User:Adam Katzenberg/Notebook/Biology 210 at AU

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Lab 5 02/24/2014

Procedure I:Observing Acoelomates, Pseudocoelomates, and Coelomates

The three types of worms we observed in lab were the acoelomates Platyhelminthes or flat worms, the pseudocoleums, Nemotodas, and the coelmates, the earthworm. Because the flat worms are acoelomates they do not have a fluid filled cavity or coelem and are bilaterally shaped, I observed their movement to be stagnant. The Nemotodas are pseudocoleums, emaning they have an incompletely lined body cavity. I noted that their movement was a slow slither through the soil. However, the earthworm which is a coelmate, have a fully lined fluid-filled coelem. I observed their movement to be very mobile in soil as if they were gliding through the dirt.

Procedure II: Analyzing the Invertebrates Collected with the Berlese Funnel

Table of Organisms: 1. Ground Spider LL- Arachnids- 1 mm in length. The ground spider has no wings, no antennae, 2 body segments, and 8 legs. 2. Millipede- 4.5 mm in length. The millipede has no wings, 1 pair of antennae, several body segments, 2 pairs of legs on each side, and a round body. 3. Centipede- 5 mm in length. The centipede has no wings, 1 pair of antennae, several body segments, 1 pair of legs on each side, and a flat body.

  • We were not able to see anymore organisms.

The range of the sizes of organisms that we collected in our Berlese funnel were from the 1 mm length spider to the 5 mm length centipede. The largest organism we found was the centipede. The smallest organism we found was the spider. The most common organisms in leaf litter are centipedes.

Procedure III: Vertebrates and Niches

Five animals who might inhabit the transect:

1. American Robin- Chordata>Vertebrata>Reptilia>Aves>Turdus>Turdus Migartorius Biotic Characteristics that would benefit-Food such as insects and seeds Abiotic Characteristic that would benefit- Dirt for food to live in, man made feeders hanging from the trees for food

2. Song Sparrow- Chordata>Vertebrata>Reptilia>Aves>Melospiza>Melospiza Melodia Biotic Characteristics that would benefit-Food such as centipedes and arachnids and seeds Abiotic Characteristic that would benefit-Dirt for food to live in, man made feeders hanging from the trees for food

3. Voles- Chordata>Vertebrata>Mammalia>Eutheria>Aborimus>Aborimus Albipes Biotic Characteristics that would benefit- Plants and seeds for food. High bushes and trees for cover from predators. Abiotic Characteristic that would benefit-Dirt to house food

4. Wood Frog- Chordata>Vertebrata>Amphibia>Anura>Ranidae>Lithobates>Lithobates Sylvaticus Biotic Characteristics that would benefit- Food such as insects, high plants to keep hidden from predators, brown trees for camouflage Abiotic Characteristic that would benefit- Dirt to house food, park bench to hide under and for shade

5. Eastern Garter Snake- Chordata>Vertebrata>Reptilia>Lepidosauria>Colubridae>Thamnophis>Thamnophis Sirtalis Biotic Characteristics that would benefit- Noles and Wood frogs for food, plants to shade and hide from predators Abiotic Characteristic that would benefit- Dirt to attract smaller mammals for food, park bench to shade when hot and to stay hidden



Lab 4 02/21/2014


Procedure V: Observing Fungus Fungi Sporangia are small, black, globelike structures. Inside the sporangia are spores which are released when the sporangia open. They are important because it allows the fungus to spread to different environments. When the spores land on new environment more fungus is able to divide its cells and grow into more fungus. The most common fungus that uses this type of reproduction are the zygomycetes.


I think its a fungus because it contains spores and mycelium which are fungi characteristics. Also it was obtained from moldy bread, and mold is a known fungus.


Lab 3 02/21/2014 The purpose of this lab was to understand the characteristics of bacteria, observe antibiotic resistance, and understand how DNA sequences are used to identify species. In order to understand the characteristics of bacteria, we will be observing three different species of bacteria under a microscope and observing their characteristics. These characteristics include, shape such as bacillus, coccus, and spirilium, size, and whether or not they are gram-positive or gram negative, which provides insight into which have peptidoglycan in their cell walls.

