11/15/18

Marci Jordan

Biology Lab 1105-31

Pre-Lab/Lecture

Morphology and Scientific Presentations

Taxonomy is the discipline of biology that names and classifies organisms.  The science of naming organisms is often credited to Carolus Linnaeus and is used universally to improve communication.  Even so, there are many ways to classify and occasionally a morphological classification scheme may not agree with a molecular or evolutionary classification scheme (phylogeny). Our goal is to learn the classification schemes used for Ciliates so that we may recognize the ciliates we collect from the soil.

The location of the of the oral cilia is used to classify ciliates Two large groups of ciliates are based on the location and characteristic of the cytostome. Ventrostomatous patterns of oral cilia describe the cytostome to one side of the cell or away from the apex. The Protostomatous pattern of oral cilia describe the cytostome at the anterior end (apex) of the cell.

Ciliates are classified by the types and arrangements of kinetids. Kineties are rows of cilia. The cilia are attached to the cell through a kinetosome, also called a basal body, which connects to bands of cytoplasmic fibril proteins. The complete structure is called a kinetid which are organized in rows called kineties. A kinetid has one (monokinetid) or sometimes 2 (dikinetid) cilia,  the subpellicle basal body region (kinetostome), and a complex array of fibrils beneath the cell surface and connected to the kinetostome.

The arrangement of the infraciliature is used to classify ciliates. One common scheme for classification is to compare the infraciliature.  This may not be easy to do with the light microscope, but understanding the characteristics will allow you to look for these patterns. What is infraciliature? Just like “infrastructure” refers to the structures and systems that are needed to keep a country or organization running properly, “infraciliature” refers to the organization and distribution of ciliary structures on the ciliate. Some cilia are located on the oral groove, also called the oral apparatus and or cytostome, (oral cilia), some near the oral groove (paroral cilia), some on the “body” of the cell (somatic cilia), and some on the caudal, or posterior tail, region (caudal cilia). The cilia are arranged in rows and are either embedded in the membrane as single (mono), pairs (di) or groups (poly) of cilia. Tufts of many cilia together are called cirri.

Information about Final Presentations

Scientific Communication is an essential skill for success in science.  One of the major goals of CILI-CURE is to give students opportunities to learn and practice 2 of the standard ways that science communicate, Scientific Reports and Scientific Presentations. Work with your team members to plan your presentation.  Do not just divide the work up into individual tasks. Work as a team so that you all contribute to the entire product. Pages in the pre-lab include some resources about science presentation skills.

Purpose

The purpose of this weeks lab is to further discover, document, and identify ciliates. The next lab meeting will be presenting all the information so it is important to gain all last minutes information this week in lab.

Objective

Objectives include ciliate biology, ciliate classification, recognizing failure in science, finding a ciliate to isolate, obtain a live image of ciliate, staining ciliate, and presentation design. All these objectives are to compact all information necessary to make a successful power point presentation.

Hypothesis

I think students will be successful today as this is the third try to obtain some ciliates. Also, we had a container with contents which held a lot of ciliates for students who could not find any in their soil. Therefore, everyone should have some photo evidence of a ciliate before they leave.

Procedures

Soil Texture

1. Use a ruler to measure the % sand, silt, and clay
2. Determine the % of each soil particle
3. Record soil type in notebook

Ciliate Identification

1. Obtain drops of soil sample from the non-flooded plate with a pipette. Be sure to try and not get any debris.
2. Search and find ciliate cells at varying magnification levels
3. Use Methyl Cellulose to slow down the ciliate and capture a picture
4. Use Iodine to stain the ciliate to have it appear more visible
5. Once a good picture is obtained try and identify based on morphology characteristics

Presentation Design

1. Define your problem/purpose- what do you want the audience to know
2. Sell your work- use a question, figure, or creative title
3. Presentation should be mostly visuals and consise
4. Everyone should contribute equally

Data

Soil Texture

My soil texture came out to be “Loam”

Ciliate Identification

So I tried to discovery more ciliates in hope to better identify or discover completely new ciliates in my soil. However today in lab was not my day as I had no luck finding many ciliates and when I did I could not photograph them. Even with Dr. Adair’s help she could not find any. Luckily I got really good detailed photos and videos of a ciliate last week in lab which I included in my last notebook post and I will be using that for my presenation.

Presentation Design

1. Introduction
   • What/why were talking about this
   • Define a ciliate
   • Backround information
2. Material and Methods
   • How we got soil
   • Non-flooded plates
   • Soil pH/ texture
   • Methyl cellulose and iodide
3. Marci
   • Add figure/pics
   • Individual pH/texture
   • % Water
   • Identify class/ciliate (why)
   • Ciliate Characterization
4. Lou
   • Add figure/pics
   • Individual pH/texture
   • % Water
   • Identify class/ciliate (why)
   • Ciliate Characterization
5. Riley
   • Add figure/pics
   • Individual pH/texture
   • % Water
   • Identify class/ciliate (why)
   • Ciliate Characterization
6. Discussion
   • Connect to tree
   • Big picture
   • Connect pH level to ciliates found

Conclusion

Overall, students have had plenty of time and resources to gain at least one ciliate. After these three lab days of preparations and the lab meetings even before while consisted of collecting the soil has now come full circle. Each student has background knowledge of ciliates, information on what we are doing and why, and how to identify ciliates which we have collected. Data such as water %, soil pH, and soil texture has also been measured and can be used for presentations.

Next Step

Following basically our last lab I am very excited to work on my presentation. Although I was hesitant and nervous before, I know feel prepared and equip to form and present this presentation. I’m excited to use the Thanksgiving Break to lay out all my data and research to piece it together and find what it all means. After that I will combine my findings with my two lab partners to create our presentation. Once we return to Baylor we will practice our presentation and present it on Thursday in class. I’m interested to see how our findings will compare to others in the same class.

11/08/2018

Marci Jordan

Biology Lab 1105-31

Pre-Lab/Lecture

Learning Theory: Creating Questions

Cilliates v.s Protistis/ Traditional Ciliates

Ciliates are different from protists as they have cilia. Cilia help them move and the diversity of infraciliature or structure of somatic and oral kinetids. Next, they have 2 types of nuclei; Nuclear dimorphism or dualism which are macronuclei (MAC) or micronuclei (MIC). Lastly, they reproduce asexually and sexually. Asexual reproduction is called fission and sexual reproduction is called conjugation; it allows the cells to exchange and reshuffle DNA.

Dichotomous Keys and Ciliate Structure

Dichotomous keys are tools that help to classify organisms by a series of questions that lead to a group or species with a defined set of characteristics. We will use a variety of resources to identify ciliates from the soil. Classification and taxonomy are always being updated, so sometimes the same organism is named differently in one source or moved to another group when further information becomes available. An atricle provided in the pre-lab shares an article titled “Ciliates” which has detailed diagrams of the ciliate structure.

Classifying Life and Taxonomy

The science of taxonomy is always changing as more species are described and new characteristics are compared.  You will find that not all classification schemes match. We will use the recent classification scheme that places all organisms into 7 Supergroups based on DNA similarity.

Where are the Ciliates in this scheme?? Where am I in this scheme?

Opishokonta –> Amoebozoa –> Excavat –> Archaeplastida –> SAR: Stramenopila, Alveolata, Rhizaria

Sometimes taxonomists put 3 groups together (The SAR Supergroup).

A recent article has described the diversity of the Phylum Ciliophora using genetic information.  Our research goal is to contribute to this database by capturing and identifying soil ciliates.  We will use the non-flooded plate method to observe and capture the ciliates.  We will characterize ciliates using size, shape, cell structures and ultimately, the DNA sequence of the variable region of the 18S ribosomal gene or other sequence that can be compared to a database of known ciliates DNA sequences.

A figure on the pre-lab demonstrates the monophyletic grouping of ciliates.  This scheme is based on all ciliates sharing a common ancestor. The phylum is divided into 2 subphyla: Intramacronucleata and Postciliodesmastophora.  The Classes of ciliates are in all caps. The article Ciliates D. Lyn Encyclopedia of Microbiology.pdf describes 11 major Classes. Read the section on the 11 Classes and review Figure 6.   Do you recognize any of the ciliates that you have already studied?

Ciliate Isolation and Characterization

During lab students will observe and record their observations from a close inspection of your non-flooded plates and capture any ciliates that you see.  Your goal is to culture these unknown ciliates and characterize them. One of the most important aspect of this lab is student ownership of their project.  It is time to discover! If you are unable to find ciliates in your soil, store the plate and use a sample provided to you by your TA. Use the Keys for Ciliate Classification included in this Module.  These Keys are based on the morphological characteristics of ciliates.

Purpose

The purpose of today’s lab is to dive further into our ciliates. To identify them based on morphological features and hopefully connect them to taxonomy. Taxonomy shows evolution and common ancestors to explain where they came from and how.

Objective

This week in lab we had several objectives. First, we gained a deeper understanding of ancestor trees, what they mean, and why they are important. Next, we performed a soil texture which we will read next week. Lastly, we put our non-flooded plates onto microscopes to try and identify more ciliates.

Hypothesis

I think a lot of people will enjoy today. Gaining more background information of ancestry trees will help students have a deeper understanding and thoughts about ciliates once they understand where they come from. I think students will be excited to see the composition of their soil next week as it is very interesting to see the diversity even when all samples were picked in a somewhat close proximity. I also think students will have a lot more success today finding their ciliates. Since more time has gone by cilliates have had time to come out of their cysts and even replication resulting in more.

