Cell Communication Lab

Purpose

The purpose of this experiment, as the title suggests, was to study cell communication. Our subject was yeast and we tracked it's cell count over fourty eight hours. The controled variable in this experiment was time, where else the independent variable was percent of total cell count. As time passed on, we hoped to see whether the percent of total cell count would decrease or increase- mostly the latter. 

Introduction

Since yeast cells are incapable of moving, they communicate through signaling molecules. These signaling molecules, after being sent by one cell, are received by a G-protein coupled receptor in another cell. The pathway of the signaling molecule can be seen in the diagram below, supplied by Pearson Education. 

The cellular response in this case, is for the yeast cell to grow and divide. In our a and alpha culture of yeast, this will be seen as a budding haploid cell, where else the cells that have yet to receive the signal to grow will be single haploid cells. In our mixed culture, single and haploid cells will be present as well, along with schmoos, and asci. In a mixed culture of a and alpha yeast cells, the cells may recognize each other's presence and beging to grow towards each other- hence what we refer to as schmoos. Asci are bundles of cells growing next to each other, another aspect unique to mixed cultures.

Methods

In this lab, we put alpha-type, a-type, and mixed-type yeast in culture tubes filled with water.  We took samples of 5 drops of the yeast after 0 minutes, 30 minutes, 24 hours, and 48 hours and put them on viewing slides, then examined them with three different fields of view with compound microscopes on a medium 400x magnification. We counted the numbers of single haploid, budding haploid, shmoos, single zygote, budding zygote, and asci cells in each field, then calculated the percentages of each. 

Data Charts

Graph

DISCUSSION

 While observing our yeast through the microscope, no noticeable differences of alpha and a yeast could be seen. Even though they do have different genetic make up. The yeast that were on their own were mating at lower rates than the mixed yeast. In the mixed yeast, schmoos were visible sooner than the individual yeast. Yeast can communicate both indirectly and directly because if some yeast change, then they can send different signals out. When the yeast mate they can receive a signal directly from the opposite gender of yeast cells. Signal transduction pathways combine A and Alpha cells. When A and Alpha cells are together they are mixed. Mixed yeast cells can be haploid, zygotes, budding haploid, or even a shmoo.  When yeast cells mate, they use signal transduction pathways because G Protein receptors help them mate. 

 In our experiment the alpha yeast had more budding yeast cells than the A yeast cells. Our percent totals were approximately the same for alpha and A because they were leaning towards budding haploid cells more. In the mixed yeast cells, since there were A and Alpha, they reproduced sexually. On the other had the individual A and Alpha cells reproduced asexually. The Mixed Yeast cells started with schmoos then by the twenty four hour observation they were haploid cells. Then the haploid cells divided to form asci by the third day. According to our data, as the time went on, many more cells were produced due to reproduction. 

CONCLUSION 

     We are studying Cell Communication in this lab and we controlled the time when we observed the progress of the yeast cells. We observed cellular responses and the growth, then dividing of the yeast. Our percent totals of cell count increased as time when on. Cells communicate with receptors, especially G Protein Coupled Receptors, which aid in the reproduction process. Signal Transduction is also a method for the cells communicate. 

Plant Pigments and Photosynthesis Lab

PURPOSE

4A
The purpose of this experiment was to use chromatography to separate and identify the various pigments in chlorophyll.

4B
The purpose of this experiment was to determine the change in the rate of photosynthesis of chloroplasts when boiled or removed from light by using DPIP to determine the chloroplasts' electron output.

