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Cao, Jasmin

BIOL 151
Professor Denmon/Vetrone
11/10/2017
Lab Questions:

Experiment 1: Observe Stimulated Diffusion

1. Hypothesis: Solute size plays a large role in the effectiveness of diffusion


a. Prediction A: If the solute size is bigger outside the cell, it will not move into the cell
b. Prediction B: If the solute size is smaller outside the cell, it will move into the cell
2. A color change from clear to dark pink/light purple was observed in Bag #1. A color change from
clear to blue was observed in Bag #2.
3. For Bag #1, the color change indicates the passage of OH- through the semipermeable membrane into
the cell. For Bag #2, the color change indicates the passage of iodine through the semipermeable
membrane into the cell.
4. For molecules to be able to pass the semipermeable membrane, they usually have to be small and
uncharged. Larger and charged molecules, on the other hand, will have a harder time passing through
the semipermeable membrane.

Experiment 2: Observe Stimulated Osmosis

5. Hypothesis: Water moves from an area of lower solute concentration to an area of higher solute
concentration
a. Prediction A: If there is only 1% concentration of sucrose inside the cell, minimal water
molecules will move into the cell
b. Prediction B: If there is only 1% concentration of sucrose inside the cell, minimal water
molecules will move into the cell
c. Prediction C: If there is 10% concentration of sucrose inside the cell, an intermediate amount of
water molecules will move into the cell
d. Prediction D: If there is 25% concentration of sucrose inside the cell, a large amount of water
molecules will move into the cell at a very fast rate
6. The movement of water will be the greatest in Bag D because it contains the greatest amount of
solutes or sucrose. A larger amount of solutes in Bag D will cause a large amount of water molecules
to move into the cell at a very fast rate. This is because water molecules move from an area of lower
solute concentration to an area of higher solute concentration
7. The movement of water will be the least in the Bags A and B because they contain the least amount of
solutes or sucrose. A minimal amount of solutes in Bags A and B will cause a minimal amount of
water molecules to move into the cell at a very slow rate. This is because there isn't much of a
difference between the solute concentration inside and outside of the cell.
8. How fast the water will move through each bag:
a. Bags A and B: Extremely slow ~ 1 min
b. Bag C: Intermediate speed ~ 30 seconds
c. Bag D: Extremely fast ~ 15 seconds
9. The change in weight between each recorded time for Bag C:
a. Final Weight (3.3g) - Initial Weight (4.3g) = 1.0g
10. Table 1: Weight of Bag C over a period of 60 minutes
Weight of Bag C (grams)

0 min 3.3 g

15 min 3.7 g

30 min 3.9 g

45 min 4.1 g

60 min 4.3 g

11. The independent variable of the experiment is the amount of solutes in each bag. This is because the
independent variable is the variable that is manipulated. The dependent variable is the amount of
water that moved into the cell, measured by recording the initial and final weights of the cell. This is
because the dependent variable is the variable that is measured.
12. Figure: Observing stimulated diffusion in Bag C. The weight of bag C over a period of 60 minutes
was measured every 15 minutes was measured. Data shows an increase in weight every 15 minutes
until it reached its heights at 60 minutes. The increase in weight indicate the movement of water into
bag C as a result of the process of osmosis.

Weight of Bag C Overtime


Total Weight vs. Time (Bag C)
4.4
Weight of Bag C in grams

3.3

2.2

1.1

0
0 15 30 45 60
Time

13. The rate of diffusion increases as the solute concentration increased.


14. The curves for bags C and D eventually became horizontal when osmosis stopped occurring or when
water molecules stopped moving into the cells. In other words, the solute concentrations inside and
outside the cell became equal and the cell and its environment reached equilibrium.
15. Osmosis occurred in all bags because as long as there is a difference between the solute concentration
inside and outside the cell, there will be movement of water molecules and thus osmosis. In other
words, the concentration gradient was present between all bags and their environments
16. Bags A and B had the lowest concentration gradient because there was not much difference in solute
concentration between inside and outside the cell. Bag D had the highest concentration gradient
because it had the largest difference in solute concentration between inside the outside the cell that
can drive the movement of water molecules across the semipermeable membrane. Bag C had a higher
concentration gradient than Bags A and B but a lower concentration gradient than Bag D because it
had an intermediate difference in solution concentration between inside and outside the cell compared
to the other bags.
17. The greatest changes in weight occurred in bags with the largest concentration gradient

Experiment 3: Effect of Temperature on Membranes

20. Figure: Effect of various temperatures on cellular membranes. The degree of color intensity,
which indicate the degradation of the cellular membrane, was measured on a scale of 0-10. (0
indicates no color, 10 indicates dark color). Data shows an increase in color intensity in the highest
and lowest temperatures such as 70C and -20C. In other words, the degradation of the cellular
membrane was extremely high in extreme temperatures.

