User:Alexandrea Cavanaugh/Notebook/Biology 210 at AU

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February 27-March 5, 2014--Lab 6: Embryology and Zebrafish Development Objective: In this lab, the stages of embryonic development were examined. The stages of development among different species was able to be compared and contrasted. The effects of altering normal conditions during embryonic development were also able to observed. Steps: Procedure I 1.)A projection of the stages of starfish development was shown to the class. 2.)Egg size, yolk, blastulation gastrulation, protection and larval stages were all observed and noted. Procedure II 1.)A fixed slide of a frog embryo was examined under a microscope. 2.)Egg size, yolk, blastulation gastrulation, protection and larval stages were all observed and noted. Procedure III 1.)A chicken egg was observed under a dissecting scope. 2.)Egg size, yolk, blastulation gastrulation, protection and larval stages were all observed and noted. Procedure IV (Embryogenesis Experiment with Zebrafish Larvae) 1.)Two petri dishes were set up with purified water, and twenty zebrafish embryos were carefully placed in each. 2.)One petri dish--the control-- was placed in normal conditions in the lab (room temperature with normal cycles of daylight). 3.)The other dish was placed in direct light for twenty-four hours a day for the duration of the experiment. 4.)The stages of development of the zebrafish placed within these dishes were observed and recorded. 5.)The development of the zebrafish was tracked every three days over the next two weeks. 6.)After one week, three embryos from each the control and the altered dish were taken for preservation in the name of closer examination and measurement. 7.)Samples of dead embryos were preserved in order for closer examination and measurement. Raw Data: Table 6.1

Table 6.2

Conclusions and Future Plans: This lab enabled us to look at stages of embryonic development in a number of species, thus allowing for comparison between the organisms examined. Embryonic development was able to be seen in action through the growth of the zebrafish, and a deeper level of understanding was gained through the alteration of development and the analysis of its effects on individuals. In further explorations, the fish could be more frequently monitored and better maintained in order to allow for embryos to develop more fully and to survive to grow to adulthood.

February 12, 2014--Lab 5: Invertebrates Objective: In this lab, the final stage of transect examination and analysis was conducted. The significance of invertebrates within this ecosystem was analyzed, and the simple systems of invertebrates were also observed. Steps: Procedure I 1.)Planaria, an acoelomate, was observed under a dissecting scope. Specifically, the method of movement was observed. A prepared cross-section of the planaria was also examined under a microscope. 2.)Nematodes, a psuedocoelomate, were also observed under a dissecting scope, with special attention paid to means and method of movement. A cross-section of the nematode was also observed. 3.)Annelida, a coelomate, was lastly observed. Attention was paid to movement as well as size, position, and orientation of internal organs and muscle layers. Procedure II 1.)The Berlese funnel setup was taken apart, and the invertebrates from the bottle were placed in a petri dish, and examined under a dissecting scope. 2.)Using a dichotomous key, the organisms were identified and noted. Procedure III 1.)Vertebrates within the transect were considered, and a food web was made including vertebrates, invertebrates, and any and all of the organisms observed within the transect or in this study. Raw Data: Procedure I The acoelomates, Planaria, moved by means of gliding. This makes sense because acoelomates are the most primitive and simple of the three forms, possessing no/very simple forms of muscles. The psuedocoelomates, nematodes, move by means of scrunching the four sections of their bodies and muscles. This makes sense because psuedocoelomates are a more complex and advanced life-form. The coelomates, earthworms, moved by extending and contracting their entire bodies. This makes sense because coelomates are the most evolved and complex of the groups examined, meaning they have multiple and more complex muscle structures than the others. Procedure II Table 5.1 (will upload) The organisms measured ranged from 0.2cm long to 2.5cm long, the smallest being the drosophila fruit fly, and the largest being the millipede. Though these organisms were not found in the transect in question, they were found in leaf litter from West Virginia. Out of these organisms, the most commonly occurring proved to be various types of flies. Procedure III Picture 5.1 (will upload) Conclusions and Future Steps: By laying out a food web for the whole transect, the significance of not only the invertebrates but of every organism studied throughout this ongoing examination was recognized and illustrated. The comparison of coelomates, acoelomates, and psuedocoelomates allowed for the analysis of specialized and evolved systems within these organisms. Further study could include analysis of vvertebrates within the transect in order to form a more accurate representation in the food web or another trial of the Burlese Funnel procedure in order to gain a more accurate and wide-ranging representation of the invertebrates present.

