User:Hannah T Sherman/Notebook/Biology 210 at AU

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" Embryology & Zebrafish Development" Lab 6 March 22, 2014 Embryology

Introduction: It was hypothesized that the addition of vitamin C would affect the zebra fish. This affect would cause developmental issues and death. To prove this there was a control poetry dish of zebra fish with out vitamin C added and then one with vitamin C. This made it possible to observe the differences between the control and the independent variable and how the zebra fish reacted to the difference. These reactions where observed over two week and looked at every three days. Watching mostly fisical changes and noting if there was difference between the control and independent variable.

Methods and Martials: To start the lab the development of starfish are observed along with the development frog development. For the starfish focus on cleavage and gastrulation. Observing the starfish to see how they develop. Then frog’s development focusing on the same details. Also the absorbtion of the yoke should be noted all the details of development and how they are different from one to the next. This is then again done with chicken development. Then the zebra fish part of the lab was started. To begin before starting the actual experiment reading a published paper about it to better understand it. After that to start zebra fish embryos are observed to see the viability and the stage that they are currently in. Two separate groups are set up in petri dishes that are covered one is the control and the other is the independent variable. 20 healthy embryos that are translucent and are basically at the same stage need to be put in to each dish. Before putting them in to the dishes 20 mls of deer park water need to be added be sure to add the correct mix to each dish. Also be sure to so control first. Using a pipet to move the zebra fish eggs. When changing the water the same additive is used in the water every time. A schedule and procedure should be set up. Over the next two weeks the petri dishes where observed carefully. After 4-5 days remove 10 mls of water along with any empty egg casings. Then add 25mls of fresh water . Save the dead ones in paraformaldehyde. One week after the experiment was started 5mls should be removed along with any dead cases and then 5mls should be added of the test solution or fresh water. 1 to 3 embryos from each dish should be preserved using paraformaldehyde. In the next half week 5mls should be removed and 10 mls should be added. After week one two drops of to feed them add two drops of paramecium repeated only when water is changed. When the second week is up be sure to make any final observations of the fish including measurements. At that point the once who survived will be collected. Once finished all data will be looked over and conclusions will be made.

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Conclusion: In the embryology lab the observation of how vitamin C would affect the zebra fish was observed. These observations where made over a two week period of time in 3 day intervals. There where no early indications of any difference between the control and the independent variable vitamin C except for the fact that the vitamin C fish where a little bit smaller / skinner. The fact that they where skinner became apparent over the first couple days besides that there were no other apparent differences. Also most of the zebra fish survived until March 4rd, which was over a week since the experiment was started. On March 4rd almost all in the control were dead and every one of the vitamin C once where dead. When the first where observed on that day the once in vitamin C where more decomposed almost unidentifiable compared to the once in the control with where pretty intact. This makes the hypothesis hard to prove because not only did the independent variable all die but almost every one in the control died also. Though the hypothesis is slightly supported because the vitamin C variable once where more decomposed meaning they died earlier most likely. This shows that the vitamin C had an affect on the zebra fish or could have because they seemed to have died before the control based on its amount of decomposition. Though this may be true it also has to be noted and taken in to account that many fish die in any circumstances over the first couple of weeks.

“Mini Lab” on DNA sequences Due on March 16, 2014 Introduction: In this lab we looked at DNA samples that were taken from the transects. The DNA sequence was then entered in to a database and identified. This lab helped to better understand not only DNA sequence but also what the actually DNA is of. This also allows a better understanding of the transect by identifying what plant is identified. Materials and Methods: This can be seen in lab 3 below in its methods section. Screen Shot 2014-03-16 at 9.47.11 PM.png

Conclusion: In the lab DNA sequence was use to identify what species it came from and if it was similar to what was originally thought to be there. Both of the samples were from different group same transect because of which samples actually worked. According to the database that was used to decode the DNA the sample that was marked A2-T1-2, which is from the marsh transect is KF358247. This organism is called stenotrophomonas maltophilia. This organism is gram negative and found in many different water environments and soil(Sanchez, Hernandez, & Martinez ).The second one was marked as S1-T1-12 and was called AB581540.This organism is "Chryseobacterium sp. StRB028". This is gram negative and is found in wide range of areas in including fresh water, soil, food sources or, sewage and wastewater. This does not relate to anything that was found previously but it makes sense because it was a sample of soil from the transect(Zamora…). Though neither of these organisms where identified in previous results but it is logical to conclude that they both would be found in the area where the transect was located. We could improve this lab by maybe making this the main focus of the lab or bagging it all together. It was very confusing doing this and jumping back to the embryology lab .