Procedures

Determining Soil Texture

1. Remove the sticks and leaves and other debris from your soil sample
2. Add soil to about 4 mL mark in a Falcon tube
3. Add water and mix vigorously
4. Add 1 drop of dispersing agent and re-mix
5. Observe the tube after 30 seconds. The sedimentation of sand particles will have occurred
6. Let the tube sit undisturbed overnight or until next lab
7. Use a ruler to measure the % sand, silt, and clay. This procedure works best by taking a clear picture of the and zooming in on the sedimentation lines
8. How can you determine the % of each type of soil particle?
9. Record soil type in notebook and on the soil data sheet

Data

Determining Soil Texture

Each student performed the protocol listed above. Once it was performed we set the tubes in a holder and will continue the protocol next week in lab. However, I could already start to the the sedimentation lines and I’m very excited to see what type of soil I have.

Ciliate Observation

This week was a lot more successful for me to find ciliates. Last week I was not able to find any but today I was able to find one. There could of been more in my petri dish but I did not have enough time to keep searching. The one ciliate I found was fairly small, I want to get an actual size next week in the. It was oval shaped and had long cilia. Also, it was super fast, spun in circles constantly, and when it did swim it was usually in large circles. I did successfully get a video but am having trouble uploading it onto here. However, I promise it will be presented in the final group presentation in two weeks.

Conclusion

This lab was a lot of fun and a success. I think it is really cool that we get to find out what types of soil we all collected from. I wonder if there will be a correlation to soil type and type of ciliate found? I wonder how many people will have the same soil type? Also, it seemed that everyone was able to find at least one ciliate from their petri dish which was not the case last week.

Next Step

This lab was very productive and made me feel a lot more confident working toward the final presentation. I now have photographic evidence of my ciliates in addition to my lab group members that had several. I’m looking forward to seeing the soil types everyone has and comparing them. I hope my group members have different soil types and hopefully different ciliates. With that information I plan on discussing how soil type impacts the type of ciliate that is found.

11/1/18

Marci Jordan

Bio Lab 1105-31

Pre-Lab and Lecture

Learning Theory:

The purpose of this course is to intertwine the cognitive science of metacognition with the content of ciliate discovery. We must be able to transfer our ability to reflect, discuss, propose, analyze, fail, dig deeper, work together, and so on from this experience to future courses and to situations beyond formal education. Some goals to reflect on include to understand that constructivist learning requires the learner to take responsibility for constructing their knowledge by being actively involved in the process of learning. To move closer to a growth mindset in which you believe that abilities can be developed through dedication and hard work. To understand that creativity powers science and increases with diverse collaborations. To be willing to fail to succeed and take the intellectual risks required for discovery.

Background Information:

The discovery and characterization of soil ciliates is an extremely important area of biological research. The biodiversity of life in the soil is important for a healthy planet.  We depend on soil for the air we breath and the food we eat.  Even so, we are just learning about the complex ecosystems involved in nutrient recycling that are driven by the microbiota of the soil.  Very little is known about the roles of ciliates in this foodweb or the diversity and abundance of ciliates in soils. The study of ciliates is a topic that will allow us to explore the core concepts of biology: evolution, cell structure and function, genetics, biochemical pathways, and interactions in living systems.

Among the least known soil organisms are protists. Soil protists are, however, the most diverse soil eukaryotes. Their functional importance expands well beyond being bacterivorous, including their role as keystone organisms of soil ecosystems and regulators of essential processes of soil fertility such as nutrient cycling and plant growth.New sequencing methods are now available to disentangle the composition and function of the protist community. Our goal is to isolate ciliates using the non-flooded plate method and ultimately culture our isolates for DNA sequencing.

Bioskills:

Soil pH Protocol

An essential part of understanding the ciliates we collect is having a detailed understanding of their environment – the soil. While there are many biotic and abiotic factors that influence the soil microbiome, the biotic factors in these environments can be highly variable and sometimes difficult to precisely quantify. Thus, our initial soil assessment will evaluate several abiotic factors that will be helpful in evaluating the genomic data produced in this lab.

Soil acidity

Acidity is measured on the pH scale, which ranks the concentration of H+ ions on a scale from 1 (very acidic) to 14 (very basic). A pH below 7 is considered acidic and a pH above 7 is considered basic.  pH can be determined by using a pH meter or pH paper.

In this lab we will use pH paper in the range of 5.0-9.0.

1. Place a small amount of soil in a glass vial and add deionized water. Mix thoroughly for 3-5 minutes and then let the soil settle to the bottom.
2. Remove about 1 ml of liquid from the top of the tube and transfer it to a clean Petri dish lid.
3. In order to test the pH of your soil water, remove a small strip of pH paper and place it in the drop of soil water. Add the clear soil water so that you submerge the strip.
4. Wait 1 minute and record the result after comparing the color.
5. Record the pH in your notebook.

Non-flooded Plate

A non-flooded plate is a simple technique used to observe and maintain microscopic life in a collected soil sample. Since ciliates congregate along the water-air interface, using the non-flooded plate allows life to be easily maintained and observed.

As the name for this technique indicates, non-flooded plates involve adding enough water to soil in a petri dish to saturate but not flood the plate! As water evaporates from the plate over time, you may need to add additional water as you see the soil drying.

Protocol

1. Put 10-50 g of fresh or air-dried soil in a petri dish. You just need enough to cover the bottom of the plate on one side.
2. Saturate but do not flood the sample with distilled water.  Add water until about 5 ml will drain off when the petri dish is tilted.  You may need to give the soil time to be saturated.
3. Observe your soil using the dissecting microscope and record your observations.
4. In order to observe the abundance and diversity and capture a ciliate from the soil, use your micropipette and remove 100 µl or so of liquid sample.  Transfer this to a concavity slide, watch glass, or small glass petri dish in order to observe and further isolate.  Record the number of ciliates observed.
5. Isolation may be done by serial dilution (if there is more than one type of ciliate in your sample).
6. Once you have captured a ciliate, transfer it to 500 µl of media in a clean 24 well plate.

**You may need to experiment with types of culture media.

7. Put the lid on the non-flooded plate to decrease evaporation, but still allow for airflow and gas exchange.
8. Continue to check your plates after flooding as often as possible up to 30 days.

Note: There is a predictable succession of ciliates in the plate over time.  Your challenge is to observe and describe the abundance and diversity of ciliates in your sample and isolate and culture a single type of ciliate.

The process on air dried soil (in which the ciliates enter into their encysted form)  may isolate more individuals and species than a fresh sample, probably due to the ability of ciliates to tolerate this change to a greater degree than other organisms (microbiostasis).

Purpose

This purpose of lab this week is to begin part three of CILI-CURE, soil ciliate discovery. The short term purpose of this is to gain observations and data to present to the class in three weeks. Within each lab group, members will identify ciliates and discuss their finding through presentation. Long term the purpose is to continue the soil ciliates into next semester. Next semester in CILI-CURE students will use the same samples but will work with DNA sequencing of the PCR product for barcoding.

Objective

The research objective of this weeks lab meeting is to determine the soil ciliate biodiversity within soil samples collected earlier this semester. Students measured soil characteristics and ciliate discovery to later present in group scientific presentation. The plan for lab this week was to calculate the & water content, determine pH of soil extract, and observe the non-flooded plate for ciliates.

Hypothesis

I think that students will be excited about part three of CILI-CURE. It is nice to take a break from the research paper and be back in the lab. Discovering living organisms on a microscope is much more fun than searching for primary sources on a computer.

Procedures

Percent Water Content

1. Retrieve your soil samples
2. Weigh the samples and calculate the mass of water that has evaporated from the soil
3. Convert this to a percent

Wet soil-dry soil/wet soil x 100= % water content

pH of Soil Extract

1. Place about 1.5mL of liquid from soil sample and insert into a microfuge tube
2. Spin the tube in centrifuge fir 1 minute to pellet the soil
3. Remove a small strip of pH paper and place it into the liquid, not touching the soil.
4. Wait 1 minute and record the results after comparing the color.

Non-Flooded Plate Observation

1. Push soil to one side of petri dish and let water collect on the opposite side
2. Place petri dish under dissecting scope to observe any bigger ciliates
3. Take a micropipette and attempt to collect some ciliates and transfer onto a microscope slide
4. Observe cells under a compound microscope and try to collect photos or videos

Data

Percent Water Content

((17.01-13.8)/17.01) X 100 = 18.87

18.9% Water Content

pH of Soil Extract

pH: 6.0

Non-Flooded Plate Observation

So both my partner Lou and I had a unique situation. Our petri dishes did not get flooded 24 hours before like everyone else’s. Therefore, our ciliates still remained in their cysts and nothing could be seen besides rocks and dirt. However, our third partner Riley did have one ciliate and one worm found from her sample.

Conclusion

Overall this weeks lab was a great introduction to our third and final portion of CILI-CURE for this semester. It was nice to get our soil samples back and find information about them. It was unique to find the water content that was already in the soil when I collected it. Also it was interesting to see that most people had the same pH level but others differed. I’m excited for the weeks to come to see what my group will produce.