Group Class Avg.


Temperature vs. Relative Color
9
Color Intensity (Scale 0-10)

6.75

4.5

2.25

0
70C 55C 37C 22C 4C -20C
Temperature in C (Tubes 1-7)

21. Figure: Effect of various temperatures on absorbance. The degree of absorbance, which indicate
the degradation of the cellular membrane, was measured on a scale of 0-10. (0 indicates no
absorbance, 10 indicates highest absorbance). Data shows an increase in absorbance in the lowest
temperature or -20C. In other words, the degradation of the cellular membrane was extremely high
in the lowest temperature.
Group Class Avg.
Temperature vs. Absorbance
1.6

1.2
Absorbance 460nm

0.8

0.4

0
70C 55C 37C 22C 4C -20C
Temperature in C (Tubes 1-7)

22. In this experiment, the temperature and time are the independent variables because they are being
manipulated. The dependent variables are absorbance and color intensity because they are being
measured.
23. -20C damaged the membranes the most while more optimal temperatures such as 4C and 55C
damaged the membranes the least. This is because the absorbance and color indicate the degree of
damage to the cellular membrane in which the lowest temperature -20C displayed the highest
absorbance and color intensity and 4C and 55C displayed relatively the least absorbance and color
intensity. The extreme temperatures such as heat and cold can damage the cellular membrane in
numerous ways. For example, high temperatures can destroy a cellular membrane by causing violent
molecular collisions. Extremely low temperatures, on the other hand, can cause water molecules to
expand and transform into ice. The expansion of water and formation of solid ice can possibly rupture
and destroy cellular membranes.
24. The spectrophotometer was used to confirm what we observed by measuring the concentration of
substances in the sample tubes.
25. For the cooling and freezing temperature, the incubation times were extended by 10 minutes because
the tubes and its contents must adjust to the new temperatures from room temperature.

Experiment 4: Effect of Organic Solvent Stress on Membrane

26. Figure: Effect of organic solvents on color intensity. The degree of color intensity, which indicate
the degradation of the cellular membrane, was measured on a scale of 0-10. (0 indicates no color, 10
indicates darkest color). Data shows an increase in color intensity in the highest
Group Class Avg.
Concentration of Organic Solvent vs. Relative Color
10
Color Intensity (Scale 0-10)

0
1% acetone 25% acetone 50% acetone 1% methanol 25% methanol 50% methanol isotonic saline
Treatment (Tubes 1-7)

27. Figure: Effect of organic solvents on absorbance. The degree of absorbance, which indicate the
degradation of the cellular membrane, was measured on a scale of 0-10. (0 indicates no absorbance,
10 indicates highest absorbance). Data shows the greatest absorbance in the highest concentrations of
organic solvents such as 50% methanol and 50% acetone. In other words, the degradation of the
cellular membrane was extremely high in larger organic solvent concentrations.

Group Class Avg.


