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The “guidelines” tells you how to write and what questions should you answe in the paper.
The “lab 2″paper is a background about what is the experiment about,
The “procedure” is how we did the experiment step by step and the data that i took.
The ” Graphing page” is where you have to graph as what the instructor mentions on the guide line. you can use Excel foe graphing or any other thing but you have to add the graph.
Lamia/Final data ( results) x
1- label all lanes, label marker sizes, and indicate which three lanes, containing at least one BSA sample and one
E. coli sample, you are writing about.
2- lanes 2, 5, 6, 9, and 11 are BSA, lanes 14 and 15 are empty, and lanes 3, 4, 7, 8, 10, 12, and 13 are
E. coli.
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Lamia/Graphing page
Lamia/Guidelines
Biology 105 Laboratory Fall 2013
Instructor: Ayça Akal-Strader
Guidelines for Lab Report
Lab 2: Quantification of Protein (Bradford Assay)
Your report for Lab 2: Quantification of Protein (Bradford Assay) is
due the week of October 7/8/9/10. Please include the following information in your report:
Hypothesis: as usual
Introduction:
• Background/theory of Bradford Assay
• Purpose of the experiment
Results:
In addition to the specific data discussed below, your
Results section should always include one or more paragraphs of text that provide:
• A brief description of the procedure
• Explanations of any charts, graphs, figures, or calculations that are included
• Statements about the most interesting/noteworthy data
Data:
1. Table of measured absorbances (like Table 2 on p. 31).
2. Table showing protein concentrations of unknowns (like Table 3 on p. 31). Say which unknowns—1, 2, or both—you used.
**Please re-make the tables for your report. DO NOT simply tear out p. 31 from your lab manual and staple it to your report.
3. Standard Curve:
• Label with title and caption
• Label axes: x-axis = Concentration (μg/ml); y-axis = Absorbance at 595 nm. Be sure to include units on Concentration. Remember that absorbance (optical density; OD) has no units.
• Plot points, leaving room to plug in your unknown absorbances to find their concentrations
• Connect the dots
(Note: Do NOT draw a straight line—unless your data really looks like a straight line. The samples we measured did not fall into the “linear range” of the spectrophotometer, and everyone’s data that I saw flattened out a lot at the high concentration end of the range. Connect your data points with a curve.)
• Indicate by drawing horizontal and vertical lines how you found the concentration of your unknowns.
Discussion:
• Did your results match your expectations? If not, why not?
• Did you have any difficulty finding the concentration of any of your unknowns?
• Do you think your measurement of protein concentration was accurate? Did your duplicates agree well? For your standards, did your absorbances increase as your protein concentrations increased?
Conclusion: as usual
Lab Report Rewrites
You may rewrite
TWO of your
first FIVE lab reports in an effort to improve your grade.
You do not need to rewrite the entire report; just fix the problems that caused you to lose points the first time around.
You MUST hand in the original version of your report along with your corrected version. If you do not have the original attached, we will not accept your rewrite.
Your final grade on the rewritten report will be the average of your original grade and the new grade. (This can only help you. If you somehow manage to get a lower grade on the rewrite, we will keep your original grade as is.)
Rewrites are
due the week of November 18/19/20/21, which is also the week of the Wrap Up and Review & the Lab Exam.
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Lamia/Lab 2
Lamia/Lab report guidelines (1)
Biology 105 Laboratory Fall 2013
Instructor: Ayça Akal-Strader
Guidelines for Lab Report
Lab 1: Introduction to Biomolecules – A Look at Proteins Using SDS-PAGE
Your report for Lab 1: A Look at Proteins Using SDS-PAGE is
due the week of
September 30 & October 1/2/3. Please include the following information in your report:
Introduction:
• Background/theory of SDS-PAGE
• Purpose of the experiment
• Expected results
Results:
Note: In addition to the specific data discussed below, your
Results section should always include one or more paragraphs of text that provide:
• A brief description of the procedure
• Explanations of any charts, graphs, figures, or calculations that are included
• Statements about the most interesting/noteworthy data
Gel Picture:
• Include your gel picture with enough white space around it to label cleanly. Affix it to a larger sheet of blank paper, if necessary.
• Number the lanes
• Give the picture a title and a caption. The caption should briefly define what is in each lane.
• Using the key provided, label your molecular weight standards.
• In one of your BSA lanes, circle the BSA band and label it with its approximate size. This information should also be included in the written part of your Results section.
• In one of your
E. coli lanes, circle the largest and smallest bands you can see clearly, and label them with their approximate sizes. Choose another band you can see clearly, circle it and label it with its approximate size. This information should also be included in the written part of your Results section.
• Estimate the number of bands you see in one of your
E. coli lanes. This information does not need to be written on your gel picture; you should include it in the written part of your Results section.