Work cited BLAST: Basic Local Alignment Search Tool. (n.d.). Retrieved March 14, 2014, from

    Blast website:

Maria B Sanchez, Alvaro Hernandez, & Jose L Martinez. (2009). Stenotrophomonas maltophilia drug resistance. Future Microbiology, 4(6), 655. doi:10.2217/fmb.09.45 Zamora, L., Vela, A. I., Palacios, M. A., Domínguez, L., & Fernández-Garayzábal, J. F. (2012). First isolation and characterization of chryseobacterium shigense from rainbow trout. BMC Veterinary Research, 8(1), 77-77. doi:10.1186/1746-6148-8-77

Lab 5 Invertebrates State the question/ problem /Objective: There are two objectives in the “Invertebrates“ lab. The first objective of this lab was to understand the importance of invertebrates. The second objective of this lab was to understand how sample systems have evolved in to more complex systems. The first objective was fulfilled through observing multiple different invertebrates and identifying different characteristics of them. The second objective was fulfilled by observing organism from our transect or from west Virginia. There are 5 main organisms that could have been found in the transect which includes Squirrel’s, Human’s, Stray cats, American Robin and Sparrow. Each of them have different phylum’s. Squirrel’s phylum is Animalia-Chordata-Vertabrata-Mammalia-Rodentia-Sciuromorpha-Sciuridae. Humans are another animal that interacts with the transect there phylum is Animalia-Chordata- Mammalia-Primates-Hominidae. Stray cat also interact with the transect there phylum is Animalia-Chordata- Mammalia-Carnivora - Felidae. The American Robin is one of the organisms which interacts with the transect and phylum is Chordata—Passeriformes -Turdidae-Turdus-T. migratorius. Sparrow’s interact with the transect and there phylum is Animalia-Chordata-Aves-Passeriformes-Passeroidea-Passeridae. There is a food chain or feeding strategy which show which animals in the transect eat each other. There where no quaternary consumers that we really found in the transect one of the tertiary consumer is the american robin. Some of the secondary consumers in the transect where the american robin and earth warms. The primary decomposer in the transect or consumers include fungi, flies among other organisms. Finally the primary producers include plants , leaves among other organism.

Record the specific steps you performed: The lab should be started by observing the acoelomate and the planaria under a dissecting microscope. Be sure to note the movement and how complex or simple it is. Continue by looking at each of the slides that were set out for the lab of the different levels of the development and note the differences of each stage. Repeat this two more times first with pseudocoelomate and then with coelomate Annelida. Observe where the internal organs are located and the layers of muscle. Note what is observed including how it moves. Be sure to take clear notes because the movement need to be noted in the notebook. Next break down the berlese funnel and transfer the contence of the test tub in to a petri dish. Then examine under the microscope and try to identify if there is any invertebrates. Once find the invertebrate identify what group they are from an what they are using a dicotomías key and the internet. Most will probably be anthropoids. Fill in the table in the lab manual with 5 of the invertebrate that are found if you do not find five fill in the spots with the sample that are available during lab. Be sure to not all the important details to fill in the table. Once finished the final part of your experiment requires you to think about what vertebrates may inhibit your transects and then answer the questions about the animals that inhibit the transect. Include all of your raw data: Bugs hs.png

State conclusions and future plans: There are two objectives in the “Invertebrates“ lab are to understand the importance of invertebrates and to understand how sample systems have evolved in to more complex systems. The first objective was fulfilled by observing invertebrates. To fulfill the second objective we observed the invertebrates which came out of the Berlease funnel. There were two that came out of ours which included a fly and a flea. They were observed along with 3 other invertebrates from West Virginia and all of there descriptions are shown in table 1 above. The descriptions show the variance among invertebrates. The second objective was through observing the insects from the Berlease funnel and recording the observations. The second objective was fulfilled by doing the filling out the table above with the information we fulfilled both objectives with the table above. I think next time it would better if this was a more depth lab in to only one of the procedures because by doing so many different procedure each in turn is only skimmed.

