20.109(F08):DNA engineering lab report guidelines

September 30 or October 1, 2008

 * First draft is due by 11:00 a.m. on the day you have lab. Please turn in your lab reports electronically by emailing them to bevin, nlerner, nkuldell and astachow AT mit DOT edu. It is important that you name your files according to this convention: Firstinitial_Lastname_LabSection_Mod1.doc, for example: S_Hockfield_TR_Mod1.doc

October 7 or October 8th, 2008

 * Your first draft will be returned. You will have one week from the time your report is returned to address any comments and resubmit your report if you choose. Improvements can increase your grade up to one full letter grade (e.g. a B- on the first draft could become an A-). The grades on the draft and final report are *NOT* averaged.

Guidelines for Module 1 laboratory report
General information about the goal and format for each section of a lab report or research article are found here. Read this general information carefully!

Originality

 * This report must be written by you, and thus should not be written with your lab partner, though we encourage you to discuss your results with eachother.

Guidelines on length
Not counting figures, using 12 pt font, double spaced text and 1” margins (be sure to turn in your final report double spaced), the length should not exceed the following (concise writing is appreciated and rewarded!):


 * Introduction - 2 pages
 * M&M – 2 pages
 * Results – 3 pages
 * Discussion – 1-2 pages
 * Abstract – Under 300 words total
 * Total: Your final report should be ~8-9 pages (certainly not longer than 12 pages).
 * Figures: Your figures should be appended to the end of your report. If the files are large, the figures can be sent together as a file, separate from the report.

Required figures
NOTE: if your own data did not result in the expected outcomes you should include it here but ALSO include the sample data that's been posted to the wiki "talk" pages so you can illustrate your understanding of the results there too. If there are several instances when you have to include both your data and the sample data, then the length recommendations can be ratcheted up a notch...but just one notch.


 * Figure 1: HR figure in the introduction
 * You are welcomed to add figures to the introduction. Hand drawn figures are encouraged (but they would need to be scanned in order to email them as part of your report).  You can use published figures, but if you do so, you must include the reference to the published work. Figures you use should not be downloaded from a web site.
 * The introduction should not include pictures of the plasmids, since that is too much detail for the introduction.
 * Figure 2: Diagram showing what you did. There are two parts to this figure, one general and one more specific. Part A of this figure should offer a high level diagram for how the plasmids can be used to detect homologous recombination in mammalian cells. The reader needs to know what the goal is: which plasmid did you set out to create? Part B of this figure would ideally be a simplified diagram that shows the design strategy for the ligation (e.g., such as the 'Roadmap for Plasmid Construction' that appears on the wiki for Day 1).
 * Figure 3: Your gel showing your PCR results. Keep in mind that all images of gels must include markers indicating the estimated sizes of the observed fragments (these can be estimated by eye based by comparing to the markers). You will certainly need labels indicating the sizes of two or three fragments in the ladder.
 * Figure 4: The gel showing the purified products. See above comments on for Fig. 3  regarding correct labeling.
 * Table 1: Data showing your transformation results, including data for the transformation controls and the calculated transformation efficiency (in the table legend or key if you like).
 * Figure 5: Diagrams showing rough plasmid maps to explain the logic of how you know whether or not you obtained the desired vector. You should indicate the positions of the restriction sites that you used for your analysis, and indicate the sizes that you expect for digestion. Ideally you would also indicate the position of the coding sequences. You will need diagrams of the parental backbone and the correct product.  Please explain to the reader how you know that you do not have multiple inserts (if you indeed know this to be the case).
 * The restriction site positions need not be perfect to the base pair, but should be correct within ~50 bp (e.g., the reader need not know the fragment is 2.543 kb – it is sufficient to state that the fragment is 2.5 kb).
 * Figure 6: The gel showing the results of your restriction analysis. See above comments on for Fig. 3 regarding correct labeling.

Order of assembly
It is recommended that you assemble your lab report in the following order:


 * 1) Figures and tables: Put together the figures and tables, as well as their legends, so you know exactly what you will be talking about.
 * 2) Results: Write the results to describe how and why you did what you did, in a logical way (tell the ‘story’ of what you did). Before you start writing anything, make an outline and think about the logic of the text.  Decide what your subsection headings will be (yes you should have subsections!).
 * 3) *The results should interpret the data. For example, you might write:  “As expected for the correct d5 vector (see map Fig. 1A), Lane 2 of Figure 1B shows that we observed two fragments that were ~x and ~y kb.  Thus, the ligation resulted in construction of the desired plasmid.”  Or you might write: “Unexpectedly, we observed an extra band that was ~3 kb (Fig. 1B, lane 2), which may be the result of a partial digestion.”
 * 4) *Tell the reader what to look at – walk them through your results. For example, for the PCR results, you might write “Figure X shows the results of our PCR.  Results for the No DNA control are shown in lane 2, with one small bright band of primers but no longer products."
 * 5) Discussion: After you write your results, you will have thought about the discussion and you’ll be ready to write it. The discussion is where you accomplish two major objectives: 1) talk about results that weren’t as expected, and come up with ideas about what could have happened.  Try to avoid simplistic explanations (e.g., the enzyme was not added), unless you have very good reason to think that you did.  2) Put the results into the context of the bigger picture.  What would be the next useful thing to try with this plasmid you've worked so hard to make? How sure are you about what it is and how it will work? How generalizable or reliable do you think these molecular approaches are?
 * 6) Introduction: Write the introduction. Knowing the whole story, you now can ask yourself ‘what does the reader need to know to understand what I’ve written?’.
 * 7) *Your introduction should address the following questions: What is homologous recombination?  Why should the reader care about this repair pathway? What is the objective of the experiments? What is the value of the plasmid in an assay you are helping to develop?
 * 8) *Generally, people usually end the introduction with a paragraph summarizing in very brief terms the major results. For example, “Here, we have created a system for measuring xxxx, and we have compared xxxx.  Our results show xxxx, indicating that xxxx.”  You will certainly want to tell the reader what you set out to do - the extent to which you disclose results here is a matter of style, so we will leave this choice up to you.
 * 9) Abstract: Write the abstract last. It’s easiest to write the abstract just after writing the main text, when everything is fresh in your mind, and you have a good sense of how confidently you can make your statements.
 * 10) Materials and Methods: This can be written at any time.  It should be brief!
 * 11) *Assume the reader can refer to the NEB catalog for temperatures and amounts of enzyme to add to cut DNA. They just need to know what enzyme was used. Similarly you can assume folks can perform Qiagen columns as directed, and that they know how to load gels.
 * 12) *Regarding PCR, they need to know what the key features of the primers were. The sequence itself is not necessary and is generally not meaningful to the reader.. ideally you would state something like: "The forward primer anneals to the first 22 nucleotides, beginning at the ATG. To the 5’ end of the primer was added the X and Y restriction sites.  An additional X nucleotides were added to the X end in order to assure that ... Sequence is available upon request.”