INTRODUCTIONS

4A Paper chromatography separates pigments by dissolving them in a solvent that moves up a strip of paper by capillary action. Different pigments are carried different distances by the solvent because of their varying solubility and attraction to the paper. Pigments like beta carotene that are more soluble and less attracted to the paper will be carried further than less soluble, more attractive pigments like xanthophyll. The distance traveled by the pigments is called Rf and is always the same for a certain pigment. It can be calculated as the quotient of the distance the pigment moved divided by the distant the solvent front moved. There are many light-harvesting pigments in chloroplasts. Chlorophyll a is the main photosynthetic pigment, absorbing blue-violet and red light best. Chlorophyll b and carotenoids are accessory pigments that are used to extend the plant's light absorption and to protect the chlorophyll a from too much harmful high-frequency light. 4B
Chloroplasts are found in plants and absorb light energy to "excite" electrons removed from water to produce ATP and NADPH, which are then used to fixate carbon into sugars in the Calvin cycle. However, the high-energy electrons produced that reduce NADP+ to NADPH can also reduce other electron acceptors, like the dye DPIP. DPIP is blue before reduction, but becomes colorless as it is reduced. DPIP, when blue, only transmits blue light, while colorless substances transmit all wavelengths of light on the visible spectrum, so as DPIP becomes colorless, it transmits more and more light. This transmittance can be measured using spectrophotometer.

METHODS

4A

Using a quarter to scratch off pigment off a spinach leave and placed that on a pencil line on the paper.  We took the chromotography paper and placed it into a cylinder with solvent. We waited time to see the pigments spread out over the paper. We mark where the bottom of the pigment band is. We measured the distance the pigment band moved. 

4B

In this experiment we set up a flood light, heat sink, and cuvettes. There were boiled and unboiled chloroplasts. We had five cuvettes the first was our control group. We put phosphate buffer and Distilled water to test tubes then DPIP to certain test tubes. We transferred the solutions to the correct cuvettes then added chloroplast to the cuvettes.  Immediately after adding the necessary chloroplasts we placed them behind the heat sink. We waited to place each in the colorimeter to read the rate of photosynthesis. Then tracked it on the logger pro. After taking them out of the colorimeter the cuvettes were placed behind the heat sink again.

GRAPHS

4A:

4B:

DISCUSSIONS

4A

From our trail, it appeared that the spinach sample separated into five different pigments. Due to the capillary action habits of plant pigments discussed in the introduction, we can guess that the top segment outlined in the corresponding picture above is beta carotene and the lowest segment is possibly xanthopyll. 

4B

To be completely honest, our experiment ended up being a complete flop due to incorrect usage of the colorimeter as can be even seen by our rather pointless data. However, I would like to ask the reader to focus on line 3 in Table 4.4 above. After realizing the mistake that had been made, we added a few more drops of DPIP into the cuvette containing the unboiled chlorophyl that was exposed to light. As can be seen from this set of data, the longer it was exposed to the light, the more transmittance was detected. This shows that over time, the DPIP was being used up in photosynthetic like reactions at a moderate rate. However, we don't have any other reliable data to compare it with but we can assume that transmittance in this cuvette was increasing at a faster rate than cuvette #2 yet slower than the boiled samples. 

CONCLUSION

4A 

From our trial, chlorophyll divided into five pigments.

4B

...I don't know what to say other than if time had been permitted we would have probably redone the experiment.

Cell Respiration Lab

PURPOSE 

This lab is based on Cellular Respiration and how temperature of germinating and dominant seeds affect the temperature. We are going to be able to relate gas production with the rate of respiration. 

INTRODUCTION

The germinating seeds are the seeds that are in the process of growing. Cellular Respiration is a catabolic process that produces ATP.  The general formula is 

C6H12O6 + CO2 ----> 6CO2 + 6H2O + energy. 

There are three steps to Cellular Respiration Glycolosis, Krebs Cycle, and Electron Transport Chain. Glycolosis occurs in the cytosol and oxideses glucose partially into two pyruvates. Glycolosis unlike Krebs Cycle (Citric Acid Cycle) does not require oxygen.  The Krebs Cycle takes place in the Mitochondria. This process starts with the pyruvates from glycolosis and the pyruvate is now Acetyl-CoA. It is broken down into carbon dioxide. Both Glycolosis and Krebs Cycle produce ATP, but small amounts via Substrate Level Phosphorylization. The Electron Transport Chain is in the inner membrane of the mitochondria. This couples the transfer of electrons between the donor and acceptors of electrons. Consumption and production of carbon dioxide is used to measure Cellular Respiration. 