Lab 4 Plantae and Fungi State the question/ problem /Objective: There are two objectives in the “Plantae and Fungi “ lab. The first objective of this lab is to allow us to better understand the characteristics and diversity of plants. The second objective is to appreciate the function and importance of fungi. The objective was fulfilled by getting sample from our transect and identify what plant they are. We fulfilled the second objective by observing fungi in the fifth step of the procedure.The one seed that we brought back from the transect we were unable to identify the seed was wrinkled because of this I am not sure if it is monocot or dicot. There seems to be no evidence of flower or spores. The fungi sporangia is when a upward hyphae grows and it will form a black small sphere structure. The reasons why they are so important is because inside it is something called a spore and when it opened it is released. Also it helps to identify what exactly is being looked at.

Record the specific steps you performed: To start the lab you will be going out to your transect with three bags. The first bag will be filled with a leaf litter sample not only containing leaves but also soil samples from different depths and areas. The first bag will be used in the berlese funnel in next weeks lab. Five representative samples should be taken form the transect they should be diverse. Be sure to make down where you got them from. If there is trees or a tree in the transect be sure to take a picture of it or find an old leave and ID it genus. The a seed should be found of any type. When you get back to the lab observe all that you brought back and try to identify the specimens. Continue by observing the lily and it cross section. Identify the different parts of it. Next the vascularization will be identify of each of the plants. Continue by examining the leaves closely. Bryophytes probably appears to have distinct characteristics which are simple photosynthetic appendages. Be sure to note everything about the leaves including shape among other things. Observed the reproductive cycle of bryophyte from the diagram. The moss Polytrichum need to be examined. Gametophytes and the sporophyte should be identified of male and female. While observing be sure to note the life cycle haploid or diploid. Its important to keep in mind that when in reproductive cycle the bryophytes are still dependent on water during this cycle. The next step was to observe the already dissected the lily flower and identify the following parts anther, stamen among other parts. Also identify the different parts inside the soaking seeds. After looking at the lily look at the seeds that where brought back from your transect. Identify the seed and it is monocot or dicot. Next you should observe the agar plates. Start by turning the ager plate over then focus downward to identify the root like hyphae which are called rhizoids which grown in to the agar plate. Be sure to look at the agar plate under the microscope and identify the three groups of fungi draw them in your note book and describe. Next set up the berlese Funnel. Start by pouring 25ml of 50:50 ethanol/water solution in to the flask. After that cut out a piece of the screen so it covers the whole in the funnel. Then tape it in to the funnel so nothing can get through. The funnel then should be placed in the neck in the square-sided funnel bottle. Gently add the contents of the first bag that was collected at the transect to the funnel. Then put a 40 watt lamp 1-2 inches above the funnel then cover it with foil. Leave this for the next week.

Include all of your raw data:

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State conclusions and future plans: In this lab called “Plantae and Fungi “our two objectives where to observe the transect and to gain a appreciation for fungi. We fulfilled our objective first by observing the transect. While observing the transect 5 samples were taken and identified. The five samples show more about the transect and what affects the transect. Through what live in the transect. Also the next lab was set up the berlease funnel which will give an even better understanding of the transect. We fulfilled the second one by hearing about what fungi do during lecture and observing fungi under the microscope. I think next time it would be better if we when more focused on one or two procedure because we are unable to look in-depth in to the experiment we are doing. Lab 3 Microbiology and Identifying Bacteria with DNA State the question/ problem /Objective: There are three objectives in the “Microbiology and Identifying Bacterial with DNA “ lab. The first objective of this lab was to better understand the characteristics of bacteria. The second thing that was observed was antibiotic resistance. The third one addresses how species are identified by sequencing DNA. We fulfilled these three objectives in this lab by observing our agar plates from last weeks experiment and seeing how much growth it had and seeing the different between plates with resistance and without . Then taking a sample from a specific one and making a gram stain. Then by preparing DNA for PCR. Over the last two weeks the hay infusion has change how it smells and looks. This week when we observed how the hay infusion smelled it smell a lot less bitter and a lot more like a camp lake. The appearance. The hey infusion is a lot less cloudy and more clear. I am not completely sure why is smells less potent but maybe it is because it has finally settled or microorganisms in the culture the reason why it is so much more clear is because the particles in the infusion have settled. The different that is seen between the plates that has been seen between the Nutrient plates and the one that has antibiotics on it is to begin with there are smaller colonies or no colonies on the anti biotic plates. There was only one antibiotic plat that had growth and the growth was very different from the rest of the plats. The growth was yellow and all in one spot all together. This could indicate one of two things first that the growth grow all together because that is the only way it can grow on that plat ore that it only grow certain things one the plat. They thrive by being close together. Tetracycline an antibiotic which affects the colony size which is what is put on the anti bacteria plates affected the colonies size by causing no growth or only one large growth. The only one that we found was fusiturm on the 10-2 plat. Tetracycline works by being an inhibiter decreasing growth. This inhibits cholera , rickettsia infections, trachoma. psittacosis , brucellosis, tularemic and other bacteria. Record the specific steps you performed:

Start the lab by checking your hay infusion for one last time note any changes in the hay infusion. After that look at the agar plates from the week before see if there was any growth note the differences in growth between the ones that had antibiotic and antibiotic resistance. The next part of the experiment is to practice observing tiny cells. Start by obtaining a slide that contains different types and shapes of bacteria. Be sure to observe the stained parts of the slide and observe at 40X objective. Then continue by using the 100X oil immersion objective be sure to only use a very little bit of oil . Then be sure to clean the oil objective. After this each group will observe four samples of microorganisms. From the colonies that are observed choose one of the isolated ones. The isolated organism will be used for PCR and species identification.Each of of our sample we will make a wet mount and a gram stain. To make a wet mount use a clean loop and then put a group of water on the slide and then put a cover slip over the slide. Then make another slide doing the exact same thing minus the cover slip and then put a red circle around the sample. Then take the first slide and observe it under the microscope. This should be looked at at 10 X and 40 X. Make observations through it may be hard to see. Next you make a gram stain start by labeling the slide that you made before with the red circle. Once it is labeled heat fix it by passing it through a flam 3 times. Then cover the slide with crystal violet for one minute then rinse off with distilled water. Next cover it with iodine molder for a minute and rinse that off. Flood the slide for 10 to 20 seconds with 95% alcohol it is complete when it comes off colorless. For 20-30 seconds cover the slide with safranin stain and rinse gently. Blot excess water carefully with dry paper towel. Once finished look at it under the microscope at 40X and at the oil objective. Finally we start our PCR which is done by getting a colony of bacteria from the plates and putting it into 100 ul of water in a sterile tube. The it should be incubated for 10 mins 100C then centrifuge. Using 5ul of the supernatant in the PCR reaction. Next week these PCRs will be ran in the gel. 

Include all of your raw data : Table 1 Lab1 5.png

Table 2 Hs Lab1 1.png

State conclusions and future plans: In this lab our objectives where to understand the characteristics of bacteria, antibiotic observing resistance, how species are identified by sequencing DNA. We fulfilled these objectives first by procedure one through 3 by observing the Ager plates and seeing what growth happened. For the most part the majority of the growth was on the Nutrient plates which is was I thought would be the result. Then we characterized three after making a gram stain of this fulfilled the objective of characterizing bacteria and observing antibiotic resistance. Procedure four started to fulfill the last objective by starting to isolate the DNA from the hey infusions. This lab helped use to better understand bacteria. Citation: tetracycline. (2014). In Encyclopaedia Britannica. Retrieved from Lab 2: State the question/problem/objective:

The objective of this lab is to understand how to use a dichotomous key and characteristics of Algae and protists. A dichotomous key are basically keys which you answer another question which leads to the identification of Algea or what every was trying to be found. In this experiment a dichotomous key was used to find what Algae was found and protests, which is where, the they come in. To used the dichotomous key the characteristics must be identified. 

It was observed that the culture smelled like sour swamp water. The culture contained green flooting particles, there appears to be life forms on the top, the liquid is cloudy and brownish, there are brownish growths which are most likely mold. The reason why organism might differ from different parts is because some organisms may only be able to survive near plants or off them which is why it may be only found near plants or away from. One of the organisms were found in the culture was Chlamydomonas. Chlamydomonas fulfills all of the needs of life through different processes. It fulfills the energy through photosynthesis also through the nutrients being absorbed through the cell surface. The next part of being alive is cells this organism is a cell. To be alive information has to be present by having and eyespot it is able to sense light. A key part of being alive is reproduction, which they are involved in both asexual reproduction and sexual reproduction. The ability to adapt to an enlivenment as in evolution, which they seem to be successful with because there are about 500 different species of the Chlamydonas. If we left the hey infusion for another two months it would be come a more prevalent habitat meaning that there would be some organisms that did not survives because of the habitat and other organism would get stronger and more prevalent over time. The selective pressure that affects in the sample are natural selection. The selection of the fittest so if one organism need to live off something that is not available or is limited in the culture then it is highly likely that it will die out.