February 5, 2014-- Lab 4: Plantae and Fungi Objective: The objectives of this lab coincided with the further study and understanding of the transects. Specifically, the roles and functions of both fungi and plant-life found within the transect was examined and noted. Steps: Procedure I 1.)A leaf litter sample, comprised of topsoil and plant-life living or growing atop it, was collected. 2.)A sample from each of five types of vegetation was collected. 3.)The location and description of each plant collected was recorded. 4.)Using a guide, the plant type and genera was determined for each of the five samples. Procedure II 1.)The moss sample, Mnium, was observed, and its height measured. 2.)The cross section slide of the lily stem was examined, and the height of the lily plant stem was measured. 3.)The vascularization of each of the plant samples taken from the transect was determined and noted. Procedure III 1.)The leaves of the moss were observed closely and carefully in order to identify cuticles. 2.)The cross-section of the angiosperm leaf was examined in order to observe stomata and the arrangement of the mesophyll, palisade and parenchyma cells. 3.)The shape, size, and cluster arrangement of each of the plants with leaves from the transect was also observed and determined. Procedure IV 1.)The moss was examined, this time in order to identify the male and female reproductive structures and the sporophytes. 2.)A dissected lily flower sporophyte was examined in order to observe the anther, the site of meiotic pollen production. The ovary, the site of ovule development, was also examined. 3.)Monocot and dicot seeds were observed. Each of the seeds from the five samples taken from the transect were determined to be either monocot or dicot themselves. Procedure V 1.)Black bread mold was observed under a dissecting scope in order to look closely at the mycelium, individual hyphae, rhizoids, and sporangia of the fungus. 2.)Samples of fungi found in any of the transects were also observed. Procedure VI 1.)A Berlese Funnel was prepared: a.)25mL of 50:50 ethanol/water solution was poured into the flask. b.)A piece of screening to prevent the passage of leaf litter was taped over the bottom opening of the funnel. c.)The prepared funnel was placed in the opening of the ethanol/water-filled bottle. d.)The previously collected leaf litter sample from the transect was placed inside thee funnel. e.)The whole funnel setup was placed under a constructed lighting fixture, and the entire setup was covered with foil and left to sit undisturbed for a week. Raw Data: Procedure I Table 4.1 (still to be uploaded) Procedure II (see Table 4.1) Procedure III Sample 1=Pine tree. This tree has thin pine needles rather than leaves which are clustered closely together while on the tree branches. Sample 2=Holly tree. This tree has small, rounded leaves, about 1-2 inches in length, clustered closely together at the end of the tree branches. Sample 3=Sycamore tree. This tree has larger, branched leaves, situated rather far apart on branches each leaf on its own small stem. Sample 4=Undetermined vine. These vines have small green leaves that are pointed at the ends and are very widely spaced out from each other. Sample 5=Oak tree. This tree has leaves similar to those of the sycamore, large and branched and situated slightly spaced out from each other each on its own stem toward the ends of the tree branches. Procedure IV Fungi sporangia contain spores, which are a means of reproduction for the fungi. This is important because without spore production and dispersal, fungi could not reproduce during the sporophyte stage of alternation of generations. Picture 4.1 (to be uploaded) This looks like a fungus because of the sting-like structure that is the hypha. The individual cells of each hypha are also visible and appear squared off and thin like the cells of a typical fungus. Conclusions and Future Plans: In this lab, plant-life was observed through the examination of leaf and plant structures as well as classifications of genera and vasculation type. Fungi cells and hyphae were looked at in order to study structure and function. Further courses of study could be conducted on different samples of plant and fungal life in the transect.