Identifying and Studying Bacteria

Yes, I think archaea species will have grown on the agar plates. This is because of their extremophilic nature, meaning they can live in extreme environments such as hydrothermal vents near methane gas, and in environments with high concentrations of saline. Therefore, they should be able to grow on our lab agar plates.

Procedure I: Quantifying and Observing Microorganisms After a week of incubation, the agar plates were observed. The amount of colonies on each plate were counted and inputted into Table 3.

I noticed that the smell of our hay infusion culture decreased in potency. This could be due to the increased life that has grown inside of the jar eating the spoiled milk that initially made the hay infusion smell. Another reason could be that the smell from all of the hay infusion cultures overtook the smell of the room so it seems that there was no smell apparent.

Table 1 100-fold.jpg

Procedure II: Antibiotic Resistance The color of the colonies, the amount of the colonies, the shape of the colonies were observed from each plate and compared to the “tet” positive colonies on the agar plates.

Yes, there was a clear difference in the colony types between the plates with vs. without antibiotic. The plates with the antibiotic contained only white, yellow, and orange colonies, whereas the plate without the antibiotic had white, yellow, and purple colonies. This indicates that purple bacteria colonies are not antibiotic resistant, and white, orange, and yellow bacteria colonies are antibiotic resistance. Furthermore, orange colonies needed the antibiotic in order to grow. The effect of tetracycline on the total number of bacteria lowers the amount of colonies on the agar plates, this is because the bacteria have to be antibiotic resistant. For example, on the dilution plate 10^-3 with agar nutrient there was an uncountable amount of bacteria colonies growing, in comparison on the 10^-3 with agar nutrients and tetracycline, there were around 48 X 10^3 per ml. Although there were still a lot on the tetracycline plate it did not match the amount of on the non tetracycline plate. We did not have any fungi growing on our plates so I am not sure how the tetracycline affected fungi. I found that three species are unaffected by tetracycline, orange, white, and yellow. This is because they appeared on the agar plates with tetracycline.

Figure 1- Agar Plates.jpg

Tetracycline inhibit enzyme reactions needed for bacterial cells. It works by binding to 30S ribosome of bacteria, and then stops the attachment of the aminoacyl tRNA to the RNA-ribosome complex. The types of bacteria that are sensitive to this antibiotic are E. coli, Haemophilus influenzae, Mycobacterium tuberculosis, and Pseudomonas aeruginosa.

Table 2-ColonyInfo.jpg


Procedure III:Bacteria Cell Morphology Observations Out of the colonies obtained, three were chosen to have the DNA analyzed, wet mounted, and Gram Stained. Two were chosen from the regular plates and one was chosen from a “tet” positive plate. All three colonies were observed on a wet mount and the observations were noted. Afterwards a gram stain was made for each colony. This process began by first labeling slides. Then the colonies were heated and smeared onto the labeled slides. For each slide the same process occurred. The slide was covered with crystal violet for one minute, and then rinsed with water. Then the slide covered with Gram’s Iodine mordant for one minutes and rinsed gently with water. Next the slides were flooded 95% alcohol for 10-20 seconds in order to decolorize the slides. The colony was then stained with safranin for 20-30 seconds and rinsed. Finally, once the slide was dried it was observed under 40x magnification under a microscope. If the bacteria were gram positive a blue color would show, conversely, if the bacteria were gram negative a pink color would appear.


Procedure IV: Start PCR Preparation for DNA Sequence Identification After the group characterized the three bacteria from their Hay Infusion Culture. One colony with the best characterization was chosen to be DNA sequenced using PCR reaction. The DNA was isolated using two primer sequences 27F and 519R in order to amplify the 16s rRNA gene. Process required obtaining a sterile tube and adding the colony in 100 uL of water. The tube was then incubated at 100°C for 10 minutes and centrifuged. The 5 uL of the supernatant in the PCR reaction was sent to a lab for DNA sequencing.


Lab 2 02/09/2014 The purpose of this lab is to understand how to use a dichotomous key as well as understanding the characteristics of algae and protists. A dichotomous key is a tool used in order to identify any group of organisms. It works by observing key features of an organism and comparing it to verified data in order to figure out what group the organism belongs to. This will be done by observing Red algae, Brown Algae, and Euglenids and identifying each based on a dichotomous key. I hypothesize that the red and brown algae will be identified by its color and the euglenids will be identified by its flagellum.