Concentration of Organic Solvent vs. Absorbance
0.7

0.6
Absorbance 460nm

0.5

0.4

0.3

0.2

0.1

0
1% acetone 25% acetone 50% acetone 1% methanol 25% methanol 50% methanol isotonic saline
Treatment (Tubes 1-7)
28. The dependent variables are the absorbance and color intensities because they are being measured.
The independent variables are the concentration of each organic solvent and the types of organic
solvents used because they are being manipulated.
29. Based on the data, lipids are soluble in both methanol and acetone. This is indicated by the high color
intensity and absorbance values in both acetone and methanol solutions.
30. 50% acetone damages membranes more than 25% acetone. This is because 50% acetone displayed a
higher absorbance value and color intensity than 25% acetone in which a higher absorbance and color
intensity indicates a large degree of damage and harm to the cellular membrane of beets.
31. Acetone solvent solution, more specifically 50% acetone, caused the most harm to lipids. This is
because acetone displayed the highest absorbance values and color intensity compared to those of
methanol, indicating the large emission of color and thus a high degree of harm to the cellular
membrane.
32. Yes, based on the data, the highest concentration of both solvents caused the more damage. For
example, 50% acetone displayed the highest absorbance values and color intensity compared to lesser
concentrations of acetone such as 25% and 1% acetone. Similarly, 50% methanol displayed the
highest absorbance values and color intensity compared to lesser concentrations of methanol such as
25% and 1% methanol.
33. Tube 7 is an isotonic saline solution that did not contain any organic solvents. The purpose of this
tube was to utilize its results and compare them with the results of the various other tubes that did
contain concentrations of organic solvents. According to the data, the tubes that contained organic
solvents had higher absorbance values and color intensity than those of the isotonic saline solution,
indicating the power of organic solvents in degrading cellular membranes.
34. No, my conclusions about membrane structure and stress is valid to most other cell membranes. This
is because beet cell membranes and the membranes of other plants and animals are similar in
structure and function. Therefore, we can make a valid assumption and prediction that if organic
solvents degrade beets membranes, they will also have the ability to degrade many other cell
membranes of similar structure and function.
35. In freezing temperatures, the cellular membranes in the food we preserve will be degraded or
damaged. However, the bacteria or other organisms that may cause the food to become spoiled may
be degraded as well in extreme temperatures due to the ability of temperatures to disintegrate cellular
membranes. Thus, food can be safely preserved by freezing it.
Results

In this lab activity, the effect of organic solvent stress on membranes were determined by
the addition of organic solvents such as acetone and methanol on cell membranes and observing
their color intensity as well as absorbance levels. There was a strong increase in the color
intensity as the concentration of the organic solvents increased. Similarly, there was an evident
increase in the absorbance as the concentration of organic solvents increased as well. For
example, 50% acetone displayed the highest absorbance values and color intensity compared to
lesser concentrations of acetone such as 25% and 1% acetone. Similarly, 50% methanol
displayed the highest absorbance values and color intensity compared to lesser concentrations of
methanol such as 25% and 1% methanol. In addition, acetone displayed the highest absorbance
values and color intensity compared to those of methanol. These observations were found in both
the group and class average datas.
Discussion/Conclusion

In this lab experiment, the effect of organic solvent on cellular membranes was observed.
Beets were used for this experiment. Each beet cylinder was placed into one of the seven test
tubes, each tube varying in the type and concentration of organic solvents. After all the beets
were removed, the color intensity of the solutions in all the tubes were observed and recorded on
a scale of 0 to 10, with 0 being colorless and 10 being the darkest shade of red. Finally, the
absorbance of each test tube was read at 460nm.
The data obtained highlight a strong increase in the color intensity as the concentration of
the organic solvents increased. Similarly, there was an evident increase in the absorbance as the
concentration of organic solvents increased as well. For example, 50% acetone displayed the
highest absorbance values and color intensity compared to lesser concentrations of acetone such
as 25% and 1% acetone. From these datas, we can determine that an increase in the concentration
of organic solvents caused greater harm to the cellular membrane. Because of the beets red
pigments in its cellular membrane, the disintegration of its cellular membrane will cause the
release of its pigments into its environment or solution. Therefore, color intensity and absorbance
levels indicate the degradation of the beets cellular membrane. In addition, acetone displayed the
highest absorbance values and color intensity compared to those of methanol. These observations
were found in both the group and class average datas. In other words, acetone was evidenced to
have a greater ability to destroy cellular membranes than methanol. Furthermore, Tube 7 is an
isotonic saline solution that did not contain any organic solvents. The purpose of this tube was to
utilize its results and compare them with the results of the various other tubes that did contain
concentrations of organic solvents. According to the data, the tubes that contained organic
solvents had higher absorbance values and color intensity than those of the isotonic saline
solution. Therefore, organic solvents have the ability to disintegrate cellular membranes of
organisms.
Organic solvents such as acetone and methanol can cause stress on membranes by
dissolving lipids. More specifically, organic solvents consisting of hydrocarbons are able to
accumulate in the membrane lipid bilayer are able to negatively influence the structural and
functional properties of cellular membranes. Accumulated hydrocarbon molecules can cause the
membrane to lose its integrity as well as an increase in permeability to protons and ions,
highlighting the destructive and toxic effects of organic solvents on cellular membranes. A
numerous number of chemical and biological processes are obstructed and hindered by the
harmful and destructive effects of organic solvents on cells. This destructive and negative effect
of organic solvent on cells were evidenced in this experiment as the color intensity and
absorbance values of beets increased with the concentration of organic solvents.
Gel Electrophoresis Lab
Results