January 29, 2014-- Lab 3: Microbiology and Identifying Bacteria with DNA Objective: The objectives of this lab directed learning and observation toward a further understanding of bacteria. During the lab procedures, characteristics of bacteria were closely examined and used to identify and distinguish bacteria cultures from the transect. In addition, antibiotic resistance was observed by means of cultures grown on tetracycline-treated agar plates, and bacterial DNA was observed by performing PCR in order to run electrophoresis on a bacteria sample from the transect in the next lab. Steps: Procedure I 1.)Observe the state of the hay infusion culture jar, noting smell, appearance, and any noticeable changes. 2.)Note bacterial colonies that have grown on each of the agar plates prepared in the last lab. 3.)Count and record colony size in order to find the number of colonies per milliliter of dilution on each plate. Procedure II 1.)Observe the tetracycline-treated agar plates, and compare the bacteria on these with the bacteria on the plates without treatment. 2.)Note similarities and differences between the treated and non-treated plates. Procedure III 1.)Obtain a fixed slide with different bacteria samples. Observe the samples, moving up to the 100x objective. 2.)Scrape a small amount of bacteria from a particular colony in an untreated agar plate with a sterilized loop, and mix this with a drop of water on a slide. Place a cover slip over the sample. 3.)Mark the colony the sample was taken from on the bottom of the plate. 4.)Observe the organisms under the microscope, adding a drop of stain to the side of the cover slip if no microorganisms are visible. 5.)Repeat this process to make three more slides, one more from a different colony on the untreated plate and two more from different colonies on the antibiotic-treated plates. 6.)Use these four slides to do a Gram stain of the organisms: a.)Ensure that each slide is labeled. b.)With the bacterial smear side up, pass the slide through a flame three times in order to heat fix the sample. c.)Cover the smear with crystal violet for one minute using a staining tray, then rinse using a bottle of water. d.)Cover the smear with Gram's Iodine for one minute, then rinse gently. e.)Decolorize the smear by rinsing with 95% alcohol for 10-20 seconds, then rinse gently. Repeat until the solvent rinses without color. f.)Cover the smear with with safranin stain for 20-30 seconds, rinse gently. g.)Blot excess water carefully with a paper towel, let air dry. 7.)Observe the stained slides, moving gradually up from the 4x to the 100x objective. Observe and record cell morphologies. Procedure IV 1.)Prepare DNA samples for PCR: a.)Transfer one colony of bacteria to 100microliters of water in a sterile tube. b.)Incubate mixture at 100 degrees for ten minutes and centrifuge. c.)Use 5microliters of the supernatant in the PCR reaction. Raw Data: Procedure I Archaea species will not have grown on the agar plates because they tend to live in only extreme environments, very unlike those found within the transect. The appearance and smell of the hay infusion culture might change week after week because bacteria levels may grow and shift as one species may grow more than another and vice versa. This could cause changes in both smell and appearance, causing the water to appear ore murky and cloudy and the smell to become more pungent. Table 3.1 (to be inserted later) Procedure II The agar plates that were not treated with antibiotics had a much more diverse appearance of bacteria, ranging in color from beige to yellow to bright yellow-orange. The antibiotic-treated plates, on the other hand, showed only one type of bacterial growth. This indicates that many more bacterial species are affected by tetracycline and only one found in this transect was not affected. Tetracycline seems to be very effective on the great majority of bacteria. No fungi samples were shown in this culture. Tetracycline is a protein synthesis-inhibitor, meaning that it prevents the synthesis and expression of proteins needed for the processes of life. Typically, tetracycline is used in urinary and intestinal infections, but its ranges of use cover a broad spectrum of infections. Procedure III Table 3.2 (to be inserted later) Figure 3.1 Figure 3.2 Figure 3.3 Conclusions and Future Plans: This lab demonstrated the differences in bacterial DNA even in just the differences in antibiotic resistance between species of bacteria. In addition, the rapid growth and spread of antibiotic-resistant bacteria was illustrated. A possible path of further study would be to test out different types of antibiotics with the same sampling of bacterial species in order to see which antibiotics were most effective and which bacteria were resistant to which types of antibiotics.