Record the specific steps you performed: The first part of the lab will be basically looking at a sample of a known organism under a microscope and characterizing it. The second part part of the lab is done by taking a sample from the culture and putting it on a wet mount and look at it under a microscope. Be sure to take samples from different parts and be sure to mark down where each sample was obtained. While looking at it under the microscope find organisms the measure and characterize the organisms. This should be repeated 2 times with each sample if you are in a group of three for a total of 6. The third part of the lab requires the jar to be shaken up then 10 mls of the culture and put it in a tub that contain 10 mls of sterile broth which is labeled 2 once added mix up and take 10mls out of the tub in to the next and repeat two more times. There should be four dilution by the end. After doing that you will be given 8 pieties dishes . Obtain four agar nutrient and four agar plus tetracycline plates. The tetracycline plates need to be labeled with “tet”. Label plates with 10-3,10-5,10-7, and 10-9. Make sure to mark the plates with group identifications. Each of the dilutions should be spread on there corresponding plates

Include all of your raw data: Lab1 3.png

State Conclusions and future plans: The objective of this lab is to understand how to use a dichotomous key and characteristics of Algae and protists. We fulfilled both of these goals by taking samples from the transect and observing them under the microscope. Then using a dichotomous key to identify what we where looking at under the microscope. Though nothing went wrong many things could have including but not limited to mixing up the sample and josling your jar/culture. Something that would be interesting to see next time is to take samples from the culture a week later in the same spot and see if there was any change in the habitat in that sense. In conclusion this lab gave use the chance to learn how to use a dichotomous key to identify Algae and protists. Citations Chlamydomonas. (2014). In Encyclopaedia Britannica. Retrieved from

Lab 1: State the question / problem / objective: This lab was set up to help use understand natural selection and the characteristics of a niche in biotic and abiotic. This lab addresses these objectives through by observing and taking notes on a transact in nature and by us taking samples and setting them up thus allowing us to look at them next time. The topography of the of the transect is grass, rocks, cement and mud on a slight slant. Image 5 lab 3.jpg

Record the specific steps you performed: 1. To start the lab a 20 by 20 feet dimensions are measured for a transact which is marked by putting a popsicle stick in each corner. 2. Once the transact is layed out it should be noted the location to begin then topography among other obserations. 3. Observe and write down in a list the abiotic and biotic components in the transact. 4. Sample should be taken of the soil and ground vegetation using a sterile 50 ml conical tube. Its key to make sure to get reprehensive of the ground 5. Stay at the transact until the instructor has checked your work and said to go 6. 10 to 12 grams of the sample should be weighed out and placed in a plastic jar. The plastic jar should also contain 500 mls of deer park water. 7. Then add in 0.1gm of dried mild. Once added the jar should be gently mixed up for 10 seconds. 8. Once mixed the led should be removed and the jar should be placed in a safe place 9. Be sure to label the jar clearly.

Table 1 Evolutionary Specialization of Members of the Volvocine Line:

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State Conclusions and future plans: The goal of this lab was to help use understand natural selection and the characteristics of a niche in biotic and abiotic. We where able to observe natural selection by looking at the animals that are still around . Biotic and abiotic organisms where observed by observing the transects and identifying what was abiotic and biotic. I don’t know how but if we did this experiment again and one of the goals was to understand natural selection then I would find away to make the goal more clearly understood. Though nothing went wrong many things could of including messing up the mixing of the jar. Something that would be interesting to see if we did is to take samples in different temperatures and weather conditions to see how that would change our results. The different variables would add more veriaty to the results thus giving us a better understanding of the life that the transect supports

Bentley/Walters-Conte/Zeller. (2014). Biological Life at AU. AU. Chlamydomonas. (2014). In Encyclopaedia Britannica. Retrieved from Nozaki, H. (1986). Sexual reproduction in gonium sociale (chlorophyta, volvocales). Phycologia, 25(1), 29-35. Retrieved from volvocid. (2014). In Encyclopaedia Britannica. Retrieved from Volvox. (2014). In Encyclopaedia Britannica. Retrieved from