Hanna Kim Lab 3 - Cells Report_Sp24 (1)

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Feb 20, 2024

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Lab Report LAB 3: Cells Bio 1AL Spring 2024 Last Name Kim First Name Hanna Lab Section # 415 Lab 3 Report 1) (1 pt) In the field of view circle below, draw an Amoeba at 200X total magnification (using the 20X objective). a) Label the following structures: pseudopods, food vesicles (and contractile vacuole if observed ) . b) Include a scale bar with clearly labeled length and units. (use reasonable values). Note that m (distance/length) is a different unit than M (concentration). µ µ Drawing instructions: If you hand-drew the drawings, you can take a picture of the drawings and insert the image below. Total Magnification: 200X 3-1

Lab Report LAB 3: Cells Bio 1AL Spring 2024 2) (3 pt) In the left field of view circle below, draw a Paramecium cell and a few surrounding yeast cells. a) Label the following structures: cilia, contractile vacuole, yeast cell and oral groove b) Include a scale bar with clearly labeled length and units. c) Use the reticle calibration from Lab 2 to determine the length of the Paramecium , and the diameter of a yeast cell. Include appropriate units in your answers in Table 1 below. On the right field of view circle below, draw a stained cheek cell from your slide. d) Label the following structures: nucleus, cell membrane, cytoplasm e) Include a scale bar with clearly labeled length and units (use reasonable values). f) Use the reticle calibration from Lab 2 to determine the widest part of the cheek cell, and the diameter of the nucleus. Include appropriate units in your answers in Table 1 below. Drawing instructions: If you hand-drew the drawings, you can take a picture of the drawings and insert the image below. Total Magnification 400X Total Magnification 400X Table 1. Calculated Dimensions of Cellular Structures Paramecium Cheek Cell (measured Total magnification: 400X Total magnification: 400X Length (long axis): 250um Cheek cell diameter: 82.5um Yeast cell diameter: 5 um Nucleus diameter: 10 um 3) Follow the Lab 3 procedures to measure rates of cytoplasmic streaming in Nitella . Record your streaming rates below and on the class data sheet - use two decimals for your rates. For the class data sheet, record your streaming rate in the corresponding green cells in the class data sheet by station number. 3-2

Lab Report LAB 3: Cells Bio 1AL Spring 2024 Baseline rate (pre-FCCP) Rate After FCCP Treatment Experiment 1 93.02 2.37 Experiment 2 53.76 2.91 4a) Once all streaming times and streaming rates have been entered for all lab station groups in your lab section, download your class datasheet as an Excel File ( File > Download > Microsoft Excel ). 4b) In Excel, calculate the average, standard deviation, and standard error of the mean (See Excel Graphing Resources page) for the cytoplasmic streaming rates before (baseline) and after FCCP addition (cells highlighted blue ), using your lab section’s class data. Copy the values for the average streaming rates and SEM into Table 2 below. 4c) Calculate the percentage change between the two means and enter into Table 2. See the Comparing Two Means page in bCourses. 4d) Run a Correlated (Paired) Sample t-test in VassarStats comparing the average baseline cytoplasmic streaming rate to the average streaming rate after FCCP addition. See the Paired/Correlated Sample t-test page in bCourses. Use the one-tailed p-value , since we are predicting that FCCP will decrease cytoplasmic streaming rate (i.e. we are predicting an effect in one specific direction). Record the relevant values in Table 2 below. Table 2. Effect of FCCP on cytoplasmic streaming rate in Nitella Baseline After FCCP Mean streaming rate ± SEM (μm/sec) 43.8 ± 2.7 10.6 ± 1.4 Percentage change (%) -75.9% Paired t-test results t = +10.24, df = 27, p (1-tailed) < 0.0001 5a) (0.5 pt) What is your null hypothesis for the t-test? There is no statistically significant difference between baseline rates of cytoplasmic streaming and the rates of cytoplasmic streaming after the addition of FCCP, and any differences are due to random chance. 5b) (0.5 pt) Based on the results of your t-test, does FCCP addition significantly decrease cytoplasmic streaming rate? Circle/highlight: YES / NO 6) (1.5 pt) Using Excel, graph the average cytoplasmic streaming rate before (baseline) and after FCCP addition from the class data. Use a column graph in Excel, and add custom error bars, using your calculated SEM values. Save the graph in Excel as an image (Ctrl Click on the graph > Save as 3-3

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Lab Report LAB 3: Cells Bio 1AL Spring 2024 Picture or take a screenshot) and paste the image below. 7) (0.5 pt) We are missing a DMSO-only control experiment in our experimental design. In a few sentences, briefly describe the key steps of this control experiment. 1) Make the slide as usual, following the previous steps to create a slide (creating a wet mount with Nitella and 90mL of pond water → 100ml total). 2) Calibrate the microscope and follow all steps (Kohler illumination, focusing, etc) until reaching the wanted objective (200X or 400X, depending on size of slide). 3) Measure the baseline streaming rate as previously followed. 4) Add 100ul 2% DMSO/Total volume (200ul) to create a 1% DMSO concentration. 5) Wait ten-twelve minutes for DMSO to spread to the nitella. 6) Measure the streaming rate again. - To calculate the rate of FCCP only: (Rate FCCP + DMSO) - (Rate of only DMSO) = Rate of FCCP only 8a) (0.25 pt) How did the cytoplasmic streaming rate in Elodea compare to in Nitella? The streaming rate in Elodea was: Circle/highlight one: FASTER SLOWER 8b) (0.25 pt) Briefly discuss one factor that may cause a different streaming rate in Elodea . (1-2 sentences) The elodea may have a slower streaming rate due to slower myosin ATPase activity. Slower ATPase activity means that the fibers contract slower, resulting in slower rates of cytoplasmic streaming and transport. 3-4

Lab Report LAB 3: Cells Bio 1AL Spring 2024 9) This question refers to the Ronald Vale paper that was discussed in Lecture 3. The abstract and Figure 4 are shown below. 9a) (0.5 pt) Let’s start by only looking at the bars labeled “N” in figure 4, which are the results of experiments using the normal myosin purified directly from scallops. Describe what you can conclude from comparing only the bars labeled “N” on the left side and right side of figure 4. The addition of Ca2+ (calcium) ions increases and regulates scallop myosin movement on actin, increasing their velocity. When Ca2+ is absent, there is little to no movement of the myosin. When Ca2+ is present, there is high movement. This indicates that Ca2+ is needed for the normal movement of scallop myosin. 9b) (0.5 pt) Now look only at the right half of figure 4. Compare only the “N” bar to the “D” bar. What does this tell us about the role of the regulatory light chain? The regulatory light chain is required for regulation of motility by Ca2+. Although calcium was present, desensitized scallop myosin had a significant decrease in velocity compared to the normal scallop myosin. Based on this fact and the data above, we can infer that calcium binds to the regulatory light chain in scallop myosin, allowing for motility. If the regulatory light chain is removed from scallop myosin, Ca2+ is unable to bind to the scallop myosin, decreasing its velocity. 3-5

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