Next Step

After this lab I have a lot of thoughts for the next coming weeks. First, I a little worried that not having my dish already flooded set me back. Two out of three group members could not record observations as our ciliates were still in cysts. I hope next week there is time to catch up as I want as much material to present as possible since our last group presentation did not go well. I wonder what characteristics we will be observing next week and we still have two more lab meeting before presentations. Overall, I’m excited to see what everyone discoveries but I feel like I have some catching up to do.

October 25, 2018

Marci Jordan

Bio-lab 1105-31

Lecture and Pre-lab; What goes on in each section and responses

Final Report

The scientific literature is written in a standard AIMRaD format of an Abstract, Introduction, Materials and Methods, Results and Discussion followed by Acknowledgements and Citations. The following information is a brief outline of each section in the order that it may be written.

1. Experimental Design: As you plan your experiment, read the literature and record your references and make notes of relevant background information.

The Introduction gives the background of your experiment and is often formatted in the way of an argument.  You are giving references to prior knowledge in order to build to your question and hypothesis. The introduction should begin by giving the reader all the information needed to understand the article and why it is important and novel. This description should be as brief as possible, while still giving all pertinent information.

2. Running the experiment: As you perform the experiment, keep detailed notes on your protocols. Record the materials that were used, the concentrations, time, and any other important information required in order to repeat the procedure.

The Materials and Methods section can be the easiest to write.  Write in a concise manner explaining everything that you did.  The materials and methods should include a very detailed account of all the procedures done and an explanation of why they were chosen. When finished, the reader should be able to replicate the experiments simply from the information given. No results or analyses should be included in this section.  Include the chemical information about your media and your treatment. Write in the past tense giving details of the organism, chemicals, media, volumes, concentrations, etc.

3. Analysis: Analyze your data by comparing the results of the Control samples and the Treatment samples. Common analyses include mean, median, mode, standard deviation, and student t-test.  Visualize your results using graphs and charts.

The Results sections should parallel your Methods. Each step in the procedure that has an outcome should be mentioned in the text of results. For your Lab Report, begin by analyzing your data and making your figures.  Remember for your report you may be including all the results for the class.  The final figures and tables should summarize the results in a cohesive manner. The narrative of the Results should point the reader to the Figures and describe the figures in words.

Statistical analysis of your data is essential and should be clearly explained and shown in all Tables and Figures. When comparing mean values among treatments, use of standard error (SE) is correct. The size of the SE is a function of the “n” so SE values are better than standard deviation (SD), because the SD does not depend on the “n”.

4. Discussion and Conclusion: Draw conclusions from your results. Return to the literature to look for similar experiments. Compare your results to what has been previously reported.  Propose future experiments and note any difficulties with your experimental design.

The Discussion should consider the significance of the results in regards to the problem that is at hand and should also consider the significance in comparison to other research. This section will offer a synthesis of the author(s)’ conclusions based on their experimental data and potentially what important questions should be addressed next.

5. Formatting: Check your paper for the correct scientific style and APA format, especially citations and references. Be sure not to plagiarize.

6. Abstract: After your paper is written, write a clear and concise Abstract.

The last thing to write is the Abstract. An abstract is a single paragraph composed of no more than 250 words. It briefly outlines the entire article, excluding details of the methods and should not contain reference citations.  It contains some background, the purpose, information about the general methods used, the most important results, and a conclusion or recommendation.

7. Title Page: The title page should include: the title of the article, your name and group members, the class and section, and semester. The title should provide as much relevant information as possible, but be concise.

Plagiarism is a form of Academic Dishonesty, and we take it very seriously. Note that any behavior that we feel constitutes Academic Dishonesty will be handled as recommended by the University.  Please see the course syllabus for information on Baylor’s Academic Dishonesty policies.

Examples of plagiarism:

1. Directly quoting text without attributing credit to the author
2. Paraphrasing text without attributing credit to the author
3. Copying a figure or table without attributing credit to the author
4. Using information in a figure or table (in your own revised version) without attributing credit to the author
5. General note regarding in-text citations: be sure the reader understands what the citation covers.  Also be sure everything you refer to in the body of the paper is listed in literature cited, and be sure there is nothing in literature cited which is not included in the body of the paper.
6. This assignment will use the Turnitin app which compares your writing with a very large database of writing samples, web pages, and journal articles.

Putting it Together

The Abstract

The instructions for writing an Abstract follows these general points:

• Some background information (Introduction)
• The purpose statement (Introduction)
• Some information (brief) about the Methods
• Overall and most important Results
• A statement about the Conclusions

For your report there is a 250 word limit.

Discussion and Conclusion

The Discussion should not be repetitive of the Introduction, but it should clearly connect with the issues and questions that you raised.    Here is a list of elements that are often found in the Discussion section.  Each sub section may be 1 or 2 paragraphs and may introduce new references.  The first 2 summarize the Introduction and Results.  The last 3 form an argument to explain the results and their importance.

1. The purpose of the study.
2. A summary of the findings in order of importance and whether they agree with the hypothesis and/or the literature.
3. Explanations for the findings with references to support your claims.
4. Implications and context in the broader field.

1. Recommendations or plans for future studies.

Citations and References

Journals vary in the style of citations they use.  For this report we will use the numerical citation system as seen in the following paragraph from the article  “EFFECT OF LIGHT ON TOXICITY OF NANOSILVER TO TETRAHYMENA PYRIFORMIS”.(On module 10) Check out the formats on Zotero and choose the one that matches this style.

The references are listed in numerical order according to the first time the reference is used and they are listed at the end of the report under a section titled REFERENCES.  Each Reference is listed in a standard format with required parts. The references should be up-to-date and essential in providing information required to understand the research.  For a research report, it is not necessary to write a review of the literature.  It is important to point the reader to the background information and relevant studies and of course- no plagiarism!

Besides numerical citations, many journals use (Author, Date) as seen below. These types of citations are also listed at the end of the article under REFERENCES, but this time in alphabetical order. Many text now also include an active link and a Digital Object Identifier (DOI) so the reader can link out to the references.

The overall point of citations and references is so the reader can know whose ideas are being described and where to go to find the evidence for the idea. It is important to be consistent in style. For this paper, use the APA format for citations and references. Alphabetize your references and number them.  Insert the number in the report whenever you refer to that reference. Use Zotero to make this a relatively easy task.

Purpose

The purpose of lab this week was to present our result figures with our lab group to the class. This gave us the chance to have incomplete or incorrect observations pointed out to us. The purpose was to also notice what other groups did differently and how it compared to ours.

Conclusion; Responses to results

In class we only went over results so I reflected a lot on that section. I have a lot to change! For cell counts a lot of group put their n value as 3 as each group used three drops however, you used everyone’s results so we must multiply 3 by the number of people. Next, each group had varying numbers for their p value while they should have the same. I now must go back and check that I used the correct values for each step to get my p values. With the p value I need to include that if it is less than 0.05 it is significant and rejects the null hypothesis. I also need to create a graph for vacuole formation and 15 minutes and combine it to the graph for 5 minutes.

Next Step

With all this new information I have a lot of checking and changes to make. I will use these two days before the rough draft is due to make it as perfect as possible. Therefore, I have fewer changes to make before the final draft is due. I will use the descriptive explanations of each section to make sure every aspect of my research paper is up to par.

10/18/18

Marci Jordan

Bio Lab 1105-31

Pre-Lab and Lecture

Nature of Science: Uncertainty of Science

New students of science work hard to find the correct answer. However, experimental questions only result in answers, not correct or incorrect answers. By now in this lab we have learned that science is not about being correct. We must use this realization to motivate us to think about experimental design and more scientific inquiry. Scientific uncertainty is inherent in the scientific process. Scientific certainty is only achieved when multiple trials of an experiment or a project give repeatable results.  Even so, there is also inherent uncertainty when measuring a variable trait in a population or counting survivors in a culture, for example. Biological systems have an uncertainty associated with the range of natural variability and interactions. A scientist understands that at any time, someone else may provide evidence that questions their findings or refines what they have done. This is how scientist move forward. Scientist introduce authentic science to the public so they can experience uncertainty and reflect on this important aspect of science

The Writing Process

The true story of research is told in the results. It is easiest to begin by first making the result figures. The easiest order to write a research paper is listed below:

Results: Figures, captions, and narrative.

Methods and Materials:  To explain and parallel each result (The first draft of the Materials and Methods can be written before the Results are analyzed, but the M&M are usually edited after all the Results are completed so that the 2 sections are written in the same order and are easy to follow.)

Introduction: To frame the story and give needed background (Citations required)

Discussion: To explain the results and discuss possible errors or future experiments (If you compare to other studies, more citations are required.)

Abstract and Title: Summary of the paper

References (If they are in Zotero, this will be an easy step.)

The results section of a paper should be parallel to the methods section. Not everything is copied sub-section for sub-section as meathods describe the preparation and results report what was measured. However they should be very similar. When writing the results section there are two main difficulties that arise. First, a results section that contains only tables and figures. This tells the reader nothing about why the results are presented in this way or this particular order, or what the reader should notice about the results. Second, results section that contains explanations of the significance of the results. This belongs in the discussion section.

Somethings that belong in the results section include: refer to your tables and figures and introduce their content, highlight the most important result(s), summarize and compare results, write with certainty about your results, and identify unexpected results. Make sure to avoid simply repeating the content of the figures and tables, generalize or interpret your results, and just list the results. Figures should be supported by text that highlights the important findings, helps the reader find the figure, and comments briefly on the results.  The comments are normally used for comparison or a clarification.  The full discussion is saved for later.

