to modify the existing XML files. Finally, those files were moved to the CDIAC development server, where they were used nightly by the CDIAC version of Mercury to build a locator file; this is also where the catalog is built. Mercury indexes the catalog and makes it searchable on the CDIAC web-site. Mercury then allows users to query the catalog by keyword, subject, title, and author to find the specific data sets they will need for research.

Characterization of Sulfate-Reducing Bacterial Populations in a Uranium-Contaminated Aquifer in Oak Ridge, Tennessee. AMY VOSS (Auburn University Auburn, AL 36849) TERRY GENTRY (Oak Ridge National Laboratory, Oak Ridge, TN, 37831)

The Field Research Center (FRC), located near Oak Ridge National Laboratory in Oak Ridge, Tennessee, contained four unlined ponds that were used for waste disposal during the years 1951 to 1983. As a result, the soil and groundwater of the FRC are contaminated with uranium-containing waste that has potential to infiltrate local water supply. Bioremediation procedures currently under development involve the use of microorganisms to reduce uranium from U(VI) to U(IV), thus lowering its solubility and reducing the potential for offsite migration. Many sulfate-reducers have been verified to also reduce uranium. The dissimilatory sulfite reduction (dsrA/B) genes were used to detect the presence of sulfate-reducing bacteria potentially capable of U(VI) reduction. The Polymerase Chain Reaction (PCR) was used to amplify the prospective dsrA/B genes in samples collected from the groundwater well FW-106. Gel electrophoresis was performed to confirm that PCR amplification produced the correct-sized fragment. A clone library was constructed from 16S rDNA and dsrA/B PCR products, creating approximately 300 clones containing the genes. These clones were screened to determine the size of the insert. Gel electrophoresis indicated that the inserted fragments were the correct size, and these inserts were then sequenced. A phylogenetic tree constructed for the dsrA/B clones showed that FW-106 was dominated by a single population. Comparison of the putative dsrA/B sequences with those from known organisms indicated that the clones were likely not from dsrA/B genes. However, genes that were found are highly unique and will be used along with a metagenomic sequencing project to determine if indigenous microbial populations have potential to reduce uranium. Knowledge of specific sulfate-reducing populations at the site will allow conditions to be managed for optimal U(VI) reduction.

Exploring Optimal Growing Conditions For Hydrogen Production by Rhodopseudomonas palustris. SARAH WOODS (Lipscomb University Nashville, TN 37204) QIANG HE (Oak Ridge National Laboratory, Oak Ridge, TN, 37831)

Rhodopseudomonas palustris is widely distributed in natural environments and is one of the most metabolically versatile bacteria ever described. It is capable of phototrophic hydrogen production and CO2 assimilation, which are of great significance for H2 production as an alternative fuel and carbon sequestration to reduce greenhouse gas emission. However, many questions remain unanswered before technologically feasible processes can be developed for hydrogen production. In this study various growth conditions were explored with the aim to determine the effect of carbon source, nitrogen source and oxygen on the phototrophic growth and hydrogen production potential of R. palustris During this study, R. palustris was grown in 30-ml anaerobic culture tubes with 10-ml of degassed medium and closed with rubber stoppers and aluminum seals. Cultures were incubated under light at 30 C under anaerobic conditions (100% N2 headspace) to simulate photosynthesis. Various amounts of different organic substrates, nitrogen sources or oxygen were added to the cultures to test their impact on growth. Growth was monitored spectrophotometrically at 660nm to avoid optical interference from the cell pigments. Growth curves of R. palustris revealed that acetate allowed optimal growth with the most rapid growth rate (doubling time˜3.5h) and the highest yield (OD660>2). More moderate growth on fumarate and succinate were also observed. Therefore, acetate being a more reduced substrate is a better carbon source for cell mass than fumarate and succinate. In contrast, growth on pyruvate was minimal with final OD660 of 0.091. Because pyruvate contains more reducing equivalents than acetate yet provided much less growth, this indicates the significant energetic obstacles that prevent the efficient assimilation of pyruvate. Nitrogen source is another growth impacting factor for hydrogen production of R. palustris. Compared with nitrogen-fixing growth, the presence of fixed nitrogen (NH4+-N) increased the growth yield by 35%, explaining the ammonia inhibition of hydrogen production is due to the inhibition of nitrogen fixation process, which is responsible for hydrogen evolution. Results show that the phototrophic growth of R. palustris and its potential for hydrogen production is affected by the type of organic carbon source, the presence of oxygen, and the form of nitrogen. For hydrogen production processes to make way in the future, these factors should be taken into consideration.