Method

The first batch that we tested were 25 germinated radish seeds kept at room temperature (which using a thermometer we measured to be 22 degrees Celsius). We placed them into a plastic chamber which got sealed by the CO2 sensor we had connected to our Lab Quest. We let the sensor run for 10 min throughout which we had the Labquest collecting data. Once we had sufficient amount of data, we took the seeds out and placed them into a glass beaker of ice cold water (15 degrees Celsius). After giving them some time to cool off, we took them out, blotted them dry, and repeated the CO2 measurements we had done previously. We then repeated the original process (meaning not including the ice water) with non-germinated radish seeds kept at room temperature and then separately glass beads for the purpose of having a control group.

Data

Graphs Germinated radish seeds, room temperature

 

Germinated radish seeds, ice water

 

Non-germinated radish seeds, room temperature

 

Glass Beads

 

Combined Graph

Discussion

  When the rate of respiration of the germinated radish seeds at 22 degrees Celsius was tested, approximately 0.61 ppm of CO2 per second was found to be produced through the seeds' cellular respiration. The rate of respiration of those same seeds after being soaked in water at 15 degrees Celsius was 0.70 ppm per second. Although most other germinated seeds started to become dormant at lower temperatures, with a reduction in respiration, the radish seeds became more active, suggesting that their optimal temperature for germination is relatively cold. The dormant, non-germinated radish seeds respirated less, with only 0.20 ppm per second, evincing their decreased metabolic activity. The change in CO2 per second with glass beads instead of seeds was -.02 ppm, a very minor change that may have been due to a small inaccuracy of the CO2 sensor or the movement of the CO2 molecules within the bottle, as the amount of CO2 would not expect to change without an organism present. The experiment did not seem to have any other glaring errors. However, if done again, a better method for removing the seeds from the cold water might save time and prevent any possible damage or stress to the seeds during handling. It also may have been prudent to use objects more similar to the radish seeds in size than the glass beads to prevent the differences in pressure from affecting the CO2 reading. The radish seeds were originally expected to respirate less at low temperatures, as most seeds become dormant when the temperature cools during the winter, but some types of seeds do grow better in the cooler months. The rest of the results followed the predictions.

Conclusion

 In this lab, it was found that germinated seeds respirate and produce CO2 at a greater rate than ungerminated seeds, and that germinated radish seeds respirate more at cool temperatures. From this, we can conclude that radish seeds have an optimal temperature for growth of less than 22 degrees Celsius.

References

Our wonderful Pearson AP Biology book :)

Enzyme Lab

Purpose

    In this lab we are observing the concentration of Hydrogen Peroxide (H2O2) in water and oxygen gas by the enzyme catalase. Then we are going to measure the amount of oxygen generated and calculate the rate of the reaction. Also, we will observe environmental factors such as pH and temperature changes.  

Introduction

   Enzymes are proteins that can speed up or slow down reactions. They are known as catalysts in reactions. Enzymes are the only thing not changed throughout the reaction. Enzymes can become denatured due to heat and pH changes. When they are denatured the enzymes become biologically inactive. Enzymes have specific duties and their active sites interact with certain substrates.

Methods

Part 2B: In this part, we put 10 mL of a 1.5% H2O2 solution in a clear plastic cup and added 1 mL of water, then 10 mL of 1 M H2SO4 solution, using a 1 mL and 10 mL syringe respectively. We mixed the resulting solution, then took a 5 mL sample of it and placed it in a separate clear plastic cup. Using a burette, we titrated the sample drop by drop with a 2% KMnO4 solution until the sample turned pink then brown and measured the amount of KMnO4 had been used.

Part 2C: In this part, we followed the same procedure as part 2A, but used a sample of 1.5% H2O2 solution that had been decomposing for 24 hours instead of the fresh 1.5% H2O2 solution.