Procedure I: How to use a dichotomous key First we made a wet mount of the sample with the known organisms and observed with the microscope at 4x and 10x. Then we characterized the organisms and described their size using the ocular micrometer. We then obtained a key with eight known organisms and used the key to determine the identity of the organism. We repeated these steps with a second organism as well.

Procedure II:Hay Infusion Culture A week after the Hay Infusion Cultures were prepared, they were observed, describing the smell and appearance. A few samples were gathered from two different niches within the jar for microscopic observation. The location from where the samples were taken was then noted. A dropper was used to place a small drop of liquid from the culture onto the microscope slides and a cover slip was placed on top. Three organisms from two different niches were observed. While observing the organisms under the microscope a picture was drawn of each, the size was observed, the mobility was noted, the organism type was identified, and whether each organism utilized photosynthesis as a food source was described. When we first created the Hay Infusion Cultures Jars the water was clear, the plants were alive, and the smell was clean. After a week, when the observed the jars there was a stark transformation. The water was murky and dirty, the smell of the jar had a strong spoiled milk smell. A thin sludge film covered the top of the water in the jar, and a thick sediment of sludge settled at the bottom of the jar. The plants were no longer vibrant greens, they turned brown and dead with the water.

The organisms near vs away from the plant will be different because on a microscopic scale they are two different habitats therefore the species inhabiting the area will be different. For example, the species closer to the plant might be using the plant as a food source, whereas the species away from the plant may eat the denser sediment on the bottom of the jar.

We obtained our samples from the top film layer and from the bottom sediment sludge of the hay infusion cultures. Hay Infusion Culture Species.jpg

3 Organisms from Bottom Niche- Gunk on Bottom 1. Pandorina- 250 micrometers, green algae, pandorina genus, mobile using flagella. 2. Paramecium Aurelia-140 micrometers, paramecium genus, mobile using cilia. 3. Paramecium Bursaria- 90 micrometers, protozoan, paramecium genus, mobile mobile.

3 Organisms from Top Niche-top film layer of water 1. Arcella- 60 micrometers, Arcella genus, mobile 2. Chlamydomonas- 12 micrometers, green algae, Chlamydomonas genus, mobile with flagella 3. Chilomonas - 32 micrometers, chryptophyte, protozoa, chilomonas genus, mobile with flagella

Paramecium Bursaria Freemen describes five things essential to life; energy, cells, information, replication, evolution. The paramecium bursaria meets all of the criteria. It is a protozoa, meaning it is motile and is unicellular. Within the cell there is a nucleus that contains its DNA or information. Because it is a paramecium it replicates asexually through binary fission. It consumes algae for energy therefore it is heterotrophic.

If the hay infusion culture had been observed for another two months I would predict that the water would be darker in color because of the constant replication of the organisms already there the water will become denser with algae and protozoans. The smell would be harsher because of the build up of algae. I would also predict that mold will start to develop on the top layer of the water, because it is a habitat conducive for mold; wet, warm, and plenty of food for it to consume.

Selected pressures that affected the composition of the culture would include size of the environment. Relative to the size of the organisms, if it is considered a big environment for smaller organisms there is more space for different habitats such as near the plants, top of the water, or bottom of the water, however if the organisms are bigger there is a higher competition for food sources and habitats therefore the fittest organisms would survive. Another selected pressure would be reproduction for the organisms. If there is not a lot of room for bigger organisms as they reproduce there is more competition for food and space therefore not many of them would survive at a given time.

