20.109(F15): DNA engineering summary

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20.109(F15): Laboratory Fundamentals of Biological Engineering

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Contents

Overview

The culminating assignment for Module 1 will consist of two elements: an abstract that succinctly describes your DNA engineering investigation, and a thorough summary of your data in figures and supporting text – including context for understanding the work’s broader implications.

The figure format is similar to that of a scientific journal article, but the traditional Results and Discussion sections found in journal articles are to be condensed into succinct bullet point form accompanying each figure. The purpose of this assignment is to prepare you to write a full journal article at the end of Module 2 by encouraging you to practice writing concisely using bullet points.

The target audience for this report is a scientifically literate reader who is unfamiliar with your specific field. Thus, you can assume rapid comprehension – but not a priori knowledge – of technical information, and consequently should strive to present your work in a logical, step-by-step fashion.

Logistics

You will complete this assignment with your partner. Be sure to review the 20.109 statement on collaboration and integrity as you proceed.

Method of submission

Please submit your completed summary on Stellar, with filename Names_TeamColor_LabSection_Mod1.ppt (for example, EngelwardBelcher_Rainbow_TR_Mod1.ppt).

Be sure to review the class late policy (link) as well as the further clarification below.

Draft submission: Oct 13th

Your DNA engineering summary is due by 5 pm on Tuesday, Oct 13th.

Dr. Engelward will comment on your submission and assign it a grade. The BE Communication Instructor, Dr. Siegel, will provide feedback about abstract structure and comprehensibility -- she will also assign the grade for the Abstract portion of the assignment. You will receive all comments by Monday, Oct 19th.

Revision submission: Oct 24th

Your DNA engineering summary revision is due by 5 pm on Saturday, Oct 24th.

You will then have the opportunity to revise your report for up to a one and one-third letter grade improvement. In other words, a C can be revised up to a B+, a C+ to an A-, a B- to an A, etc)

For your final report, all changes need to be in a different colored font so the improvements you made are clear. You should also include a cover letter with your final draft that explains how you addressed the concerns raised (e.g. "paragraph x was completely rewritten to better explain….” or “Results for the agarose gel analysis were clarified by …."). You will receive additional comments and your final grade on the assignment by Friday, Nov 6th.

Guidelines on formatting and length

You will prepare your report as a series of PowerPoint slides, using a portrait rather than landscape layout. This approach will allow you to create figures that are representative of those found in the literature (i.e. sized appropriately and may contain several sub-panels). The text should be embedded in the slides no smaller than Arial 14 pt font (12 pt font for figure captions). In addition, the text should be in included as bullet points rather than full sentences and paragraphs.

Please see this example for an appropriate format.

Core document length should be about 10 pages, and certainly not exceed 12 pages. Though somewhat variable, typical section lengths (including both text and figures) might be:

  • Background and motivation: ~2 slides
  • Results and interpretation: ~5-8 slides
  • Implications and future work: ~1-2 slides

The first page of the document should include an informative title and author information (section/color/names); the second page should include just an abstract. These two pages will not count toward the suggested 10-12 pages. A typical introductory or conclusion/summary page will be enhanced by a supporting figure, though some might include only text. A typical data page should include a figure or two and its/their caption on the top half, and a few bullet points/short blocks of text interpreting that piece of data below. (Reminder: Your figure captions themselves should avoid interpretation.)

Take some time to look at how Figures are presented in the scientific literature -- they are almost never full page and each data plot is typically on the order of 3 inches x 3 inches. Use these visual guidelines to help you construct data pages that fit both your figure, caption, and interpretation bullet points. Remember that when you shrink a figure, you must make sure it remains interpretable!

More detailed suggestions for content (as opposed to style) are below.

Content guidelines

Begin by reading the general guidelines for scientific writing. In particular, the sections on Title, Abstract, Figures, and Holistic View of Data are particularly applicable to this assignment.