January 22, 2014-- Lab 2: Identifying Algae and Protists Objective: The objectives of this lab were in line with further understanding of the niche and function of organisms within an ecosystem. Specifically, this lab illustrated the proper use of a dichotomous key in identifying organisms like the algae and protists observed i the lab. In addition, the characteristics of both algae and protists were also examined and observed. Steps: Procedure I 1.)A wet mount taken from a sample of known organisms was prepared and examined, first at the 4x objective, then at the 10x. 2.)Once an organism was found, it was noted and drawn. 3.)The dichotomous key was utilized to characterize and name the organism based on its characteristics upon observation. 4.)Steps 1-3 were repeated for one more organism on the slide. Procedure II 1.)Without disturbing the jar, a first impression of appearance and smell of the hay infusion culture jar created in Lab I were observed and recorded. 2.)Slides were prepared for observation with two samples from the jar, each sample from a different location within (ex. One from the top of the water, one from the very bottom of the jar.) 3.)Three organisms from each sample were observed and drawn, and the dichotomous key was used in an effort to possibly identify any of the organisms. Procedure III 1.)A 100 microliter sample of mixed solution from the hay culture jar was taken and transferred to a tube, labeled 2, of 10 milliliters of broth for dilution, then swirled to thoroughly mix the liquids. 2.)A 100 microliter sample of solution from tube 2 was then taken and diluted in 10mL of broth in a tube labeled "4" and mixed well. 3.)A 100 microliter sample of solution from tube 4 was then taken and diluted in 10mL of broth in a tube labeled "6" and mixed well. 4.)A 100 microliter sample of solution from tube 6 was then taken and diluted in 10mL of broth in a tube labeled "8" and mixed well. 5.)A 100 microliter sample was taken from each tube and each placed on its own nutrient agar plate labeled accordingly to dilution. 6.)A 100 microliter sample was taken from each the 2, 4, and 6 tubes and each was transferred to a nutrient agar plate treated with tetracycline and labeled according to dilution. 7.)The agar plates were incubated at room temperature for a week until the next lab. Raw Data: Procedure I Figure 2.1-Amoeba (can't add pictures yet) Figure 2.2-Euglena (can't add pictures yet) Procedure II Figure 2.3-Organisms from Sample 1 (top of the hay infusion) (can't add pictures yet) Figure 2.4-Organisms from Sample 2 (bottom of the hay infusion) (can't add pictures yet) After a week of incubating, the culture smelled very faintly of mold and must. A very thin film had developed across the top of the water, a little bit of mold or algae had started to grow on the sides of the jar, and the water had become slightly brown and cloudy. Of the two samples examined, one was taken from the very surface of the water, while thee other was taken from the very bottom of the jar. The location of these samples is relevant because different organisms thrive in different environments, no matter ho slight the difference or proximal difference may be. For example, some organisms live toward the surface of the water or cling to the walls of the jar, while others live closely to the plant matter that had settled to the bottom of the jar. Given any of these single-cell organisms, the needs of life can be met and described: these organisms are made of cells, reproducing most likely by dividing themselves in two. By means of both this reproduction and horizontal gene transfer, evolutionary changes may be made. No evidence of genetic information was necessarily evident, however reproduction of cells depends upon the ability to copy and transfer genetic information, so it can be assumed that, because they do, in fact, reproduce, they do, also, have genetic information. In terms of energy creation and conversion, none of these organisms appeared to be photosynthesizing cells, so it can be assumed that they intake energy from some form of various food sources. If the hay infusion culture was observed for another two months, it can be assumed that mold, algae and bacteria growth would continue to increase until the ecosystem hit the limit of its carrying capacity and could not sustain any more life. The compositions of these samples were affected by the environment from which they originally came. The material used to create the hay infusion culture was taken from transect 2, an area shaded by trees, back from the sidewalk in the grass, and covered with a layering of some type of ivy across the ground. These conditions allow for much different organisms to grow and thrive than would an open, frequently sunny area because there are different resources available. Procedure III Figure 2.5-Set-up of nutrient agar plates (can't add pictures yet) Conclusions and Future Plans: The examination of just a few microorganisms from these samples demonstrates the immense diversity that exists among them throughout the planet. The characteristics of algae and protists, though diverse even in this tiny sample size, were able to be observed in this process in addition to size, manner of locomotion, shape, etc. The use of a dichotomous key to identify known samples of protists and algae was very helpful in that it helped to provide experience with distinguishing between features and determining what is notable about an organism. Further explorations on this topic could include working to characterize and name the more diverse organisms found in the hay infusion cultures.