**Note: If you cannot clearly see your own results due to technical problems with the gel, you may analyze another group’s results that are shown on the same gel. If you do this, say so!
Discussion:
• Did your results match your expectations? If not, why not?
• How many different proteins do you think are present in an
E. coli? Did you see that many protein bands in your
E. coli lane? If not, why not?
• Why are some of your
E. coli bands thicker/more intense than others?
Lab Report Rewrites
You may rewrite
TWO of your
first FIVE lab reports in an effort to improve your grade.
You do not need to rewrite the entire report; just fix the problems that caused you to lose points the first time around.
You MUST hand in the original version of your report along with your corrected version. If you do not have the original attached, we will not accept your rewrite.
Your final grade on the rewritten report will be the average of your original grade and the new grade. (This can only help you. If you somehow manage to get a lower grade on the rewrite, we will keep your original grade as is.)
Rewrites are
due the week of November 18/19/20/21, which is also the week of the Wrap Up and Review & the Lab Exam.
Page
2 of
2
Lamia/lab_report_complete_photo
Biology 105 Laboratory Fall 2013
Instructor: Ayça Akal-Strader
Guidelines for Lab Report
Lab 1: Introduction to Biomolecules – A Look at Proteins Using SDS-PAGE
Introduction:
Background/theory of SDS-PAGE
A major goal of biochemistry is to study the cellular processes of living organisms and how these processes relate to the functioning of the organism. Research in the area of biochemistry has been extremely successful over the last century; we now know the atoms and biomolecules that make up living organisms, the central dogma around which biological information is transferred and how this information results in a greater understanding of ourselves and the world in which we live. One of the most important results from research in biochemistry is that all organisms are uniform at the molecular level and the diversity we see today is a result of evolution.
A procedure for extracting plasmid DNA from bacterial cells is described. The method is simple enough to permit the analysis by gel electrophoresis of 100 or more clones per day yet yields plasmid DNA which is pure enough to be digestible by restriction enzymes. The principle of the method is selective alkaline denaturation of high molecular weight chromosomal DNA while covalently closed circular DNA remains double-stranded. Adequate pH control is accomplished without using a pH meter. Upon neutralization, chromosomal DNA renatures to form an insoluble clot, leaving plasmid DNA in the supernatant. Large and small plasmid DNAs have been extracted by this method.
By heterologous expression of the pore-forming transmembrane-protein (or porine) OprF from Pseudomona fluorescens a further optimization of the E. coli membrane will be achieved. E. coli possesses several own porines, for example OmpF and OmpC. But these naturally occurring porines are only permeable for molecules smaller than 600 Da, which decreases the range of usable mediators significantly. Opposed to that, porine OprF from P. fluorescence forms one of the biggest known pores in the outer bacterial membrane with a permeability of up to 3000 Da, thus improving the electron shuttle mediated extracellular electron transfer (ETT) and enabling the usage of alternative mediators such as riboflavin (vitamine B2).
Results
The SDS-PAGE shows a significantly higher protein concentration for E.coli with OprF and T7 promoter. It seems to be that the higher membrane permeability (shown with NPN and ONPG uptake assay) allows a better release of membrane proteins by 0.2 % SDS. Nevertheless, we can see a strong over-expression band at the expected OprF size of about 36 kDa for, which is equated with a strong expression and overproduction of OprF. Furthermore we were able to identify the overexpressed outer membrane porin (Figure. 2) with MALDI-TOF MS/MS. Tryptic digest of the gel lane for analysis with MALDI-TOF could examine the outer membrane porin with a Mascot Score of 222 against bacteria database.
The heterologous expression of the outer membrane porin OprF will enhance the hydrophobicity of cell membrane. The outward-facing side groups on each of the β-strands of the OprF monomer are hydrophobic. Therefore a positive expression should be visible by an increase in hydrophobicity. An increasing hydrophobicity of cell membrane changes the physicochemical properties of the cell. This could effect for example the cell-electrode interaction.
Discussion
In summary we can say that the heterologous expression of OprF from Pseudomonas fluorescens in Escherichia coli significantly improves the membrane permeability. NPN and ONPG assay show correlating results. Escherichia coli outer membrane permeability is enhanced with increasing promotor strength for OprF expression. The Wildtyp shows only a weak uptake rate of chemical molecules (NPN) but no product secretion as quantified with ONPG assay. Therefore, Wildtyp is not suitable for a usage in the MFC. Whereas membrane optimized Escherichia coli with OprF shows great diffusion processes in and out of the cellular membrane, indicating a great optimization of electron shuttle-mediated electron transfer (EET) to the anode and increasing current production.
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Lamia/Procedure .crdownload