Generation of Populus Aux/IAA RNAi Constructs, Soil Acclimation of Transgenics and Standardization of RT-PCR Technique for Confirmation of RNAi Effect. MEGAN O'SHAUGHNESSEY (Purdue University West Lafayette, IN 47906) UDAYA C. KALLURI (Oak Ridge National Laboratory, Oak Ridge, TN, 37831)

Over the past centuries CO2 levels have been increasing causing increased global warming and climate changes. Plants play an important role in terrestrial carbon sequestration owing to their natural ability to fix atmospheric CO2 through photosynthesis. This study concentrates on identifying and characterizing genes that control the fate of belowground carbon in plants. Populus is an excellent model for this study because it is a fast growing tree, has its genome entirely sequenced, and has well established molecular biology and transformation tools. Auxin is a hormone that plays a major role in plant bioprocesses such as cell division, cell elongation, vascular development and root formation. Cell walls are a long-term sink for carbon from photosynthates, thus auxin helps to indirectly control carbon storage levels. The AUX/IAAs gene family is involved in auxin signaling and response. Therefore, in this project, Populus AUX/IAA RNAi constructs were created out of transgenic plants. The functional role of the specifically down-regulated AUX/IAA gene can be elucidated through observation of phenotypic changes in the transgenic plants. Hence, RNAi constructs for specific genes will ultimately help in understanding the possible functional roles these genes play in carbon sequestration. To this end, gene-specific fragments were amplified from Populus genomic DNA though PCR techniques and then directionally cloned using a D-TOPO pENTR reaction, resulting in an entry clone that is subsequently used to create an expression clone using pCAPT vector in a gateway technology based recombination reaction. Upon sequence confirmation, the expression clone was introduced into Agrobacterium C58 cells for use in Populus transgenesis. Transgenic plants were generated from this work. Acclimation of tissue culture plants to the greenhouse conditions was carried out gently and carefully. Transgenic tissue samples were studied microscopically to screen for phenotypic differences. RT-PCR was used to measure the extent of down-regulation achieved in the transgenic plants. These studies will allow us to directly observe whether the targeted gene has been down regulated as expected and will also provide further insight into the role of AUX/IAA genes in carbon allocation.

Identification of an Arabidopsis thaliana Mutant Having a T-DNA Insertion in an NIA Gene. PETER ANTHOPOLOS (University of Tennessee Knoxville, TN 37996) LEE E. GUNTER (Oak Ridge National Laboratory, Oak Ridge, TN, 37831)

The overarching hypothesis of this study is that a single-enzyme change affecting an important biological process (e.g., nitrate assimilation) will translate across multiple levels of biological organization to produce predictable responses at the ecosystem level. Long term plans consist of exposing wild-type (WT) and transformed Arabidopsis thaliana (nitrate reductase deficient or nia^-) to ambient and elevated levels of atmospheric CO₂. Additionally, the competitive interaction between the WT and nia^-) will be recorded at the various CO₂ levels, as will the interaction between the carbon and nitrogen cycles. The goal of the project described herein was to identify additional nia^- mutants for the study. Two genes in Arabidopsis code for nitrate reductase, and the objective was to identify two classes of Arabidopsis mutants: the first class consisted of mutants having an Agrobacterium T-DNA insertion in NIA1, the first gene for nitrate reductase, and the second class consisted of mutants having an insertion in NIA2, the second gene for nitrate reductase. Each of the seed lines acquired for this project had a putative T-DNA insertion in either NIA1 or NIA2. The seeds were germinated and DNA was extracted from the seedlings. Polymerase chain reactions (PCR) were performed on the DNA using two types of primer pairs: (1) primer pairs designed to detect only the wild-type allele of NIA1 (but not NIA2) or vice versa and (2) primer pairs designed to detect only the presence of a T-DNA insertion in NIA1 (but not NIA2) or vice versa. Initial analysis was completed on three seed lines: Salk 004181 (putative insertion in NIA1), Salk 075996 (putative insertion in NIA2) and Salk 138297 (putative insertion in NIA2). Through PCR, the presence of a T-DNA insertion was confirmed in the Salk 138297 seed line and an individual homozygous for the insertion was isolated from that population. Southern blotting will be performed using DNA from Salk 138297 seedlings to ensure that no other T-DNA insertions are present in the genome, as there is a significant chance that two or more T-DNA insertions resulted from the transformation. PCR indicated that neither of the remaining two seed lines had an insertion at the specified location. The project was successful in that one of the two desired classes of mutants, i.e., the NIA2 mutant, was isolated. Additional seed lines will be tested in an ongoing effort to identify a mutant having an insertion in NIA1.