A few prompts to get you started are below, but note that this list is not exhaustive and also that several elements could reasonably be included in more than one section.

Background and motivation: potential topics and figures

  • Topic: Why is measuring DNA damage interesting and/or useful?
  • Topic: How does HR work?
  • Figure: Depiction of HR
  • Topic: How does the HR assay work?
  • Schematic: HR assay approach
    • You may prepare something similar to the assay depiction from the lecture notes, but should NOT copy and insert it directly. Your goal should be to make a figure tailored specifically to this assignment and audience. What elements might be cut or added? How can you modify the figure to best highlight key takeaways?
  • Topic: What kinds of questions can the HR assay address?

Data: potential topics and figures

Figures and topics are listed below according to the two major phases of your experiment. Within each phase, you should look for sub-groupings of interest, rather than treat each piece of data in isolation. In other words, try to both interpret and communicate outcomes holistically.

Keep in mind that you are describing the detailed methods in a separate assignment. Therefore, figure captions and/or supporting text should include only the most relevant aspects of the methods, such as the names of the diagnostic enzymes, a clear description of any normalization or statistics done on the flow cytometry data, etc.

System construction: making and verifying plasmid

  • Schematic: Overall approach
    • You may prepare something similar to the M1D1 Intro figure, but should NOT copy and insert it directly.
  • Topic: Apparent success of PCR, digestion, and recovery, including role of controls when applicable
  • Figure: Gel of digested DNA prior to cloning
  • Figure: Recovery gel of purified, digested DNA
  • Topic: Apparent success of ligation and transformation, including role of controls when applicable
  • Topic: Apparent success of cloning, explicitly including predicted vs. observed sizes, extraneous bands, and role of controls when applicable
  • Schematic: Diagnostic digest plan, for example in marked up plasmid map form
  • Figure: Diagnostic digest gel

DNA repair assay

  • Schematic: Overall approach and question being asked
    • You may further modify the background section figure (or create a new one) to emphasize specific samples, if you did not do so before
  • Topic: Rationale for gating choices, particularly on FL1-FL2 plot.
  • Topic: Understanding controls: What is the purpose of the negative control? the positive control? the two single plasmid controls?
  • Figure: Sample raw flow cytometry data from own experiment
    • at a minimum, include one FSC-SSC plot, as well as fluorescence plots for a negative control, a positive control, and one experimental sample
  • Figure: Processed individual flow cytometry data (e.g., bar chart and error bars, where applicable)
  • Figure: Processed class-wide flow cytometry data (e.g., bar chart and error bars)
    • For class-wide data, make sure you are making meaningful comparisons. One plot that shows everyone's data without a thoughtful purpose or interpretation will result in a poor score.
    • Please use a separate plot or dual-axis plot to keep the positive control from dwarfing the other data
  • Topic: Flow data analysis: for the sub-topics below, be as specific as you can be. When possible, use supporting statistics to make your argument.
    • In more depth than you may have described above, what can you learn from the positive control? If two groups have different GFP+ values (say, 60% and 80%), what does that outcome mean and how might it influence your later data analysis?
    • Did the outcomes for the single plasmid controls match your expectations? How might you explain any discrepancies?
    • Did the relative amounts of HR (both individual and classwide) match your expectations/hypotheses? How might you explain any discrepancies between outcomes and expectations?
    • For repeated samples, does the class-wide data mark any of your own data as suspect?

Implications and future work: potential topics and figures

  • Topic: Based on the results, whether they matched your expectations or not, what experiments might you recommend next? Follow-up experiments could distinguish between competing explanations of a given outcome or broaden the sample set for a question you already asked, to give just two examples.
  • Topic: How might this assay be improved?
  • Topic: How might this assay be used as a research tool? in the clinic? in the pharmaceutical industry?
  • Topic: If you were studying two cell lines that showed different levels of HR with this assay, what more could you learn by tagging BRCA2 protein with RFP (red fluorescent proteins) in the same two cell populations?
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