10-Photosynthesis Report_2023

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Colorado State University, Fort Collins *

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212

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Biology

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Jan 9, 2024

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LIFE 212 Laboratory Report 10 Name: _____________________________ Due: _____________________________ Section: _____________________________ The Study of Cellular Functions Part 3: Isolation of Chloroplasts from Spinach Leaves Photosynthesis: Hill Reaction 50 pts total plus Notebook Conclusion (4 pts) 1. Part B: Determining Chlorophyll Concentration a. (Step 13): Absorbance of chlorophyll: A 652 = ____________________ (1 point) b. (Step 14): Calculate the amount of chlorophyll in 1 mL (in mg/mL) of your chloroplast suspension. Show your calculations. (1 point) 2. Part C: Hill Reaction Assay a. (Step 15): What is the volume of the chloroplast suspension needed to make 10 ml of a 0.01 mg/ml diluted suspension? (4 points) b. What is the dilution factor needed to yield a final chlorophyll concentration of 0.01 mg/ml? Hint: dilution factor is C1/C2 or V2/V1. (2 points)

LIFE 212 Laboratory Report 10 Name: _____________________________ Due: _____________________________ Section: _____________________________ 3. Record the absorbance of the chloroplast reaction tubes below: (30 points total) Absorbance (A 600 ) / time Reaction Number 1 (control) 2 (dead) 3 (active) 4 (dark) 5 (DCMU) 0 min after adding DCPIP - 10 min after - - 20 min after - - 30 min after - a. Calculate the amount (in mg) of dye reduced at each 10 minute interval for Reactions 2, 3, 4 and 5 (only after 30 minutes for Tube 4) using the equation below. The molecular weight of DCPIP is 290.08 g/mol. Show your work with one sample calculation. Look up the DCPIP concentration in the Materials. (7 points) Note: The reaction volume is 6 mL. Cr = Co(Ao-At)/Ao Where: Cr is the dye reduced in mmol/liter (mM) Co is the original dye concentration in mmol/liter (mM) Ao is the absorbance of the reaction at the start At is the absorbance of the reaction at each 10 minute interval Reaction Number 2 (dead) (mg) 3 (active) (mg) 4(dark) (mg) 5 (DCMU) (mg) 10 min after 20 min after 30 min after

LIFE 212 Laboratory Report 10 Name: _____________________________ Due: _____________________________ Section: _____________________________ Sample calculation:

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LIFE212 LAB REPORT 10 Name: ______________________________ Due: ___________________________ Section: ______________________________ b. Create and paste in the space below a line graph showing the change in the amount (mg) of reduced dye in each of the reactions versus time. (7 points) Note: Use a scatter graph with connected lines, NOT a best-fit trendline. c. Explain your results in each of the reactions. What is the trend and why? Reaction 2: (4 points) Reaction 3: (4 points) Reaction 4: (4 points)

LIFE212 LAB REPORT 10 Name: ______________________________ Due: ___________________________ Section: ______________________________

LIFE212 LAB REPORT 10 Name: ______________________________ Due: ___________________________ Section: ______________________________ Reaction 5: The photodissociation of water occurs within photosystem II with electrons ultimately flowing to photosystem I. DCMU stops the flow of electrons between photosystem II and I. Comparing the amount of dye reduced in reaction 3 to reaction 5, what do you conclude about the action of DCMU in photosynthesis? How does it affect plant growth? Explain and support your answer. (4 points) 4. Reactions 2 and 4 were included as experimental controls. What exactly do they tell us about the experimental results? (4 points) 5. Why did you chill your glassware and complete all of your centrifugation at 4 o C? (4 points)

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LIFE212 LAB REPORT 10 Name: ______________________________ Due: ___________________________ Section: ______________________________

LIFE212 LAB REPORT 10 Name: ______________________________ Due: ___________________________ Section: ______________________________ 6. Why did you centrifuge the supernatant in Step 7 at a higher g force? Would a greater g force yield more chloroplasts? (4 points) Notebook Conclusion (4pts) The conclusion should be a concisely written summary of the experimental results using technical language. Describe each objective from your Introduction individually and frame in context with the experimental results. Vague and/or incomplete conclusions will not receive any credit.

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