Identification of Metal Resistance Genes in Microbial Communities from Atlantic Fleet Weapons Training Area (AFWTA) Facility in Vieques, Puerto Rico using Microarray Techniques. ERNIE PEREZ (University of Puerto Rico at Mayaguez Mayaguez, PR 00681) CHRISTOPHER W. SCHADT, TERRY GENTRY (Oak Ridge National Laboratory, Oak Ridge, TN, 37831)

The Atlantic Fleet Weapons Training Area (AFWTA) facility, which is located in the eastern part of Vieques, PR, includes land areas, waters and cays impacted by 63 years of military training operations, largely by the U.S. Navy. Samples from the island of Vieques, Guánica, PR and Adjuntas, PR (same geoclimatic zone) were collected in May, 2005. The Functional Gene Array (FGA) technique was used to study the microbial community. Metal resistance genes for Cd, Pb, Hg, Al, and others were included in the analysis. Using different methods, DNA was extracted and purified from the microbial communities in the samples. The community DNA was amplified using the Whole Community Genome Amplification (WCGA) with different quantities (50ng/uL, 100ng/uL and 200ng/uL) of template. The amplified template was labeled with Cy5 fluorochrome dye. Hybridization was performed between the samples and a FGA probes encompassing functional genes for a variety of microbial processes including approximately 4,546 probes for metal resistance gene targets. The objective is to identify metal resistance genes from the AFWTA facility in Vieques, Puerto Rico and compare the results with the ones obtained from the forests in Guánica and Adjuntas. It is believed that a low biomass due to contamination at the AFWTA facility caused difficulties in its DNA extraction. The MoBio-Soil Extraction-Protocol seemed to be the method with the best extraction results. After many attempts, it was concluded that the WCGA made better amplifications using a 50ng/uL concentration of template. The labeling was performed with the expected outcomes. The results obtained from this analysis will provide the tools to identify the profile of genes involved in metal resistance and also develop some strategies for the island's restoration.

Physiological Adjustments of Leaf Respiration in Betula alleghaniensisand Quercus rubra Grown at Varying Temperatures. ASHLEY VOLLMAR (Pellissippi State Technical Community College Knoxville, TN 37933) CARLA A. GUNDERSON (Oak Ridge National Laboratory, Oak Ridge, TN, 37831)

Global air temperatures are predicted to rise 1° to 4.5° Celsius by the year 2100. This climatic change can have a great effect on the succession and migration of temperate deciduous forest species. Most physiologically based models of forest response to climatic change focus on the ecosystems as a whole instead of on individual tree species, assuming that the effects of warming on respiration are generally the same for each species, and that processes can not adjust to a changing climate. Experimental data suggest that physiological adjustments are possible, but there is a lack of data in deciduous species. In order to correctly model the effects of climate change on temperate species, species-specific respiration acclimation (adjustment) to rising temperatures is being determined in this experiment. Two temperate deciduous tree species Betula alleghaniensis (BA) and Quercus rubra (QR) were grown over a span of four years in open-top chambers and subjected to two different temperature treatments; ambient and ambient plus 4° Celsius (E4). Between 0530 hours and 1100 hours, respiration was measured over a range of leaf temperatures on several comparable, fully expanded leaves in each treatment. Circular punches were taken from the leaves and dried at 60° C to determine leaf mass per area (LMA). Respiration rates at a common temperature in both species decreased by at least 4° C with increasing growth temperature, indicating a large degree of physiological acclimation. Foliar mass per area decreased with increasing growth temperature for both species. It can be concluded that there is a relationship between leaf respiration and foliar mass as it relates to respiratory acclimation, and that these two species had similar patterns of adjustment to warming.