It is very important to know when to use what kind of figure. Use a table when it involves: recording raw data, explaining calculated data, showing the actual values, to make comparisons in many directions, and often a way to summarize the text. The most common figures are: pie charts, used for highlighting proportions; column or bar charts, used for comparing different categories; and line charts, used for displaying a sequence of variables and illustrating trends. It is very important to be careful when formatting figures. Make sure to use the same colors, types of scales, label axes, and write descriptive titles. To make a figure easy to interpret it must be clean and uncluttered.

Excel Help

A video a little over 2 minutes was given to us in the pre-lab. This video is for excel for beginners. Seeing how I have worked with excel, I did not watch the video but it is nice that it is there for reference. We had to make a practice figure in excel and upload it as part of our pre-lab. On the following page was another link to a page which helped with basic Microsoft functions for MAC computers, I personally have a Dell.

Purpose

The purpose of this week in lab is to further extend our research paper. With methods already done, now is results, and then everything else is due by next Friday. The purpose in lab is to create figures under supervision to make sure we have all the correct bells and whistles to make our figures complete.

Objective

The objective is to successfully make figures up to the correct standards. To make sure they have a good color scheme and all the information needed at a basic level

Hypothesis

My hypothesis is that student will have a lot of fun in this lab. Although it is a lot of work it is fun and easy to make colorful grapgh which depict the result in a way to see what they mean. Also, since we are breaking up the work load there will be more time to put into each figure.

Procedures

1. Get into lab groups
2. Each person will create a figure for their assay
3. A person must also make a figure for the cell counts and optical density
4. Follow the outline/rules on how to make a proper figure.

Data

On the link below is all the figures I made. I could not find a way to copy and paste them but hey theres the link. On the link it includes all the figures as my lab group broke up the work. Since my assay was vacuole formation I completed that gragh and also did the cell count to help my team. Although I cant copy the figures (again they are on the link) I will write the description that are under each figure.

Cell Counts

Figure 1: Mean cells counted in the control and treatment Tetrahymena environments. (n=3; p=1.27E-07). The treatment includes microplastics (polypropylene) to see if it would affect the number of cells.

Vacuole Formation

Figure  5: Mean of vacuoles formed in the control and treatment Tetrahymena environments. (n=10; p=0.07). The treatment includes microplastics (polypropylene) to see if it would affect amount of vacuoles formed.

Reserch Project Figures-18xgg8n

Conclusion

In conclusion, this lab was a sucess. My lab group completed all our figures and got them checked by Felicia. They include all the criteria and are ready to be included in the research paper. I really like how with our figures it is easy to see what is significant and what the results mean.

Next Step

Now that these figures are done there is a lot in store for them. They will be inserted in the appropriate sub-section in the results section of my research paper. They should be so good that someone can look only at the figures and know what my research was on. Once the figures are added the methods and results sections will be complete and then I will add the other sections of AIMRAD to complete my paper for the rough draft which is due Friday. I definitely need to find sometime this week to dedicate to writing the other sections.

10-12-18

Marci Jordan

Bio Lab 1105-31

Pre-Lab and Lecture

Learning Theory: Fail to Succeed

Dr. Adair gave us a short podcast to listen to called “Why Science Needs Failure to Succeed” by Science Fridays. This podcast was very interesting as the host and two guest, Stuart Firestein and Helen Snodgrass made comments on the subjects. Some messages I liked and took away are as listed. Success in science gets all the attention. People are awarded the noble peace prize for their discoveries but without failure those discoveries never would of been found. To be a scientist you must have the stomach for failure but also love the taste of failure. Failures lead scientists down the correct path as it takes them to new places in which they find new discoveries. An example of this is when the G-Protein receptor was discovered. The experiment kept failing because of the washing techniques of the supplies. There was aluminum fluoride in the washing solution which disrupted the experiment but lead to the discovery that trace metals are activators of enzymes. In school, they only teach about the discoveries. But those discoveries that everyone learns about all came from failures. In Helen Snodgrass’s classroom she have “Failure is not an option, it is a requirement” posted on wall. I really enjoyed this podcast because it not only explained that its okay to fail in science, but also emphasized that failure is necessary to be successful.

Bioskills:

Data Analysis

Following tutorials/data is not using our class lab data. In statistical analysis we will use four tools on the Excel Toolpak. These tools include Descriptive Statistics, Histogram, The F-Test Two-Sample for Variances analysis, and The Two-Sample t-Test analysis tool. Each of these tools have different purposes so I will list each purpose of the four below. The Descriptive Statistics analysis tool generates a report of univariate statistics for data in the input range, providing information about the central tendency and variability of your data. The Histogram analysis tool calculates individual and cumulative frequencies for a cell range of data and data bins. This tool generates data for the number of occurrences of a value in a data set. We can use it to generate a graph to test for. The F-Test Two-Sample for Variances analysis tool performs a two-sample F-test to compare two population variances. This result will be used to choose the type of t-Test. Lastly, The Two-Sample t-Test analysis toolstest for equality of the population means that underlie each sample. The three tools employ different assumptions: that the population variances are equal, that the population variances are not equal, and that the two samples represent before-treatment and after-treatment observations on the same subjects.

Data analysis is simply the conversion of raw data into meaningful information. Remember that statistical analysis indicates correlation, not causality. The experimental design can contribute to the ability to draw inferences from an experiment.  For example, a controlled experiment that has only 1 variable, is easier to interpret than a comparison of cell counts from a variety of natural environments. Below are nine definitions of common used terms in data analysis. A proportion is a fraction – for example: out of the 100 organisms seen in the soil, there were 3 ciliates (3/100). A percentage is another way to express a proportion by multiplying it by 100% (the proportion above would be .03 x 100% = 3%). The mean is the average of a set of data, and can be obtained by dividing the sum of the set by the number of data points. The mean is sensitive to extreme values – when there are outliers in a set of data, the mean is less helpful as an analysis tool. The median is the middle of a set of data, and can be found by arranging the data in numerical order and finding the middle-most data point. The median is less sensitive to extreme values, and can be useful despite the presence of outliers. The range of a data set is the difference between the extreme measurements (smallest & largest) of the sample. The Standard Deviation is a measure that is used to quantify the amount of variation or dispersion of a set of data values. Basically- how spread out are the values? Variance is the expectation of the squared deviation of a random variable from its mean. Informally, it measures how far a set of (random) numbers are spread out from their average value. Charts and graphs are used in data analysis to display trends, relationships, and comparisons. Different types of graphs are used for displaying different types of information. For instance, a line graph displays trends over time, a bar chart allows comparisons between categories of data, and a pie chart shows proportional share. All charts and graphs should be clearly labeled and should enhance the viewer’s understanding of the topic.  Tables are an excellent alternative to charts and graphs. While it is easier to detect patterns in graphs, tables allow you to display much larger quantities of data.

Statistical Analysis Overview and Descriptive Analysis

A screen capture performed by Dr. Adair showed the steps to get the statistical analysis data. First, Open the excel file generated in class. The click the data table and on the right side click on “Data Analysis”. On the box that pops up by scrolling down you can see the four tools we will be using in lab which I talked about individually above.

Another screen capture performed by Dr. Adair showed the steps to get the descriptive analysis data. She performed this using the cell count data from class. First she opened up a new sheet on excel(on the bottom). Next, she copied and pasted different cell counts from different sections. Then, click the data tab, click on the data analysis button, and select descriptive statsitics. Lastly, she selected the input range and output range. Also, be sure to check the “summary statistic” box as it will give you all the information you need. Once the new data is generated you can compare the results side by side.

The Null Hypothesis, P-Values, and Statistical Testing

Nothing in science is proven until it is disproven. In other words, we cannot prove that a certain chemical causes Tetrahymena to grow or not, because this proof requires that we have thought of, successfully tested, and have the results of every possible observations concerning this hypothesis. Instead, we can state the negative and say the chemical will have no effect on the growth of Tetrahymena. If we detect that there is a difference in growth then we can reject the null hypothesis. This falsification of the null hypothesis supports the idea that there is an effect caused by the chemical. If this is the result we have reason to investigate the effect further. If we do not detect a difference in growth, than we accept the null hypothesis and conclude there was no effect.  We did not prove there was no effect; we collected evidence that supported our null hypothesis.

There are many statistical tests and rules concerning hypothesis testing. In most cases, these tests ask in a mathematical manner how probable are the data if the null hypothesis is true. The probability is reported as a P-value. The smaller the P-value, the stronger the statistical evidence against the null hypothesis. A common critical value that a test statistic must exceed in order to accept the null hypothesis is 0.05.

So, if our null hypothesis predicts that the difference between the control cell counts and the treatment cell counts is 0 (there is no effect caused by our treatment) and we compare the values using a statistical test (such as the Students T-test) and calculate a P-value of 0.001, then we would reject the null hypothesis and support the idea that the treatment did have an effect. It does not prove that the null hypothesis is false; it just indicates that it is unlikely to be true.

The T-Test

In our experiment, the effect of a substance was measured through measuring cell counts. Our null hypothesis states that there was no difference between the control and the treatment at a certain time period. In order to determine if the mean averages are the same or different, you need to use statistical tests. One of the most common ways to test the difference between 2 means is called the Students T-test. This test can be found in the list of statistical tests in Excel (T.TEST) and is listed in the Data Analysis tools along with the other tools we will be using.