Part 2D: In this part, we followed the same procedure as part 2A, but instead of the 1 mL of water, added 1 mL of a yeast solution that acted as a catalase to the H2SO4. We allowed the mixture to sit for 10 seconds  for reactions to occur before titrating a 5 mL sample. We then repeated the process seven times, allowing the reaction to be catalyzed for each of 30, 60, 90, 120, 180, and 380 seconds before titration.

Data

Graphs & Charts

Discussion

Part 2B: The initial reading of the burette was 13.5 mL. After completion of the experiment, the burette’s reading had dropped down to 10 mL. The baseline was calculated by subtracting the initial reading from the final reading, giving us the result of 3.5 mL of KMnO(4). This means that the initial amount of H(2)O(2) present in the 1.5% solution was also 3.5 mL. The other two groups in our class had results ranging from 3-4 mL so it’s safe to say that our results were rather valid. To ensure results, it’s never a bad idea to redo a baseline but in this case we didn’t see any reason to have to do so.

Part 2C: The initial reading of the burette was 26.5 mL and and after the completion of this experiment, the burette reading was at 31.0 mL. This resulted in 4.5 mL of KMnO(4) titrant used. Therefore, the amount of H(2)O(2) spontaneously decomposed (mL baseline-mL KMnO(4)) was 1.2 mL. The percent of the H(2)O(2) that spontaneously decomposes in 24 hours [(mL baseline-mL 24 hours) / mL baseline] x100 was approximately 3.5%.

Part 2D: Before starting the experiment, we once again conducted a baseline with the result this time being 3.3 mL of H(2)O(2) being present in the 1.5% solution. This was still in the range that most other groups were getting so there was no concerns with that. Even if the change had been slightly greater than that what we got the day before, it wouldn’t have caused much alarm because it was expected that some chemical changes may have taken place in the bottle that held the solution with it being constantly opened and closed. For both my sake and the reader’s, I will now direct you to look at the table above with all our data for this experiment, rather than me typing it all out once again. The biggest trend seen here is that there is an inverse relationship between the amount of KMnO(4) consumed and the amount of H(2)O(2) used. As the time given for the reaction to occur increases, the amount of KMnO(4) consumed decreases while the amount of H(2)O(2) used increases. The results obtained were within the range that we were looking for as well as the inverse relationship I just mentioned. We definitely ran into a little error the first time we tested the 30 sec reaction because the results we obtained did not match the pattern seen in the other experiments, but it was easily solved by simply redoing that one, so we strongly suggest to anyone else doing these experiments to keep the solution samples until you are 100% satisfied with the results you obtain :)

Conclusion

In summary, we learned the importance of conducting a baseline and that the amount of KMnO(4) used also represents the initial amount of H(2)O(2) in the 1.5% solution. Also, if you let a solution of 1.5%H(2)O(2) stand for 24 hours, a portion of it will spontaneously decompose. The most important thing that we got out of this those, is the concept that the longer an enzyme has to do its work, the more it can produce in form of the desired reactions.

 

Diffusion-Osmosis Lab

Purpose

1A: The purpose of this experiment was to determine the permeability of the dialysis tubing by identifying which molecules would or would not diffuse through it.

1B: The purpose of this experiment was to measure the effects of solute concentration on the osmosis of water through a selectively permeable membrane like a dialysis tube.

1C: The purpose of this experiment was to determine the water potential of potato cells and by measuring the diffusion of water into and out of pieces of potato in solutions of various molar concentrations of sugar.

1E: The purposes of this experiment was to observe the effects of immersion in solutions of varying tonicity on onion epidermis cells.

Introduction

1A: Diffusion is a kind of passive transport in which molecules move from an areas of high concentration to low concentration. Dialysis is the diffusion of molecules through a selectively permeable membrane, one that allows some substances but not all to pass through. Dialysis tubing is selectively permeable, allowing only substances small enough to fit through its pores to diffuse. Glucose,water, iodine, and potassium iodide are all relatively small molecules, while starch is a macromolecule consisting of many monosaccharides like glucose linked together.

