20.109(S15):Data analysis (Day7)

From OpenWetWare
Jump to: navigation, search


20.109(S15): Laboratory Fundamentals of Biological Engineering

Dilution curves F13.jpg

Home        People        Schedule Spring 2015        Assignments        Lab Basics        OWW Basics       
DNA Engineering        System Engineering        Biomaterials Engineering              

Introduction

Protocols

Part 1: Visit by Leslie to discuss the Discussion section

We will start the day with a visit by our writing instructor, Leslie, who will talk about the structure of a Discussion section, the only part of a formal research paper you have yet to write.

Part 2: Statistics practice

You may find averages, variance (standard deviation or 95% confidence intervals), and t-tests useful when you report on class results.

You can practice the steps below using the male and female heights that we collect during pre-lab lecture.

  1. Begin by downloading the following Excel file as a framework to carry out the basic statistical manipulations we discussed in pre-lab lecture. The file is modified from one originally written by Professor Bevin Engelward.
  2. Find and plot 95% confidence intervals for the male and female heights.
  3. Compare the means of these two populations. At what confidence level (if any) are they different?
    • Would a one-tailed or two-tailed test be more appropriate for this comparison?

Part 3: Stain irradiated cell colonies

Your irradiated cells have now been growing for 7 days. Time to count colonies!

  1. Briefly observe your irradiated cells on the TC microscopes. Are you able to find some colonies? About how many cells are in some of these small dispersed clusters?
  2. Take your plate to the main lab for the remaining steps. First, aspirate the media. You don't need to change the yellow tip between samples if you move from highest drug dose to lowest, as that should be the order of least to most colonies.
  3. Rinse each well with about 2 mL of PBS.
  4. After removing the PBS, use a serological pipet to consistently add 2 mL of Coomassie to each well.
  5. Place your plate on the shaker at 80 rpm for about 1 hour.
  6. Repeat the PBS rinse.
  7. Let the well plate dry for a short time after aspirating the PBS.
    • If you wait a very long time the stain will begin to fade.
  8. Finally, count the colonies in each well and document these on today's Talk page.
    • Do your best to apply a consistent standard for threshold colony size and threshold staining intensity. As long as you are consistent, there is not one right answer as to what constitutes a colony.
    • One of your instructors finds it easiest to count colonies by making a dot with a lab marker as she counts each colony, right on the underside of the 6-well plate at the colony location, and then writing down each decade (10, 20, etc.) on the plate as she reaches that number so she doesn't lose her place.
  9. Whether today or on your own, you should plot your data by surviving fraction (as described in the Nature Protocols paper linked in the M2D6 introduction) for the Module 2 report. If you are feeling ambitious (and your data is appropriate) you can estimate the IC50 for your inhibitor. Does it agree with the published values?

Reagent list

  • PBS
  • Bio-Safe Coomassie Stain (Bio-Rad)
  • Mostly your brains!

Navigation Links

Next Day: Growth of phage nanowires

Previous Day: Flow cytometry and paper discussion