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Masters degree: 2003-2006
My masters program took me north from the scorching heat of Arizona to the frigid cold of Wyoming. I went north to work with David Williams on a masters degree in rangeland ecology and watershed management. The University of Wyoming impressed me immediately because it worked much more efficiently than the bureaucratic mess that plagued the University of Arizona. The school was much smaller than U of A, but it had a lot to offer. I decided to stay in Dave’s lab when we moved to UWyo to finish an unfinished part of my previous research in Arizona. I wanted to know why mesquite (Prosopis velutina) fractionated hydrogen isotopes during water uptake (pdf of published paper).
Cryogenic vacuum distillation line for extracting water from stem and soil samples. This line is partially built. We still had not installed the automatic heaters to heat the sample vials, the pressure gauge system, and vacuum pump.
Here are the plants from my greenhouse study to measure the magnitude of isotopic separation of 16 xerophytic and halophytic species. These species are mesquite (Prosopis spp.)
Magnitude of isotopic separation (d2H = 2Hsoil water + 2Hxylem water) for 16 species of semi-arid and arid shrubs and trees and the herbaceous mesophyte Lycopersicon esculentum
After the greenhouse study, I looked at the variation in d18O and d2H of xylem water in various plant parts. The two outcomes of these two experiments was that found that fractionation was only measured in hydrogen and the location of fractionation was the roots. The roots had the lowest d2H values of any plant parts.
The relationship between the magnitude of isotopic separation (d2H = 2Hsoil water + 2Hxylem water) and salinity tolerance, defined as the minimum soil electrical conductivity negatively affecting plant growth. Values of salinity tolerance were obtained from published studies. Information on salinity tolerance for five species (Acacia constricta, Acacia greggii, Artemisia ludoviciana, Flourensia cernua, and Olneya tesota) were not available
Mean isotopic composition of water extracted from woody, green stems (young), woody stems with well-developed bark (old), leaves and soil from five potted individuals of P. velutina. Two-tailed paired t-tests were used to determine the statistical significance of mean differences between the isotopic composition of water from stems and soil samples. 2H and 18O values of plant organs that were significantly different from that of the soil are marked with an asterisk (*P < 0.01, **P < 0.001, ***P < 0.0001). Mean 18O values of water extracted from old stems were slightly more positive than that of soil water (P = 0.07)
Mean isotopic composition of leaves, whole stem segments, sapwood, bark, and roots of four potted Prosopis velutina plants. The isotopic composition of the leaf was calculated a modified form of the Craig and Gordon model (Yakir and Sternberg 2000). Two-tailed paired t-tests were used to determine the statistical significance of mean differences between the isotopic composition of water from plant tissue and soil samples. 2H and 18O values of plant organs that were significantly different from that of the soil are marked with an asterisk (*P < 0.05, **P < 0.01, ***P < 0.001). The inset represents sapwood (inner region) and bark (outer region) of the stem and their respective isotopic compositions
After conducting the pot experiments, I wanted to identify the mechanism responsible for fractionation. The hypothesis was that the symplastic movement of water through root especially through the endodermis fractionates hydrogen isotopes. Root pressurization was designed to measure the isotopic composition of water pushed from the root system. The pressure chamber was filled with water and pressure was applied to the water head space. Water was collected from the top of the root system. Under our hypothesis the d2H values of the water leaving the root system would be less than the d2H values of the source water in the chamber. Also as the pressure increased, resulting an increase in flow rate through the root system, the d2H values of the water leaving the root system would decrease.
The relationship between the 2H of water expressed from whole, severed root systems and flow rate per unit root mass of three xerophytic woody species. Data are composites from three individuals of each species. Statistically significant linear regressions (solid lines) were observed for data from live root systems of Artemisia tridentata (y = -2.8x - 128.5, R2 = 0.34, P < 0.001) and Atriplex canescens (y = -61.3x - 122.2, R2 = 0.2, P < 0.01). Disruption of the plasma membranes from excessive heat by boiling did not significantly alter the relationship between flow rate and the 2H of water expressed from the root systems in these species (dashed lines)
My first research project in plant physiological ecology was with velvet mesquite (Prosopis velutina), which I began in 2000 at the University of Arizona. I was able to complete my study as part of a Undergraduate Scholar Fellowship from the Arnold and Mabel Beckman Foundation. My project was part of a larger Upper San Pedro Partnership Water Needs study to quantify the water use of the San Pedro Riparian National Conservation Area (SPRNCA).