1B: Osmosis is the diffusion of water molecules through a selectively permeable membrane. Isotonic solutions are at dynamic equilibrium, with molecules moving at equal rates in both directions through a membrane for a net change of zero on both sides. Hypertonic solutions have a greater concentration of solute than the opposite side of the membrane, so water rushes into them, while water leaves hypotonic solutions because of their comparatively low solute concentrations; thus, in standard conditions, water will always move from a hypotonic solution to a hypertonic solution until they reach equilibrium and become isotonic.

1C: Water potential  is a measure of the tendency of water to move to or from a place. It is calculated through a combination of the pressure and the solute concentration of the area. Water potential is directly related to pressure potential, but inversely related to solute potential. Addition of solute always decreases water potential so solute potential is always negative. The water potential of pure water at standard atmospheric pressure is zero. Areas of high water potential have high free energy and many water molecules, and water always moves out of areas of high water potential to ones of lower water potential.

1E: Onions are plants and have a rigid cell wall surrounding their cell membranes. When in hypertonic solutions, plant cells experience plasmolysis, in which water diffuses out of the cell and the cytoplasm shrinks. Because the cell wall does not change shape, the cell membrane pulls inwards and becomes detached from the cell wall. In hypotonic solutions, water flows into the cell and is stored in its central vacuole.

Method

1A: For this experiment, we used an approximately 15cm long dialysis tubing; a clear plastic cup; a 15% glucose and 1% starch solution; glucose testing strips; and an iodine potassium-iodide solution. After soaking and opening the dialysis tubing, we tied one end and poured the 15%glucose/1%starch solution into it. The iodine potassium-iodide solution was poured into the clear plastic cup. Both solutions were tested for glucose, and then (after tying up the other end of the dialysis tubing) we submerged the dialysis tubing into the iodine potassium-iodide solution. After 30 mins, each solution was once again tested for the presence of glucose.

1B:This experiment required 6 clear plastic cups; 6 pieces of approx. 15cm long dialysis tubing; distilled water; 0.2, 0.4, 0.6, 0.8, and 1 M sucrose; and a weight. After soaking and opening the pieces of dialysis tubing, one end of each was tied up and each of the previously listed solutions was poured separately into each dialysis tubing. The other ends were then tied up and each “baggie” was weighed and recorded. Meanwhile, each plastic cup was filled with distilled water, and once all the baggies were weighed, they were each placed into a plastic cup for 30mins. Once the time passed, each baggie was taken out of the cups, blotted dry and weighed again.

1C: The materials used in this experiment were as followed: a potato; a cork borer; six clear plastic cups; distilled water; 0.2, 0.4, 0.6, 0.8, and 1 M sucrose; plastic cling wrap; and a weight. Using the cork borer, 24 potato cylinders were cut out (and any pieces of skin removed!) and separated into groups of 4.Each group was weighed and recorded, while the solutions listed above were poured into each plastic cup separately. Once that was taken cared of, each group was placed into a solution filled cup, which were then wrapped closed with the cling wrap and left overnight. The next day, the groups of potato cylinders were taken out, blotted dry, and then each group was weighed and recorded.

1E: This experiment required: an onion; 15% NaCl solution; microscope; distilled water. First, we had to prepare a wet mount of a small piece of the epidermis of the onion which was then observed under 100X magnification. Once that was recorded, we added two drops of the 15% NaCl solution onto the specimen and recorded what we saw. After that we removed the cover slip, put water on the onion, and recorded the result.