This test has some assumptions which can themselves be tested using a histogram and an F test, also found in the Data Analysis tools.

1. The individual observations are independent of each other. (The occurrence of one does not affect the probability of occurrence of other)- TRUE
2. The distribution of observations are continuous (not categorical) TRUE
3. The observations are normally distributed. Make a Histogram to test
4. And the variances of the control sample and treatment samples have to be considered. Use the F test.

Once you have determined if you have equal or unequal variances, run the appropriate T-test.

Purpose

The purpose of this lab was to run statistics of our results. Statistics involves estimations, inferences, and study designs which help us see patterns. Students will not only get to assess statistics but actually compare their results and find what they mean. The statistics might not show anything exciting but it puts it in a format to compare which is the purpose.

Objective

The objective this week was for students to perform a series of statistic processes. These statistic processes will be done with the combination of everyone’s data from the section from last week. Students will properly perform a descriptive statistics, histograms, F test, and a T test. Each of these tools will be used twice; once for the cell counts (control and treatment) and twice for the assay which they performed (either cell speed, directional change, or vacuole formation).

Hypothesis

I think that students will be very intrigued during this lab. It was a lot of fun to finally perform the experiment and get data; but now we can compare and see what the results mean. However, I think that students who did not complete the pre-lab will have difficulty as the pre-lab allowed students to use these tools before class.

Procedures

Descriptive Statistics

1. Perform descriptive statistics of cell counts
2. Repeat; once for the control, second for treatment
3. Perform descriptive statistics for vacuole formation
4. Repeat; once for the control, second for treatment

Histogram

1. First, make a column of data and a column of bin numbers
2. Bin numbers are the X point values you want on your graph
3. Create four histograms (cell count control, cell count treatment, vacuole control, and vacuole treatment)

F-Test

1. Use a F-Test to determine the variance
2. First, run a F-Test for cell counts
3. Second, run a F-Test for vacuole formations
4. Then look at the values of variance to determine which T-test to use

T-Test

1. Use the appropriate T-test based on variance
2. First, run a T-test for cell counts
3. Second, run a T-test for vacuole formations

Once all these tools have been utilized and you have all the tables and graphs, transfer the final answers onto a word document.

Data

Cell Count Descriptive Statistics:

Vacuole Formation Descriptive Statistics (5 mins)

Histograms for Cell Counts

Histograms for Vacuole Formations

F-Tests for Cell Counts

F-Tests for Vacuole Formation

T- Tests for Cell Counts

T- Tests for Vacuole Formation

Conclusion

In conclusion this lab was a lot of fun. It was very convenient using excel to do all the work and getting the statistics ran right away. From the results my conclusion is that Tetrahymena are affected by polypropylene. I say this because seen in the results above the data is higher for the treatment. For cell counts, there was much more cells in the treatment then there was in the control. In the vacuole formation assay there were more vacuoles formed in the treatment than in control. This was not as huge of an increase as the cell counts were but it was much more. I’m not sure what will come next but from this weeks lab I can confidently say that polypropylene makes cells reproduce more and form more vacuoles.

Next Step

With this new information I am looking forward to next weeks lab, whats next? Now that the research has begun I am thoroughly enjoying it. I have this data and I can see that the polypropylene affected the Tetrahymena. I will take these statistics and apply them to other research that has been done. Can my results possibly support someone else’s findings? Are my results popular with other groups in my section? I know my results are that it changed the cell count and vacuole formation but is it enough to reject the null hypothesis? My next step is to find answers to these questions as I continue research and form my research paper.

10/4/2018

Marci Jordan

Bio Lab 1105-31

Pre-Lab

Learning Theory: The Importance of Stupidity

We were given an article in the pre-lab which I really enjoyed. First the question was presented; do you like feeling smart or challenged? It then went on to discuss how science is based on failure. You must except mistakes and being wrong to make new discoveries. Research is hard as no one knows the answers. The purpose of lab is to learn the process of science and experiment the reality of failed experiments and challenges. “Failure to succeed” is acceptable in science. This made me feel better as I have definitely not been getting everything right. Having this article to reassure me that everything will be okay was a nice weight taken off my shoulders.

Bioskills

Serological Pipette:

We have spent weeks working with pipettes. Learning how to properly change the volume, switch tips, and what each part does. This week we were introduced to a new kind of pipette. Serological pipettes are more recent and more accurate. They can in several different volumes and work different ways. Serological pipettes are used for volumes greater than 1mL. One pipette carrying 1000ml three separate times would equal using one serological pipette once.

Practicing Calculations and Mean/Standard Diviation:

For practicing calculations we were given some practice problems and a reference link. This is so if we ever forget how to do any calculations. Calculations included finding cell counts and serial dilutions. Next, we were given a mock experiment with four separate students who performed 3 trials of cell counts. With 12 different counts we were then asked to find the mean and standard deviation using excel. In the excel data analysis tab there are formulas which will do all the work for you. I plugged in the 12 numbers and it gave me the information I was looking for in seconds.

Protocols and The Format of a Scientific Paper:

Protocols are methods that are used to perform an experiment. Similar to a recipe when cooking, protocols offer step by step detailed instructions. It gives you a plan to follow, specific amounts, and times. It’s always a good idea to have a protocol ready before you start an experiment. Writing down the protocol prevents issues or needed supplies later in the experiment. When formatting a paper, papers are not always written beginning to end. It is important that students look and follow “AIMRAD” to have the correct format. A research paper is very different then a English paper. Students need to start finding articles similar to the research and save them in ZOTERO. Articles will be used in the research paper’s introduction and discussion.

Methods and Materials (Style and Content):

The methods section should derive from your lab notebook but it should not read like a notebook. Subheadings are used to organize the methods. The methods section should make it easy for someone else to follow the experimental design and should be identical or similar subheadings as found in the results. It is up to the writer about what kind of voice they want to write in. It is recommended to use a past tense passive voice instead of an active voice. An example of an active voice is “The Scientist examines strains of bacteria” while a passive voice looks like “Strains of bacteria are examined by the Scientist”. To help develop our own methods section we were given a couple links to observe how it is done. Attached below is my Methods and Materials portion I typed for the pre-lab to be included in my final research paper. It is not complete by any means but it is what I have created in this point of the experiment.

Materials and Methods

General Experimental Design

In CILI-CURE 2018 Tetrahymena soil cells will be tested against the microplastics in baling twine by the students. Different assays like vacuole formation, direction change, and swim speed that the Tetrahymena perform will be observed as data.

Bailing Twine

Baling twine is the polypropylene (PP) plastic that wraps around bales of hay to tie it together. It is a problem as much of this twine gets disregarded on the ground and makes way into the soil either by soil tilling or environmental factors like weather. These are long pieces of twine and once they are in the soil, it is not easy to pull back up. Devastating images of baling twine polluting Osprey’s nest can be seen by the twine hanging over the sides of the nest swinging the wind. Osprey get their feet caught in this twine and die frequently. Cattle and horses eat and digest this twine as it is in the grass. This effects their digestive tracks as it is chemicals that they should not eat. Not a lot of research has been done on microplastic pollution’s affect on the soil. Like the plastic in the ocean has macroscope effect, plastic is the soil might too. Therefore, testing Tetrahymena with the microplastics found in baling twine will give the students the results of how plastic affects the soil.

Polypropylene

PP is a thermoplastic material resulting from the polymerization of the propylene by catalysts. PP is the plastic which makes up baling twine. It is a very resistant material and normally tough and flexible. PP twine can have knotting strength range from 130lbs to 700lbs, sometimes even up to 1000kg without breaking any knots. It is low in weight and easy to handle so farmer love it. It comes in different colors for identification, but the color also provides UV protection, so it will not break down in the sunlight. Lastly, PP is very inexpensive which is another reason why farmers love it so much.

Polypropylene Microplastic Production

To get the microplastics from PP a series of steps were performed by the students. First, shed or cut PP twine into small pieces with scissors. Section 31 will have the color white twine. Next, measure 0.5g of PP twine into a sterile glass beaker. Then, add 50mL of sterile proteose-peptone-tryptone (PPT) media into the beaker (this is the Tetrahymena medium). To act as heat from the sun; boil and stir the mixture for the entire lab. Afterthat autoclave the “twine juice” to kill any bacteria in the mixture and store for next week. After a week has gone by filter mixture using 5µm filter paper into a sterile 50mL tube.

Tetahymena Medium

5g of proteose-peptone, 5g of tryptone, 0.2g of K2HPO4, and 1L of distilled water combined makes up the Tetrahymena Medium (or PPT seen above). Adjust the pH to 7.2 before autoclaving to remove any bacteria in the solution. Dispense 5mL of medium per test tube.

Lecture

Fail to Succeed:

To struggle and fail are essential parts of science. If you plan your experiment carefully, then both positive and negative results will be meaningful. If your ciliates die for an unknown reason use that opportunity to hypothesize why that happened and potentially improve the protocol. There is no difference between the control and treatment you can hypothesize why and potentially improve the protocol. Part of experimental design is very “cookbook”, but collaboration, observation, record keeping, and creativity are needed skills.

Experimentation:

Experiments provide data that determine if a hypothesis should be accepted or rejected. Our treatment (independant) variable is how the PP from the twine affect the Tetrahymena’s behavior. Our response (dependant) variable is how they respond and that are the results. Our contol is the Tetrahymena alone in the media which is their food and no new factors are involved.

