Cellular Biology Lab – Homework #3
Due the week of Nov. 4th
You may use the lab manual, pre-lab lectures, and credible internet resources, however you may not use your cell bio lab classmates as a resource. You will most likely see this material again on the Final and I highly encourage you to work individually and seek help from myself or your TA. Plagiarism will result in an automatic zero.
1. In the cell bio lab, we use company manufactured gels, however you can make you own polyacrylamide gels. List all of the ingredients found in an SDS-PAGE gel. Which ingredients are responsible for polymerizing the solution? How does the percentage of acrylamide effect the migration of proteins (ex: 4% gel vs. 18% gel)?
The percent
…show more content…
In this gel above, which lanes contain the positive and negative controls? Describe the information you can deduce by comparing and contrasting the negative control and lane 2 protein bands. Lane 3 is the positive control and lane 4 is the negative control. By comparing lane 4 the negative control and lane 2 the purified protein X it is noticeable that the second band in lane 2 is the elution buffer due to the fact that the negative control only contains elution buffer and only forms one band which matches up with the second band in lane 2.
c. Describe the information you can deduce by comparing and contrasting the positive control lane and lane 2 protein bands. By comparing lane 3 the positive control to lane 2 the purified protein X it is noticeable that the first band is the unknown protein X due to the fact that the positive control only contains purified protein X and only forms one band which matches up with the first band in lane 2.
d. You don’t know the molecular weight (MW) of protein X and you are not able to find that information in the scientific literature. The best way to determine the MW of a protein using an SDS-PAGE gel is to use the protein ladder bands to create a Log(MW) vs. Rf graph and calculate the MW from the line of best fit. What is the equation to calculate the Rf of a protein band? Make a table of the Log(MW) and the Rf values for all 5 protein ladder bands. Describe any trends you see in the table
Please answer these questions then place them in the drop box for this lab. Use Microsoft word if possible.
In the experiment we used Turnip, Hydrogen Peroxide, Distilled Water, and Guaiacol as my substances. On the first activity, Effect of Enzyme concentration of Reaction Rate for low enzyme concentration, we tested three concentrations of the turnip extract, and hydrogen peroxide. For the Turnip Extract I used 0.5 ml, 1.0 ml, and 2.0 ml. For hydrogen peroxide we used 0.1 ml, 0.2 ml, and 0.4 ml. We used a control to see the standard, and used a control for each enzyme concentration used. The control contains turnip extract and the color reagent, Guaiacol. We prepared my substrate tubes separately from the enzyme tubes. My substrate tube
2. The larger value obtained when more glucose carriers were present corresponds to an increase in the rate of glucose transport. Explain why the rate increased. How well did the results compare with your prediction?
Colorimetric assay is a process of determining a concentration of a solution based on absorbance of light. The purpose of this lab is to determine if the Bradford assay is an accurate way to determine an unknown concentration of two samples of protein. The Bradford assay is done by measuring wavelength of light passing through a cuvette filled with Bradford dye and concentrations of PBS and proteins. After the cuvettes are mixed they are placed into a spectrophotometer to measure wavelength. The wavelength given will be used to plot a standard curve based on concentration (x-axis) and wavelength (y-axis). The standard curve is then used to measure an educated guess on the concentrations of unknown protein concentrations. We hypothesized that if we use the Bradford assay and colorimetric spectrophotometry we can determine an accurate concentration of two unknown concentrations of proteins. The results of this lab failed to reject our hypothesis based on accurate measurements of protein concentrations. The standard curves are drawn with a linear increasing slope. The Bradford assay is an accurate way to demine the concentration of an unknown concentration.
4. Describe the test used to detect the presence of each type of biologically important
5. What reaction would you expect when performing a negative control in the catalase assay?
This technique separates Rubisco samples based on their size. The electrophoresis has a positive and a negative end. Positive charge proteins are loaded from the positive end and migrate towards the negative end. Negative charge proteins are loaded from the negative end and migrate towards the positive end (Sakthivel & Palani, 2016). The sample that contained the highest molecular weight of Rubisco will travel the shortest distance on the gel while the protein with the smallest molecular weight will travel the longest distance (Sakthivel & Palani, 2016). The size proportion of each Rubisco molecule correlates with the distance traveled. Rubisco will be in its purest form after running through SDS-page since each technique will increase the purity of the protein. If the salting out, the ion exchange and the SDS-page protein isolation techniques are performed on protein Rubisco, then it is purified and separated by solubility, charge, and size. The rationale of this experiment is to isolate the purest form of Rubisco so that it can perform carbon fixation at an optimal
3. You test another new unknown bacterial sample, and find the G+C content is identical to one of the samples you have already identified, but the rRNA gene sequence contains one base that is different. What can you conclude: C.
Read in your lab manual about the following agar mediums: Blood Agar (pg 168), EMB Agar (pg 170), Mannitol Salt Agar (MSA)(pg 172) ), MacConkey Agar (pg 174), and PEA Agar (pg 176) to answer the following:
This document is not meant to be a substitute for a formal laboratory report. The Lab Report Assistant is simply a summary of the experiment’s questions, diagrams if needed, and data tables that should be addressed in a formal lab report. The intent is to facilitate students’ writing of lab reports by providing this information in an editable file which can be sent to an instructor.
Figure 1 contains gel electrophoresis for protein samples. The lanes were labeled from 1 to 10 from the right to the left. Lane 1 contained the ladder fragment. Lane 2 contained the filtrate. Lane 3 contained the S1 sample. Lane 4 contained the P1 sample. Lane 5 contained the P1 medium salt sample. Lane 6 contained the P1 high salt sample. Lane 7 contained the S2 sample. Lane 8 contained the P2 sample. Lane 9 contained the P2 medium salt sample. Lane 10 contained the P2 high salt sample.
This document is not meant to be a substitute for a formal laboratory report. The Lab Report Assistant is simply a summary of the experiment’s questions, diagrams if needed, and data tables that should be addressed in a formal lab report. The intent is to facilitate students’ writing of lab reports by providing this information in an editable file which can be sent to an instructor.
Gel electrophoresis method. Qualitative analysis shows protein concentrations in kidney, heart, and liver. 1-6 are kidney tissue. 7-14 are liver and 15-19 are the heart tissue.
Load the remaining protein mix into the column, and this will begin to bind within the column. Once the solution has settled, fill the column with NaCl a total of four times; as the wash is flowing through, collect and label the protein solution in fractions, and save for the
Name ____________________________ I) Introduction All cells contain four major types of macromolecules: carbohydrates, lipids, nucleic acids, and proteins. In today’s lab, we will be studying three of the four-proteins, carbohydrates and lipids. Various chemical tests can be used to detect the presence of each of these molecules. Most of the tests involve a color change visible to the eye. If a color change is observed, the test is considered positive. If the color change is not observed, the test is negative, indicating that a particular molecule is not present. In all the chemical tests we will be performing, we will also be using a control. In most cases, the control will be a sample of