January 14, 2014-- Lab 1: Biological Life at AU Objective: The objective of this lab was to understand natural selection and to understand the characteristics, both biotic and abiotic, of a niche. To achieve the, three different organisms from an evolutionary line of green algae were observed and compared. In order to begin to understand the characteristics of a niche, designated transects of land were mapped out and observed. Steps: Procedure I 1.)Prepare a slide of a living sample of Chlamydomonas, adding Protoslo to ease observation. 2.)Observe structural traits such as structures for movement, number of cells, size of cell colonies. Record findings in Table 1.1. 3.)Repeat steps 1 and 2 with a living Gonium sample. 4.)Repeat steps 1 and 2 with a living Volvox sample. Procedure II 1.)Find a 20ftx20ft transect of land, predesignated. 2.)Note the characteristics of the transect, any landmarks, topography, biotic and abiotic factors within it. Record these observations, and draw a map of the area. 3.)Use a sterile 50mL conical tube to take a sample of soil and vegetation at ground level. 4.)Create a hay infusion culture by

  a.)Place 10-12 grams (weighed) into a large plastic jar.
  b.)Add 500mL of Deer Park water.
  c.)Add 0.1grams of dried milk.
  d.)Screw on the cap, shake the mixture gently for about ten seconds.
  e.)Remove the cap, label the jar so that it may be distinguished from others, leave to sit for a week. 

Raw Data: Procedure I

Table 1.1- Evolutionary Specializations of Members of the Volvocine Line: (can't insert table...)

Th significance of cell specialization across these three genera shows the increasing complexity of the algae as it evolved. Evolution does not always move toward increased complexity. The example of these green bacteria is not representative of this because most evolutionary functions that have acted on the species have moved it toward greater complexity.

Procedure II

Figure 1.1- Transect 2: (picture of transect)

Transect number 2 is located near the Wesley Theological Seminary. It is in thee middle of a grassy area, on a slight hill. The extent of the transect is covered with some type of ground-ivy, and is shaded by trees. There are three trees located inside of the transect, and a small piece of trash was found within it as well. Abiotic components found in this transect were soil and the piece of trash, aluminum foil. Biotic factors found within the transect were the three different types of trees, believed to be a holly, a sycamore and an eastern pine, and the ivy ground-covering. Conclusions and Future Plans: The observation of the three members of the Volvocine line of green algae illustrate the increasing complexity of a line of evolutionary change. This, however, is only one example of the workings of evolution, many of which do not work toward increasing complexity of organisms. Observing the land transect showed the distinction between biotic and abiotic factors in a given area. This transect will be monitored from time to time in order to record any possible changes that could have effects on the niche of each factor within it. With careful monitoring, it will be better observed just how the members of a small ecosystem such as this interact with each other and any outsiders that enter it.

Good first lab entry. Please start each entry with a date and title in bold and add new information or the next lab entry to the top of the page pushing the old information downwards. Soil is an abiotic component. There are some typing errors. Please upload images when you can. SK

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