What Happened to Jars:

Each jar of twince was filtered using 5 µm filter paper. PPT media was added to the filtered solutions so that all the solutions had the same concentration of 0.5g/50ml, or 1%. The solutions were autoclaved to kill any bacteria or other microbes. On Tuesday at 5p.m., 5 ml of a 6.1×10^4 cells/ml Tetrahymena culture was added to 45ml of media or treatment (1:10 dilution).

Spectophotometers:

Optical density (OD) is the degree to which a refractive medium retards transmitted rays of light. As see below the spectophotometers were used in lab t see the optical density in just the PPT(media), PPT + Twine juice (PP), PPT + Tetrahymena, and PPT + Twine Juice + Tetrahymena.

Purpose:

This purpose of this weeks lab is to first master more calculations, writing skills, and obervations. Now that we have a experiment designed, it is time to test and start getting some results.

Objective:

The objective of this lab is to first start thinking of or completely write the methods portions of the research paper. We have been given enough information to start that section. Next, fine tune using tools such as microscopes and pipette. Successfully make cell counts. Lastly, perform the tests for the CILI-CURE 2018 research project!

Hypothesis:

I predict that students will be very excited and ready to start the process. I think everyone will be successful at the cell counts and feel reassured they get to do the assay that they are familiar with. My hypothesis for all the assays are that the Tetrahymena will be negativity impacted by the twine juice. It will make them reproduce slower and not form as many vacuole.

Procedures:

Spectrophotometers

1. Measure the Optical Density (OD) at wavelength 600nm of each solution
2. Record the following:

• PPT (clean media)
• PPT + “Twine Juice” Treated Media (TJ)
• PPT + Tetrahymena (TH)
• PPT + “Twine Juice” Treated Media(TJ) + Tetrahymena(TH)

Cell Counts

1. Count three drops using 2 µl of Tetrahymena control and 1 µl of iodine
2. Take a picture and count how many cells are present with a cell count data table
3. Count three drops using 2 µl of Tetrahymena treated solution and 1 µl of iodine
4. Take a picture and count how many cells are present with a cell count data table
5. Observe the differences between the control and treated samples

Vacuole Formation

Perform this twince; once with the control and once with the treatment Tetrahymena

1. Place 10 µl of Tetrahymena culture on a flat slide
2. Add 2 µl of India ink to the drop of cells and pipette up and down to mix the cells and the ink
3. Place 2 µl of methyl cellulose in the center of the drop
4. Place a cover slide on the drop and focus the slide on 100X-400X magnification on the compound microscope and start your stop watch
5. After 5 minutes, scan the slide for a cell (randomly pick the cells) and count the number of stained vacuoles in the cell
6. After 15 minutes, count the number of vacuoles for 10 more cells
7. Repeat the procedure at 30 minutes

Data:

Spectrophotometers

Cell Counts

Vacuole Formation

***What happened:

The information of vacuole formation from the control Tetrahymena to the treatment trial are different because my partner was confused about the data. While I followed the table and inputted how many vacuoles I saw in each cell. My partner counted the total number of vacuoles in 10 cells. That is why here numbers and format are very different from mine. Luckily two other tables performed this experiment and the lab can use their data for reliable results.

Conclusion:

In conclusion, all the assays were performed successfully with minor result differentiation. In the spectrophotometers we learned that the twine juice is what absorbs a lot of light. With just the media and/or Tterahymena the light can mostly all pass through. In cell counts we were surprised to find that there were more cells in the Tetrahymena treated with the twine juice than the control Tetrahymena in the media. With the vacuole formation it is diffucult to compare the data but when I add up my data from each time slot the tetrahyhmena are forming less vacuoles than the treated Tetrahymena. Overall, twine juice is absorbing the most light, there are more cells in the treated Tetrahymena, and more vacuoles are formed in the treated Tetrahymena.

Next Step:

Once we got this data I was extremely surprised from the results. All the tests I performed the twine juice caused it to have higher numbers then the control. I was shocked that in the presents of the microplastic pollution of PP Tetrahymena reproduced more and had faster vacuole formation. Is this due to a mutation? Has the pp caused mutations in the cells DNA? What is causing the increase in cell behavior? My next step is to do research to try and find an explanation for these results.

9/27/2018

Marci Jordan

Bio Lab 1105 31

Pre-Lab

Learning Theory

We already know the difference between a fixed verses a growth mindset. But in this pre-lab we focused on the metacognitive knowledge versus regulations. Metacognitive knowledge is  “cognition about cognition”, “becoming “aware of one’s awareness” and higher-order thinking skills. While Metacognitive regulation refers to the ability to regulate self learning. Self regulated learners are able to understand what a task involves, identify personal strengths and weaknesses related to the task, create a plan for completing the task, monitor how well the plan is working, and evaluate and adjust the plan as needed.

The Process of a Scientific Paper

With our own research paper report coming up it hard to make time for it. Everyone has hard classes and tests coming up but making time to research, read, and write is critical. In the pre-lab it stated that the best method is to schedule a time NOW to research, read, and write. Once articles related to the project are found, save them in Zotero (these articles will be used in the introduction and discussion). I wonder when I should start these steps in preparation of the paper? How long does it take to find articles, read, and write the paper? Will I use this system 4 years from now to write my research papers?

Bioskills: Calculations

In the pre-lab we discussed four different ways/types to calculate concentrations. These four ways include percent solutions, molar solutions, C1V1=C2V2 or ratio solutions, and lastly stock solutions/dilutions. In the pre-lab it went into very detailed descriptions and steps but since I had previous knowledge before this lab about how to calculate it all I’m just going to briefly summarize it. For percent totals, to find the concentration simple divide the percent(or part) over the total of the entire solution. Molar solutions are made by dissolving 1 mole of a substance into the solvent (usually water)and diluting to 1 liter. Molarity tells us the number of moles of a solute in one liter of solution. Two pieces of information are required to find molar concentration; moles of solute present in solution and the volume of solution in liters containing the solute. To calculate M= Moles of solute/volume of solution in liters. C1V1=C2V2 is to calculate an unknown quantity where 2 solutions or mixtures are proportional. The equation mean concentration(start)Volume(start)=concentration(final)Volume(final). Set equal and solve for the unknown. Lastly for stock solutions/dilutions refer to the last lab notebook where I discussed stock solutions and serial dilutions.

Bioskills: Excel

The last part of the pre-lab was over excel. I was fortunate enough to have an opportunity in high school to learn excel. Two years ago I took “Business Information Management” and got certified in Microsoft Excel so I already have know to make calculations in Excel. However I did learn how to access Excel through Baylor where documents in class will be uploaded. I wonder if results from our experiment will be uploaded onto Baylor’s Excel? Can I find away to use this resource in my research paper?

Lecture

Experiment Design and Results

The results of all our separate experimental designs is that us as a class want to see the effect of microplastics on Tetrahymena. Many groups used microplastics such as fibers, polyethylene, pH levels, Styrofoam, etc. Experiments monitoring lysosomes, cell counts, genetic mutations, and protein production were also very popular.

CILI-CURE 2018

Our CILI-CURE 2018 experiment will be testing the microplastics of baling twine on Tetrahymena. Baling twine is what keeps bails of hay together but once they are cut off they seep into the soil through toiling and other environmental factors. It already negativity effects macroscopic wild life such as birds like Osprey who use it for their nest but get tangled in it. But, how is it impacting on a mircoscopic level? This is why we will test the effects of microplastics from baling twine effect on the soil with soil ciliates; Tetrahymena. Baling twine is made up of polypropylene(PP) which is a very resistant material, normally tough and flexible, low in weight, easily to handle, in a variety of colors that provide UV protection, and is inexpensive.

Purpose

The Purpose of this lab is to focus on the big picture; our research. Now knowing our finalized experiment what are the expectations. Students will be given the information about the whole upcoming assignment and the purpose of this week is to wrap their head around it and starting thinking about when they will have time to devote to this research. The overall purpose of the lab is preparation.

Objective

The lab objectives were first, to practice for our experiments. More exposure to pipetting and cell counting achieved by perfoming serial dilutions and counting cells again. Students will finally see the effects of staining Tetrahymena by using Iodine. Also students will learn new assays of Tetrahymena by observing swim speed, percent directional change, and lysosomal behavior.

Hypothesis

My hypothesis is that students will be very successful at repeating and fine tuning those lab skills. I think they will be sad that they are not doing their labs they thought of in group as that was the impression I got. However, the assigned lab will be a lot of fun and I think it will uncover some great research results.