Data

1A

 

1B

 

1C

1E

 

 

Graphs and Charts

Discussion

 1A: There was 15% gluecose and 1% starch in the bag. The bag was colorless inside, but then when we put it into the iodine potassium iodine solution and it turned blue.  We know that the iodine went into the bag because iodine reacts with starch and the membrane is permeable to iodine. The bag is not permeable to starch because it did not leave the bag. Before we put the bag into the iodine, there was no gluecose outside, but after being in the iodine potassium iodine, gluecose was found outside the bag. We used the strips that test urine to find if gluecose was present or not. We found that it was, this makes sense because starch is the largest solute, then gluecose, then iodine, and the smallest solute is water. Starch was not able to go across the selectively permeable membrane because it is large, but gluecose, iodine, and water were able to go through. We thought that the membrane was selectively permeable, and we knew that water would perform osmosis through the membrane, we knew larger solutes would be less likely to go through and perform dialysis.

1B: During this experiment we ran into a few technological malfunctions. We had just finished taking the bags out of the distilled water, weighed, and read the masses of the dialysis bags so we disposed of them properly, but our data and mass numbers got deleted FOREVER! We figured out our data eventually. While trying to find the percent change in mass, final mass minus initial mass must be divided by direct initial mass, then multiplied by one hundred.

As we measured the solute concentration of the osmosis of water through a selectively permeable membrane we found that the larger the amount of solute, the larger the percent change. As an example, for(A) Distilled Water, dynamic equilibrium should occur, meaning water goes in and out at the same rate, making this isotonic. The size of the bag should stay the same and ours was 16.3g before and after.  For (B) .2 Molarity of Sucrose, the mass should increase because the water is hypotonic, meaning it has less solute than the inside of the bag. Before we put it in the water it weighed 13.3g and when we weighed it after it weighed 13.8g, consisting of a percent change of 4%. Just as (B) (C-F) were also hypotonic, the percent change ranged from 9%-15%, but the bag of (F) 1 Molarity should expand the most because there is the most sugar inside the bag. The graph shows a direct relationship, as the Molarity of Sucrose increased, the percent change increased. Since life is always trying to reach an equilibrium, the water had to keep going in and spread out.

1C: While measuring the water potential of a potato core during this experiment our data shows that our results had an inconsistency. For Distilled Water and .2 Molarity of Sucrose, our data is acceptable becausethe potato cores in Distilled Water started at 12.6g and increased in mass by 1.6g, making it 14.2g.  The percent change for the potato core in this solution was 13%. The polato cores in solution with .2 Molarity of Sucrose was first 10.2g then increased in mass by .4g, making it 10.6g. There was a percent change of 4% in .2 Molarity of Sucrose. These two were good data because the solutions were hypotonic to the potato, so the water rushed in and made it become heavier. For solutes .4-1 Molarity of Sucrose, the potato core was placed in hypertonic solution where the water potential outside the solution was less than the inside of the potatoes. The water went from high concentration (potato) to low (solution), which caused a change in mass of the potato. At the higher molar concentration water moved out of the potato and moved in at lower molar concentrations. The graph shows an inversely related line because as the Molarity increased, the percent change decreased.

1E: Plasmolysis is the shrinking of the cytoplasm of a plant cell in response to diffusion of water out or the cell and into a hypertonic solution surrounding the cell. The onion showed the difference between how animal and plant cells would react. When the onion was in a hypertonic solution of 15% NaCl, more water went inside the vacuole than the amount that left the cell. The cell wall should begin to expand and became lysed. When in an isotonic solution the amount coming in and out were at equal rates. While in an hypotonic solution since the concentration of solute is less outside the cell wall the water rushes out.

Conclusion

1A: The membrane was selectively permeable and the smaller solutes such as gluecose, iodine, and water were able to diffuse and cross over. Using IKI to show test for starch, the color change showed starch was too big to fit through the membrane because it was still where it was in the beginning.

1B: This experiment proved that water moves across the selectively permeable membrane of the dialysis bag. The sucrose does not move easily through. We can conclude that sucrose must be too large to pass through.

1C: Since water potential equals pressure potential and solute potential, this experiment helped us conclude potatoes had lower water potential because they took in water while in the Distilled Water cup.

1E: From this experiment we were able to understand polymolysis and how the plant cells react in a hypertonic solution. Since the pressure decreased, it became weak when the water left the cell.