20.109(S24):M1D8

Introduction

The goal for today is to give you time and space to get started on the Mod 1 major writing assignment, the Data summary. Be sure to take advantage of the 'work day' to discuss the assignment with your laboratory partner and to ask the Instructors any questions you may have about the material covered, data analyzed, or assignment details.

Protocols

Part 1: Image EMSA experiment

1. Obtain the membrane with your EMSA experiment from the front laboratory bench.
2. Decant the 0.5X TBE buffer from the container.
3. Wash the membrane with X mL of 1X Detection Wash Buffer.
   • To wash the membrane, rock the container 3-5 times, then decant the buffer.
4. Add X mL of Blocking Buffer, then incubate for 20 minutes at room temperature at 50 rpm.
5. Remove 1 mL of the Blocking Buffer from Step #4 and transfer to a microcentrifuge tube.
6. Add X μL of Streptavidin-HRP Conjugate to the 1 mL aliquot of Blocking Buffer.
7. Transfer the 1 mL of Blocking Buffer + Streptavidin-HRP Conjugate into the container with your membrane.
8. Incubate the membrane for 45 minutes at room temperature at 50 rpm.
9. Decant the Blocking Buffer.
10. Wash the membrane with X mL of 1X Detection Wash Buffer for 10 minutes at room temperature at 50 rpm.
    • Complete a total of three washes.
11. In a microcentrifuge tube, mix XX mL of Substrate A with XX mL of Substrate B.
12. Carefully place your membrane between the flaps of the Detection Sheet.
13. Hold the top flap of the Detection Sheet away from your mebrane, then use a pipet to transfer all of the XX mL of the Substrate Solution onto your membrane.
    • Pipet the Substrate Solution evenly across the surface of your membrane.
14. Gently lay the top flap down onto the membrane ensuring that the Substrate Solution covers the entire surface without trapping air bubbles.
15. Incubate for 5 minutes and room temperature.
16. Alert your Instructor when you are ready to image your membrane.

Part 2: Outline Data summary and divide workload accordingly

In science publications it is uncommon to find single-author papers. Most science is done in collaboration with peers and colleagues, and this is true for your Data summary assignment. You will work with your partner to prepare a draft and revision that describes the work you completed in Module 1. To ensure the work is distributed evenly you will first assess what needs to be done to complete the assignment and then decide who will be responsible for which parts.

Assess what needs to be done

Prepare an outline for your Data summary that highlights the components that should be included (i.e. "Background & Motivation" and "Results, Figure 1"). For each component, list what has been done (for example, "prompts regarding information for Background & Motivation section from due M1D2 homework") and what needs to be completed (for example, "use due M1D2 answers to write bullets for Background & Motivation section").

Distribute what needs to be done between partners

With your partner, decide who will tackle which components of the assignment. Discuss what work needs to be completed such that you are both in agreement on what is expected. Then discuss who will be responsible for that work. Review the submitted homework assignments and comments as this may help determine which partner should complete which components (hint: which homework is most easily revised based on the comments?).

Note that the assignment is graded as a whole and both partners will receive a score based on the whole and not their individual contributions. In this, it is important that both partners are involved in all components...either as the primary writer or as a reviewer. Next, coordinate times that you can meet to discuss the progress of the work and / or to review the work. It is also helpful to set deadlines for when certain work should be done so both partners are able to stay on track and are accountable to each other.

In your laboratory notebook, complete the following:

• Based on your discussion with your laboratory partner, record the following:
  • What has been done?
  • What needs to be done?
  • Who will complete which components?
  • When will you meet to discuss progress and review the completed components?

Part 3: Prepare Data summary

Use the remaining class time to prepare your Data summary!!.

Reagent list

Navigation links

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