This lab experiment was performed to visualize DNA fragments from PCR and restriction
enzyme digest. Our results from gel electrophoresis can be seen in the picture above. The first
well contained the DNA ladder. The second well, which contained the MreB master mix and the
displayed no bands. The third well contained MreB master mix and strawberry DNA. The fourth
well contained MreB master mix and bacteria DNA. The fifth well, which contained MreB
Master Mix and A+ solution, displayed an extremely faint band. The sixth well contained MreB
master mix and the M+ solution. The seventh well contained B-Actin master mix. The eighth
well contained B-Actin master mix and strawberry DNA. The ninth well contained B-Actin
master mix and bacteria DNA.The tenth well, which contained B-Actin Master Mix and A+
solution, displayed no bands. The eleventh well, which contained B-Actin master mix and the M
+ solution, also displayed no bands. Furthermore, many primer-dimers were detected in bands
3,4,8, and 9. Primers dimers are unwanted extension products that result from primers annealing
to themselves or each other at 3 ends. In addition, bands 3,4,6,7, and 8 were displayed below
0.5kb on the DNA ladder.
Discussion/Conclusion

This lab experiment was performed to visualize DNA fragments from PCR and restriction
enzyme digest by gel electrophoresis. Gel electrophoresis is an analytical technique used to
separate DNA and RNA fragments or proteins by size and charge. During this experiment, the
agarose gel solution was created and poured into the gel tray. The gel solidified and was left to
cool in the gel tray after inserting the plastic comb into the gel. As the gel was solidifying, we
prepared our samples by adding dye to each tube. The gel was then reoriented so that the wells
were towards the negative pole. TAE buffer was added to the tank and the comb was pulled out
gently. In the first well, the DNA ladder was added. Finally, our samples were also added to the
remaining wells of the gel. The gel was next left to run at 100V for around 40 minutes.
Afterwards, the gel was visualized with a bioimager. Because the DNA ladder provided did not
include kilo bases under 0.5kb and was outside the range of accuracy, we were not able to
determine the kilo bases of some bans. As a result, we were not able to determine that the
product that was produced by the beta-actin primers is in fact the correct size. The second well,
which contained the MreB master mix and the displayed no bands.This is accurate because the
second well did not contain any DNA and thus, should not display any bands. Also, the tenth
well, which contained B-Actin Master Mix and A+ solution, displayed no bands. The A+ should
have been amplified by B-Actin and produced bands. Similarly, the eleventh well, which
contained B-Actin master mix and the M+ solution, also displayed no bands. This is odd because
these two samples contained DNA and should have produced clear bands. The error in the results
may be due to contamination or possible loss of sample somehow. The seventh well contained B-
Actin master mix,which means the sample did not contain any DNA. However, the seventh well
displayed a clear band. These results, just like the tenth and eleventh wells, may be caused by
continuation or an error when performing the procedures. The fifth well, which contained MreB
Master Mix and A+ solution, displayed an extremely faint band. This is accurate because A+
solution can only be amplified by B-Actin. Furthermore, many primer-dimers were detected in
bands 3,4,8, and 9. Primers dimers are unwanted extension products that result from primers
annealing to themselves or each other at 3 ends. In addition, bands 3,4,6,7, and 8 were displayed
below 0.5kb on the DNA ladder which means we will not be able to interpret the data since it is
beyond the range of the DNA ladder.

Gel electrophoresis is a technique used to separate DNA fragments according to their size
and charge. Electrophoresis involves running a current through a gel containing the molecules of
interest. Based on their size and charge, the molecules will travel through the gel in different
directions or at different speeds, allowing them to be separated from one another. As a result, gel
electrophoresis can be used to separate DNA fragments to get a DNA fingerprint for forensic
purposes, paternity testing, checking PCR reactions and to look for evolutionary relationships
among organisms. The process is also used to test for and locate genes associated with a specific
disease and to distinguish between samples of genetic material.