cottonwood-willow gallery forest along the San Pedro River near Charleston, AZ, USA
My part of the study was to partition mesquite water sources and determine the seasonal trends of water use. My project was part of a larger Upper San Pedro Partnership Water Needs study to quantify the water use of the San Pedro Riparian National Conservation Area (SPRNCA). My part of the study was to partition mesquite water sources and determine the seasonal trends of water use. When my study began in 2000, the assumption used for riparian mesquite woodlands was that they used almost exclusively groundwater. However, this assumption would overestimate groundwater use and the important source that the deep vadose zone and monsoonal precipitation play in mesquite-water relations. I used stable isotopes to partition water resources. I partitioned daily mesquite transpiration flux by mesquites' three water sources: shallow soil water (0-1 m) from precipitation, deep vadose zone (1 m - capillary fringe), and groundwater. Mesquite transpiration fluxes were provided by Russ Scott's eddy covariance data. I was able to partition total mesquite water use by its water sources because soil moisture in the top 1 m of soil was highly correlated to water use from the top 1 m of soil. Using the line of regression from the correlation, I partitioned daily water use by the corresponding water sources.
Top two pictures: mesquite bosques near Charleston, AZ, USA. Photos by Bill Cable
Left: field site near Charleston where Enrico Yepez is conducting vapor collection for partitioning evapotransporation. Photo by Marcela Yepez
I needed to know the volumetric water content, soil texture, and most importantly the δ2H and δ18O of the deep vadose zone soil (deep unsaturated zone). The groundwater table was 9.5m down, so I had take soil samples along a core 9.5 m long. That was a lot of digging!
I made the auger using old metal water pipe and used unions to join the 7ft pipes. I used a nut welded onto a pipe to screw the bucket into the bottom of the auger. The auger broke once, but fortunately it was fixable. Bill and I collected 2 cores of 7.2 and 9.5 m deep. Thank goodness we did not hit any rocks on the way down!
Believe it or not but the bucket of this soil auger is 9.5m down.
My early research experience 1995-1997
First official research experience (summer of 1995): where I got my feet wet
I began my first research experience the summer after my junior year in high school when I was accepted to the High School Biology Research Apprenticeship Program (HSBRAP) at the University of Arizona. I worked in Dr. Kathy Taylor’s plant physiology lab, studying the disease citrus blight. My project focused on plant regulation of the suspected wound-inducible protein chitinase. Chitinase is the enzyme responsible for breaking down chitin (poly-N-acetylglucosamine) to its sugar monomers. Chitin is found in the cell walls of fungi and the exoskeleton of insects, so chitinase is a pathogenic response to wounding by these organisms. I investigated the production of chitinase produced by citrus trees as part of a wound-inducible response. I sampled leaves from blight-affected and healthy citrus trees, and then isolated the enzymes using gel filtration chromatography. The quantity of chitinase activity was analyzed by measuring the concentration of chitin reduced to its sugar monomer, N-acetylglucosamine by chitinase with a spectrophotometer. I found that chitinase activity in citrus increased if the tree had citrus blight, but the chitinase activity varied among leaves.
Molecular work continues: 1996-1997
I again worked in Dr. Taylor’s lab in HSBRAP. Another graduate student in the lab had identified and cloned a gene responsible for a suspected pathogenic response in blight-affected citrus trees. My project was to develop a procedure to introduce this gene into three citrus rootstocks, Citrus aurantifolium, C. macrophylla, and ‘Troyer’ citrange. By taking advantage of natural characteristics of Agrobacterium tumefaciens such as the process of introducing T-DNA from the Ti-plasma into the plant cell, selected genes can be transferred from the bacteria to the plant. Transformation requires infection of the epicotyls with A. tumefaciens and regeneration of transgenic plant tissue. I cultured transgenic stock of A. tumefaciens and carried out the infection and regeneration process. To verify that the Agrobacterium used was transformed with the correct gene, I used polymerase chain reaction (PCR) to amplify the bacterial T-DNA and then verified the presence of the gene using electrophoresis.