Procedure III: Preparing and Plating Serial Dilutions In order to view the bacteria growing in the transect, a serial 100-fold dilutions were made from the cultures and then inoculated on agar petri dishes. The bacteria would take a week to grow. In order to make the dilutions, four tubes of 10ml sterile broth were obtained and labeled 2,4,6,8. Then, four nutrient agar plates and four nutrient agar plates with tetracycline were collected. Each plate from the two groups were labeled 10-3, 10-5, 10-7, 10-9 with the tetracycline plates also labeled with the word “tet”. The Hay Infusion Culture was then swirled in order to maximize the chances of obtaining different types of bacteria. Then, 100 uL were then taken from the culture and aseptically added to the tube labeled 2 for a 10-2 dilution. Tube 2 was agitated and then again 100uL was taken from tube 2 and added to tube 4 for a 10-4 dilution. This procedure was repeated two more times to make the 10-6 and 10-8 dilutions. Then a 100 uL from the 10-2 tube was taken and placed on the surface of the agar plate labeled 10-3. The same process was used on the “tet” agar plate labeled 10-3. This process was repeated for the rest of the tubes with corresponding agar plates. The agar plates were then placed in an incubator that was set at room temperature for the week.

Serial Dilutions.jpg


Introduction Bio 2010 01/31/2014 The objectives of this lab were to understand natural selection and understand the biotic and abiotic characteristics of a niche. In order to understand natural selection in this lab we observed characteristics of three different algae families; Volvox, Chlamydomonas, and Gonium. By observing these algae will give us a better understanding of how a simple cellular organism evolved due to natural selection and are able to survive today. These organisms will give us context into natural selection occurring on bigger organisms as we move on in lab. Similarly, in order to understand biotic and abiotic characteristics of a niche, we observed a transect. By observing a transect will provide insight into how the biosphere works by focussing on biotic and biotic components of a home to many organisms.

Procedure 1: The Volvicine Line In this experiment we observed an isogamous, single celled, motile alga called chlamydomonas. We prepared a slide of a living Chlamydomonas and observed it under a microscope, noting the shape, stigma, nucleus, chloroplast, and flagella. Then we observed a living culture of a Volvox and Gonium on a slide and annotated our observations.

Table 1 Evolutionary Specialization of Members of the Volvocine Line From samples of the three algae a number of characteristics were observed: Number of cells, colony size, functional specialization, and reproductive specialization. The results are contained in the table below.

Characteristic: Number of Cells, Colony Size, Special Function, Reproduction Specialization Bold text Chlamydomonas: 2, 6, Motile, Isogamy

Gonium:3, 5, Still, Oogamy

Volvox:1, 1, Still, Oogamy

Conclusion This lab provided insight into natural selection. By comparing each of these cells it was apparent how evolution by natural selection had a profound effect on the development of theses cells. For example the chlamydomonas ability to photosynthesize in order to survive, give it the ability to live wherever sun and water are available. In order to further this experiment it would be interesting to view one of these single celled organisms next to a cell of a complex organism side by side, and note the differences. This would provide a good contrast on how evolution effects more developed organisms compared to simple ones.

Procedure II: Defining a Niche at AU The lab was split up into five groups of two. Each group was assigned a different transect on the American University Campus. The groups then proceeded to their transect with a 20 foot long string, notebook, and a 50mL sterile conical tube to collect a soil/surface plant sample. Once at the transect the groups were asked to describe general characteristics of the transect, draw a map of the transect, list abiotic and biotic components of the transect, and obtain soil and vegetation sample from the transect using the 50mL sterile tube.

Characteristics of the transect: The transect is located at the Washington Seminary at American University. The transect is a circular plot of plant growth with trees growing out from it. Under the plant growth was rich damp soil which would be perfect conditions to support life, however a weed type growth of plants suffocate the area. Within the growth there were fallen leaves from the trees, sticks, and roots entangling the plants. A tennis ball lay on the outskirts of the circular plot, and a ball of aluminum foil was found in the center of the plot. Surrounding the transect is a grassy hill, the road, and stone park benches.

A list of the abiotic and biotic life we see is listed in a table below:

List of Abiotic and Biotic life:

Abiotic: Soil, Tennis Ball, Aluminum Foil, Park Benches

Biotic: Weed type growth, Trees, Roots, Grass

Conclusion This lab provided insight into the different micro-environments that can occur on any plot of land within a 100 yard radius. At the transect we viewed a whole ecosystem occurring on a 20 ft by 20 ft plot of land. This included seeing a variety of plants and animals that lived in that small plot of land. Using this information as a starting point, it will be interesting over the course of the semester to continue observing the transect and see the different organisms that are currently inhabiting that niche.