Properties of Colloidal Silica Nanoparticles in Aqueous Solution With Organic and Inorganic Cations. JENNIFER XU (University of Southern California Los Angeles, CA 90007) WEI WANG (Oak Ridge National Laboratory, Oak Ridge, TN, 37831)

The adsorption of various ionic species onto silica nanoparticles causes variations in their apparent surface charge, stability, and other properties. These changes of characteristics are of especial interest because of their potential to be used in the analysis of river sediments (composed mostly of sand or silica) when different toxins are introduced and become adsorbed onto these nanoparticles. While many past studies have focused on alkali metal salts, this study has also examined the effect of varying pH, as well as different concentrations of large organic molecules and phospholipids on the zeta potential of the silica nanoparticles. Using a dynamic light scattering technique (DLS), both the effective diameter and the zeta potential of spherical silica nanoparticles (~40 nm in diameter, highly purified and monodisperse, neutral pH) in aqueous solution with concentrations of different cations ranging from 0 to 1x10-2 mol/L were investigated. These cations include four alkali chloride salts, four quaternary ammonium cations, and a phospholipids (DPPC). The effect of pH on the plain silica nanoparticles was also studied, both with and without an electrolyte ( 0.01 M KCl) in aqueous solution. From the pH variation experiment, it was observed that the aqueous KCl greatly stabilized the zeta potential fluctuation concomitant with the variation of pH. The trials with both the organic and inorganic cations indicate that the effective diameter of silica colloidal nanoparticles increases with increasing size of the cation, as does the zeta potential. With larger cations the zeta potential increased more dramatically than when the colloid is in solution with smaller cations. Therefore, larger cationic molecules cause greater flocculation and aid in the precipitation of silica nanoparticles. Additionally, a total organic carbon (TOC) analysis was done to determine the amount of adsorption of DPPC on colloidal silica. The result shows that there is a linear relationship between amount of DPPC adsorbed and the concentration of the DPPC in solution with the silica particles. If these behaviors can be predicted in river water where the alkalinity, pH, or concentrations of various disposed organic wastes can be predicted, sedimentation in rivers can be selectively formed and more accurately determined. This study is part of a larger project dealing with various properties of many other types of nanoparticles.

The Use of Agricultural Modeling to Predict the Potential Supply of Biomass Feedstocks over the Next Ten Years and to Describe Economic Impacts. BENJAMIN CLEMENS (University of Virginia Charlottesville, VA 22904) BOB PERLACK (Oak Ridge National Laboratory, Oak Ridge, TN, 37831)

As the price of oil continues to increase, the United States government is increasingly looking to alternative fuel sources to replace crude oil demand. Though hydrogen power shows great promise in the future, many analysts believe that more immediate solutions are required. One possibility to bridge this energy gap is the use of biofuels, which are refined from products ranging from corn grain to specially harvested grasses made specifically for energy use. A considerable amount of biomass feedstock is currently available, and more could become available depending on government pricing programs and advances in technology. Modeling of the results shows that the availability of cellulosic biomass feedstocks such as switchgrass, corn stover, and wheat straw is highly dependent on yield and price. Using current technology and tillage practices, 200 million dry tons of cellulosic residue could be harvested at a farmgate price of $50/dry ton by 2014. These two sources alone could replace as much as 12% of gasoline demand, providing a clean, domestic alternative to foreign oil. Regional analysis shows that regions that can produce corn stover closely mirror the largest corn-producing regions, while switchgrass production will be centered on the Mississippi River Valley, the Great Plains, and the Southeast coast. Crop residues can be profitably harvested at prices as low as $30/dry ton, while switchgrass must compete with conventional crops for acreage, and therefore does not displace a significant amount of cropland below prices of $40/dry ton. Increased acreage of switchgrass would lead to a uniform rise in the prices of major crops, resulting in a substantial increase in net returns at the same time that government payments decline. This reduction in payments allows the government to borrow against future savings to subsidize switchgrass startup expenses, a necessary incentive given the high cost of beginning switchgrass crops. Current ethanol plant technology cannot keep up with the forecasted increase in production, so new ethanol plants must be built with the ability to process cellulosic biomass.