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How to Get Started on an NMR Project

  1. Expression System/Cell types
    * BL21(DE3)
    * HMS174(DE3)
    *** We typically use pET expression vectors***
  2. Appropriate Buffers
    * Tris
    * Hepes
    * Phosphate
    *** if you have questions about a buffer, just give us a call***
  3. Sample Conditions
    * Protein Concentration: We recommend a protein concentation of between 0.2mM - 2mM (less than 0.2mM is possible for some experiments if necessary)
    * Sample Volume: Samples need to be 600ul if using a standard NMR tube, or no less than 300ul if using a Shigemi tube
    * D2O: The final NMR sample must contain between 5% and 10% D2O
    * Salt Concentrations: Try to keep this LOW (<100mM total ionic strength)
  4. Things to buy:
    * Sample Tubes:
         - Shigemi
         - Wilmad
    * Labeled Isotopes:
         - CIL for 13C-glucose
         - CIL for 15N-ammomium sulfate
    * CIL for D20

 NMR Core - Protein NMR Data Graphic

The above image illustrates typical preliminary data collected at the start of a protein NMR project. The above spectrum takes approximately 15 minutes to collect and tells much about the feasibility of a given NMR project. NMR structural work is carried out on proteins grown with nutrients labeled with both 15N and 13C.