Procedures

Polypropylene Microplastic Production

1. Shred/cut white PP with scissors
2. Measure .5g of PP into a sterial glass jar/beaker
3. Add 50ml of sterile proteose-peptone-tryptone(PPT)media.
4. Boil and stir during lab (acts as heat from sun)
5. Autoclave twince juice to kill any bacteria in the mixture.
6. Next week: Filter using a 5µm filter paper into a sterile 50ml tube
7. Aliquot so each group is using the same microplastic

The Cilliate Count Challenge

1. Using a pipettor get 20 µl of Tetrahymena cells and place on a petri dish
2. Using a pipettor get 5 µl of iodine and mix with cells on petri dish by pipetting up and down
3. Transfer 5 µl from petri dish to a microscope slide
4. Notice that 5 µl drop is too concentrated
5. Dilute solution by a factor of 10^-1 by adding 180 µl to the 20 µl solution left in the petri dish
6. Observe cell counts and record in table

Behavioral Assay: Swim Speed

1. Place a 20 µl of culture on a clean flat slide
2. Set the slide on top of a metric ruler and adjust the slide so that you can see the cells on top of the mm marks on the ruler
3. Pick a cell to watch and line it up with the inside of one mark
4. Start the timer and watch the cell until it touches the inside of the next mark, stop the timer
5. A stopwatch can be started whenever a cell passes over one of the marks and stopped when the cell touches the next mark, but this is easier if you have one person watching saying “go” and “stop” to the partner. Only cells swimming straight during the entire length of the mark should be included.
6. Record the time and repeat for at least 10 cells
7. Swim speed can be expressed in mm/s
8. Calculate the average and standard deviation.

Behavioral Assay: Percent Directional Change

1. Add a 5 µl drop of culture to a clean flat microscope slide
2. Place the slide on a dissecting scope
3. Focus the slide on the black plate using reflected light
4. Choose a cell to follow for 10 seconds (use phone timer)
5. Keep track of how many times the cell changes directions by more then 17% during this time period
6. Record the number of direction changes
7. Repeat for at least 10 cells
8. Record the numbers and calculate the average number of direction changes/second
9. Calculate the standard deviation

Lysosomal Assay: 0, 10, 20, 30 minutes

1. Place 20 µl of Tetrahymena culture on a concavity slide
2. Add 1-5 µl of India Ink to the drop of cells and pipette up and down to mix cells and the ink.
3. Quickly place a cover slide on the drop and focus the slide on a 40x magnification on a compound microscope and start a stopwatch.
4. Scan the slide for 10 cells and count any vacuoles. This is your time 0.
5. After 10 minutes, count the number of vacuoles for 10 more cells
6. Repeat the procedure for 20 mins and 30 minutes.

Data

The Cilliate Count Challenge

Average= 140 cells; this means there was 140 in the solution

Behavioral Assay: Swim Speed

Riley did this experiment, and this is her data

Behavioral Assay: Percent Directional Change

Lou Gilbert did this experiment, and this is her data

Lysosomal Assay: 0, 10, 20, 30 minutes

Conclusion

Overall, this lab was very successful. Time was a bit of an issue as my lab group had to split the Tetrahymena Behavioral Assay experiments and we each only got to do one. However, Everyone one was successful with the Cilliate Count Challenge as we discovered a new technique. If you took a picture on snapchat and used the pen on the phone to count the cells it made it a lot easier. It was a fun lab and everyone achieved the objectives.

Next Step

With this knowledge from this weeks lab I feel prepared to start the research. I definitely have learned more information and skills then I knew on the first day of class. This lab got me really excited for the research project and seeing what we find. In the upcoming week I wonder how much time will be designated to CILI-CURE research? I wonder if other students are thinking ahead and planning time to research articles, read, and write?

9/20/2018

By: Marci Jordan

Bio Lab 1105-31

Pre-Lab and Lecture

Learning Theory: Understanding Deeply

As we learned before a fixed mindset means we cant change. The brain is able to do much more than we ask, therefore the brain can grow. Understanding deeply means asking a lot of questions to better understand the subject. Through questions you re-emphasize what you know and clarify things you don’t. After completing CILI-CURE, we will recognize and experience the diversity of life, process of experimental design, connections between living things, cell structure and function, microscopy, and the joy of learning and discovery. Through the process of completing the units, we must ask questions to deeply understand the purpose of why we do this to apply it in our scientific future. To understand deeply I’ve been asking questions like “How does this equation apply?” and “Specifically what are ciliates?” “Why is there such a wide variety of types of ciliates?”

Scientific Method and Experimental Design:

The process of the scientific method include asking a question -> Research -> Forming a hypothesis -> Designing an experiment -> recording data -> and coming to a conclusion, then repeat. It is necessary that the research question that is posted is testable. For this experiment, we are designing in very classical scientific method to form an experiment that has a single variable and control group.Epistemology is the branch of philosophy concerned with theory of knowledge. Epistemology studies nature of knowledge, justification, and rationality of belief. With our experiment with Tetrahymena and microplastics, how does this affect the overall soil environment? How long has it been a problem? When did microplastics peak to where it became a negative impact?

Deductive and Inductive Reasoning

Scientific research is described as “The process of determining some property y about something, to a degree of accuracy sufficient for another person to confirm this property y” (Glass). Experiments can use both types of reasoning; experiments are designed and performed using deductive but applied using inductive. Inductive reasoning involves evolving from a series of specific cases into a overall general statement or prediction. Therefore, the conclusion in inductive arguments is never guaranteed because the experiment is yet to be performed. In our course we want to design an experiment with some kind of statistical probability for the effect of x on y. Literature will be used to help make a good design.

Hypothesis: Falsification Framework and Experimental Design: 24 well plate

Philosophy of sciences varies depending on area of study and researcher perspective. Not all science is hypothesis driven, but for our experiment it’s okay to try to falsify a hypothesis based on results. The purpose of a 24 well plate is to test the effect of substance x against Tetrahymena growth rate in a culture. As seen later in this lab report after performing serial dilutions we went to the computer lab to complete experimental designs with our lab group. My group already had an idea to it was just a mater of getting it on paper. Our detailed experiment idea is listed below but I had a few questions when thinking about the 24 well plate. When forming our experiment I didn’t know what measurements to use. How much Tetreahymena should I put in each well? How much mncroplastics should I put in each well? How will I make the mass of microplastics precise?

BioSkills: Serial Dilutions and Calculating the Concentration of Cells in a Culture

To count the number of cells in a culture, we learned to preform a dilution. A dilution involves using micropipettors and a stock culture. When reading the pre-lab I was very confused as there were detailed steps about how to dilute but I was unfamiliar with the process. Once in class we performed a skill performance task of diluting our own stock solutions in a 24 well plate . You start with one well with the full amount of stock solution given. Then you take a smaller portion of that mixture and mix it with the next well. Then you apply the same process going down the chain. I wonder if this is the only order to dilute a solution? Are there other method? Once we begin our experiment will i need to dilute anything? The normal format for recording the concentration of a culture is in cell/mL. No one wants to count the entire mL or even a small dense volume as there could be a high concentration of cells. So instead we count a small volume of a diluted sample and compute the number of cells/ml.

Soil Collection

This is where we merge pre-lab to in class lecture. Prior to the pre-lab we were instructed that this week we would need to bring soil samples into lab. During pre-lab were we instructed why we need the sample, how and where to get the sample, the location of sample gathering based on lab section, specifically where on the tree to gather the sample, instruction to upload pictures and more information about the soil gathering on another website, and how to properly label our samples. Then in class the first thing we did was soil collections. We labeled the top and bottom of a petri dish with our full name and soil identifier. Next we measured the mass in grams of the empty dish (bottom only) and then the bottom dish plus enough soil to cover the bottom. My data is shown below. Then we covered it and stored them in the fume hood. They will stay here for about a week to dry them out so the Tetrahymena form spores.

Purpose

The purpose of this lab was to get us familiar with serial dilutions. As I mentioned earlier reading about serial dilutions on the pre-lab was confusing so having hands on experience would clear up a lot of questions. We also gained more experience properly using I micropipettor which is very valuable knowledge for when working in a lab in the future. Lastly, to construct our experiment for the rest of the semester. Now we have a solid question that we can apply and research.

Objective

The objective is to successfully deeply understand how to dilute a solution. To understand what is happening each step and correctly calculate the cell concentration of the sample. Also, to properly use micropiprttors of various volumes, with different tips. Lastly, to form a proper experimental design dealing with Tetrahymena and microplastic pollution. It must be able to falsify and very detail that it is clear what is the question and can anyone replicate the experiment.

Hypothesis

I believe that students will mess up diluting the solutions are many wont read all the way through first. However, I think everyone will perfect and master using the pipettors correctly. I believe forming proper experimental designs will be hard as some groups were changed and I know some individuals already had ideas that there group might not want to do. Also, it is hard to find information or research on Tetrahymena and microplastics so difficulty of research success will impact forming a experimental design.

Procedures

In Lab

Goal: To determine the concentration of Tetrahymena in the “stock” solution.

1. Begin by observing the Tetrahymena in the “stock” well of your 24-well plate using the discecting microscope. This is your undiluted sample or 10^0
2. Record your observation in lab notebook: living, dead, concentrated….

Performing a 10-fold series dilution:

1. Label 4 wells of the 24 well plate: 10^-1, 10^-2, 10^-3, 10^-4
2. Add 900 µl of Tetrahymena culture media to the 4 wells
3. Add 100 µl of 10^0(undiluted) stock culture to the “-1” tube, mix briefly by slowly pipetting up and down. Change tips

When pipetting from the Tetrahymena stock cultures, avoid the bottom of the well where dead cells and debris have accumulated. Living Tetrahymena cells are located near the surface. Develop a consistent method for removing cells from the same vertical level. You may pipette while observing your cultures under the dissecting microscope while learning this technique. Learning to be consistent reduces sampling error.

1. Add 100 µl of 10^-1 sample to “-2” well, mix briefly by slowly pipetting up and down. Change tips.
2. Add 100 µl of 10^-2 sample to “-3” well, mix briefly by slowly pipetting up and down. Change tips.
3. Add 100 µl of 10^-3 sample to “-4” well, mix briefly by slowly pipetting up and down. Change tips.