Regeneration of the plant tissue required using the correct hormone concentration and ratio between cytokinin and auxin to regenerate roots and shoots without the formation of callus. Callus could increase the probability of genetic mutation. My experiment contained 61 sets of hormonal ratios of cytokinins to auxins for each of three rootstocks. After the correct hormone concentration was found and regeneration accomplished, the regenerated plants were tested for the presence of the selected gene using the sample method of gene amplification and verification that was used with Agrobacterium. I successfully regenerated all the rootstocks and transformed ‘Troyer’ citrange. I presented the results of these two projects in the Undergraduate Biology Research Conference in 1995 and 1996.
First field research project: flat-tailed horned lizard in the ultra-arid desert near Yuma in the summer of 1997
I assisted Kevin Young, a graduate student of Dr. Edmund Brodie at Utah State University, studying a rare lizard species, Phrynosoma mcallii, in Yuma, Arizona. The project included studying the effects of predation, drought, and depredation along roads on the lizard’s small population. I assisted in behavioral trials to identify behavioral patterns in male-female interactions and intra- and interspecific competition between P. mcallii and P. platyrhino.
I assisted in behavioral trials to identify behavioral patterns in male-female interactions and intra- and interspecific competition between P. mcallii and P. platyrhino. I tracked lizards using radio telemetry to determine their active range and daily movements. I conducted surveys of lizard populations, habitat use, and the influences of predators on population size and distribution. My work was included in the report: Scientific study of the flat-tailed horned lizard, P. mcallii prepared for the US Department of Navy Contracts and in the strategy plan: Flat-tailed horned lizard rangewide management strategy, 2003 revision prepared by Flat-tailed Horned Lizard Interagency Coordinating Committee.
A fascinating find from this study was the purpose of horns on the horned lizards. Kevin found that the larger the horns on the lizard, the less likely that they will be predated by loggerhead shrikes (Lanius ludovicianus). This shows adaptation of horns as anti-predator defense. Reference: Young et al. (2004) Science 304: 65
Loggerhead shrike - the bird responsible for many impelled horned lizards on desert shrubs. It is deserving of its nickname, the butcherbird.
(A) a picture of a skull left impelled on a creosote bush (Larrea tridentata)
(B) Flat-tailed horned lizard in defensive posture
(C) relative parietal horn length
(D) relative squamosal horn length
Another interesting aspect of horned lizards is their keen vision. I have never seen lizards that were more aware of their surroundings. We conducted behavioral trials with flat-tailed and desert horned lizards. The first attempt was to observe the lizards in their natural habitat. However, they would not move as long as someone was nearby. Next we tried to place the lizards in baby pools full of sand surrounded by camouflage mesh to hide us from view as we observed them. They could see through the mesh, so that did not work. Next the mesh was replaced with cardboard with small holes cut in them to function as observation holes. They could see use through the holes and would crouch down and would not move as long as we were there. Next we placed mesh over the holes to no avail. Last in desperation placed one-way screen. The screen has a white surface (facing toward the lizards), the back side is black and allows for easy observation. >We still could not move quickly behind the screen, or they could see us. The cardboard contained a small hole (5mm x 5mm) and the lizard crouched down because it saw my finger being brushed in front of the hole! I could not believe how good of eyesight they had.
I have owned many reptilian pets in my life, both exotic and native lizards and snakes. When I was younger, I had a large cage outside (6ft x 3ft x 2.5 ft). I kept 3 desert iguanas, 1 chuckawalla, 1 black-collard lizard, 1 Colorado Desert fringe-toed lizard, and 1 desert spiny lizard. The cage had natural rocks, plants, and sand to hide under or burrow in. They were so much fun to watch interact and eat. My firends and I would sit around the cage to watch them eat crickets and vegetables. Once in a while I would feed the black-collard lizard a little lizard. Watched the lizard run like a T-Rex when he chsed the lizard was quite an exciting sight.
One group of lizards that I will never own is the horned lizards (Phrynosoma). They are so observant that constant movement in the room where the cage is located will cause them high stress and eventually lead to death. They have to eat ants and do not survive on any other food. They also need 150-500 ants per day. Because of their nervous behavior, difficult diet to supply , and their hot temperature demands, they are not suitable as pets. Only a super large cage or enclosure outside will suffice. So please if you happen to see a horned lizard in the wild, resist the temptation to keep it as a pet.