In Computer Lab

Get together with group and create a experimental design with Tetrahymena and microplastic pollution. Use the QTM as an outline to aid in this process. The goals of today is to determine the research question that we want to address and create experimental design/method. Meet with members of your assigned team and discuss the experimental design that you would like to propose.

Data

In lab

In Computer Lab

My group decided to see how microplastics affect Tetrahymena’s behavior. A more detailed methods description is on the QTM and not recorded in my lab notebook. We will be watching 4 different enivorments within a 24 well plate. Group 1 will be the control, group 2 will have .005g of microplastic pollution, group 3 will have .1g of microplastic pollution, and group 4 will have .15g of microplastic pollution. All wells will have 1000µl  of Tetrahymena extracted from our soil samples. We will observe them for 1 week and see how the different various concentrations of microplastic pollution affects them compared to the control group (group 1) with no microplastic pollution.

What Went Wrong

In the in lab procedures above I only listed the procedures that we did. Felicia announced at the beginning that we would not be doing it all as we needed to go to the computer lab. After all we just needed to learn about dilution so it is the only part we did. The above is not the accurate data that I got from what I saw in my microscope in the lab. The whole class was having problems seeing the Tetrahymena at different dilutions; dilution D1 being the hardest. Although I couldn’t see ciliates I still learned about dilutions which was the goal. I understood that as you went down the chain the solution became more diluted which means less concentration of cells which means less cells. Therefore, I inferred my own numbers per well and had them decrease as the dilution increased. I showed Felicia my numbers and she said they were acceptable. I used the numbers I created and used them to work through the equations to find the columns in the table. Therefore, I understand what effects dilutions have on the concentration of cells and how to use the equations to find various methods of measurements.

Conclusion

After lab today I learned a lot of valuable information on a wide variety.I now deeply understand what serial dilution is and how it is performed. I consider myself a professional at micropiettes and feel more confident in a lab. I’m very excited that my group has our experimental design set up and Felicia said it was great and will be a good idea to test. I’m very excited for the rest of the semester as I can now apply what we have been learning in class.

Next Step

My next step in this process is all about application. The past weeks have been learning about skills needed for our research project. But now the time has come and I cant wait to get my hands dirty. My own first lab experiment!! Skills such as dissecting microscopes, compound microscopes, micropipettes, making wet mount slides, staining, and now series dilutions are coming together to see the effect that microplastic pollution has on Tetrehymena, the best model organism.

9/13/2018

By Marci Jordan

Bio 1105-31

Pre-Lab and Lecture

Learning Theory

Although we learned about learning theory last lab with fixed and growth mindsets we applied that to science. Through an article I learned how creativity powers science. Associative thinking is best when doing a relaxing activity and allows new great ideas. Along with the article a video was provided discussing the great discoveries that were made by using Tetrahymena. These discoveries were made by people who were thinking creatively and outside the box. Who ever would of thought these small, abundant, and cute cilliates could help with cancer research and even win two noble prizes! We also discussed in class lecture why ciliates are such good model organisms. Unlike mice or other model organisms, ciliates are cheap and reproduce quickly. Therefore an experiment that would take months is quickly advanced as results are available quicker by rapid reproduction. You can get 10 ciliate generation in a week while it would take months for other species. I would who discovered how beneficial ciliates are in a lab? How long have ciliates been chosen as a model organism? Are there any organisms that are better model organisms then ciliates?

Nature of Scientific Communication

In the pre-lab I was notified about the unique opportunity the Baylor Library offers. They often have “Science Workshops” that help students understand the process of scientific communication. This is valuable information as throughout school it will help students understand how to properly communicate and also what to look out for. Next as a scientist its important to know to evaluate scientific sources. This requires knowing where to look and what to look for. Usually primary or secondary scientific literature sources are the best choice. With a overwhelming amount of sources available on the infinite internet evaluating the validity of a source is necessary. Questions such as where source was published, was it peer reviewed, author, authors reputation, date, and type of source are good to decide if it is a good article.

Once sources are found you need to cite them in your paper. Helpful tools were listed in the pre-lab such as “Zotero” and “RefWorks” which are tools that collect, manage, and cite research sources. This is critical as most professors will deduct heavily or fail a paper if it does not have the correct citation. With improper citation comes plagiarism, either on purpose or accident but its all the same, and is against the honor code. In addition to OneSearch, Baylor also offers a database called “PubMed” which can find articles with keywords. I wonder what most Baylor students perfer, OneSearch or PubMed? I wonder if one is better for different purposes? I wonder which one has more articles about Tetrahymena?

Basics of Tetrahymena

As stated before Tetrahymena are great model organisms for experiments. They have been used many times for biochemistry and cell functions. But Tetrahymena have been used extensively in research of studying the cytoskeleton and sub-cellular components. I stated above that Tetrahymena easily reproduce and this is due to how their genome is sequenced and available for genetic manipulation. I wonder how far we will go into Tetrahymena? Will we conduct experiments to manipulate their genome?

Getting Ready for “Experimental Design”

During lab we worked with our partners to design an experiment to be performed on Tetrahymena. We decided to test how Tetrahymena are effected by microplastic pollution. Microplastics are pieces of plastic that are less then 5mm big. Our experiment will consist of 3 different enviorment. The first group being the control will have Tetrahymena only to track normal growth, function, and behavior. Then the other two environments will have different concentrations of microplastics within the environment with the Tetrahymena. One group will have a small amount of microplastics while the third group will have a high concentration. Then over time we will observe how the Tetrahymena are effected by the different concentrations. The independent variable is the amount of microplatics we decide to include in the environments. The dependent variables will be the outcome such as the Tetrahymena’s cell structure, behavior, function, reproductive rate, appearance, and life span that have been changed from the pollutants.

The question being asked is how are tetrahymena affected by microplastics in their environment? My hypothesis is that the more microplastics in the environment the less successful the Tetrahymena will be. For example, they will have slower reproduction rates, slower movement, less function, and a change in shape. I will test these ideas in the procedure listed above. Data will be taken by observing each environment over time. I will measure growth, function, behavior, reproduction rate, movement, and appearance. Results will be analyzed by comparing results at the end to the beginning.

Micropipetting

Lastly in pre-lab and lecture we learned how to use a micropipette. This was my first experience with a micropipette but I had a lot of fun. In lecture we got a demonstration about the dos and donts. We learned how to calibrate it to the desired volume. We had to be careful to not set it above or below its capacity (found on the pipette) as it would become uncalibrated. Then we learned how to put the proper tip onto the pipette shaft. We practied transporting various volumes of water. Once we mastered the skill we used the pipettes to put Tetrahymena samples onto concavity slides. A micropipette’s 1000µL=1mL and 1000mL=1L. I wonder how many student in class have previously used a micropipette? I wonder if anyone used it incorrectly?

Purpose

The purpose of this lab is to work on skills using a micropipette, dissecting scope, and compound microscope. Get aquainted with Tetrahymena and develope a scientific experiement which involves them in addition to microplastics. Lastly, find an article to enter us into the mind frame of Tetrahymena and microplastics.

Objective

To get familiar with the model organism Tetrahymena. Also to develope new skills with a micropipette in addition to reviewing the dissecting and compound microscope. Another objective is to correctly find a primary source and identify what makes it good. Lastly, to create an experiment using Tetrahymena and microplastics.

Hypothsis

My hypothesis is that students will enjoy observing a new ciliate. Collaborating with their lab partners will increase creativity to think of a experiment. This will assist in combining science and creativity in a relaxed environment. Lastly, I think students will be able to find article on Tetrahymena or microplastics but not together as I don’t think there is a lot of research done on those two combined specifically.

Procedures

Wet Mount

1. Transfer 100 µl of Tetrahymena stock culture to a clean well using the correct micropipettor.
2. Observe Tetrahymena in 24 well-plate with dissecting microscope.
3. Pick 5 µl of cells using a P-10 micropipette
4. Transfer 5 µl to concavity slide and observe on 4x and 10x using the compound microscope.
5. How many cells can you count per 5 µl?
6. Using FOV measurements, approximate the diameter of Tetrahymena
7. Record your procedure and observations in lab notebook.

Data

Trial #	1
# of cells in 5 µm	300
Approximate diameter of cells	170
Diameter in mm	Mag: 4x = 4mm
Diameter in  µm	Mag: 4x = 4,000 µm
Size of cell in  µm	23.54 µm
Drawling

 Conclusion

Tetrahymena are very very small. They are very abundant and are great model organisms. They are great for experiments as they reproduce quickly and are cheap. Micropipettes are good for exact measurements and getting a small drop on a slide. I still need to work on my skills to focus a compound microscope. Microplastic pollution is a huge problem even if it is physically small (get it, because its smaller then 5mm). Tetrahymena have been used in a wide variety of experiments so why not see the effects of microplastic pollution on them. There is not many sources on Tetrahymena so as the CILI-CURE class we should research and make our own discoveries.

Next Steps

After today in class I was motivated to help to planet. Be more aware of whats recyclable and make sure all trash is disposed of properly. I am very excited for the opportunity to create my own experiment and test it. Tetrahymena are so cool and have done so much I cant wait to test my theories on them. Once my experiment is done I can use the results to show others the effects of microplastic pollution and how serious of a problem it is.