Patricia L. Clark Research Lab: PROTOCOLS

Important Note: These protocols are intended as reference guides for experienced lab personnel only!
First-time users should get experienced hands-on assistance; this is mandatory for undergraduate researchers.

Authors: PLC, Patricia L. Clark; KF, Kay Finn; EK, Elizabeth Klimek; MSE, Mike Evans; MJ, Mirco Junker; KW, Kelli Whiteman; MMM, Michelle Munoz; BW, Beata Wielgus-Kutrowska.

Many of these protocols were inspired by protocols of the King lab at MIT; see their protocol page here.

 

Microbiology & Molecular Biology * Protein Purification * Gel Electrophoresis * Immunoassays * Spectroscopy * Images/Figures * Miscellaneous

Microbiology & Molecular Biology
  • LB Media and Plates [HTML] [PDF] (KF)
  • Minimal Medium [HTML] (PLC)
  • Phage Buffer
  • Making Plating Bacteria [HTML] [PDF] (KF)
  • Titering Phage [HTML] [PDF] (KF)
  • Streaking Phage
  • Making Phage Stocks
  • High-Titer Phage Stocks
  • Glycerol Stocks [HTML] [PDF] (KF)
  • EtOH Precipitation of DNA [HTML] [PDF] (MSE)
  • Plasmid DNA Miniprep [HTML] [PDF] (KF)
  • Plasmid DNA Midiprep [HTML] [PDF] (KF)
  • UV Determination of DNA Concentration [HTML] [PDF] (KF)
  

Purification of Proteins and Protein Complexes
  • Tailspike Purification (from Salmonella phage infection) (KF)
  • Tailspike Purification (from E. coli overexpression) (TFC)

 

  • P.69 Pertactin Purification, part one [HTML] [PDF] (MJ/KF)
  • FPLC Methods for Pertactin [HTML] [PDF] (KF)
  • Making Linear Sucrose Gradients [HTML] [PDF] (PLC)
  • Making Sucrose Gradients 2 [HTML] [PDF] (MSE)
  • Fractionating Sucrose Gradients [HTML] [PDF] (PLC)
  • Preparing Active Ribosomes [HTML] [PDF] (PLC)
  • Purifying Empty Ribosomes & Subunits (PLC)
  • GFP Prep: Part 1 [HTML] [PDF] (MMM)
  • Large GFP Prep [HTML] [PDF] (KF)
  • GFP Column Methods [PDF] (BW/KF)

Gel Electrophoresis
  • SDS Gel Recipes [PDF] (KF)
  • Native Gel Recipes [HTML] [PDF] (KF)
  • Pouring and Running SDS Gels [HTML] [PDF] (KF)
  • Native Gels

 

  • Agarose Minigels [HTML] [PDF] (KF)
  • Coomassie Staining
  • Silver Staining [HTML] [PDF] (KF/PLC)
  • Air Drying Gels with Cellophane
  • Using the Gel Dryer [HTML] [PDF] (KF)
  • Using the Phosphorimager [HTML] [PDF] (PLC)
  • Using the Densitometer [HTML] [PDF] (KF)
  • Scanning Gels:
  • Huber Lab [HTML] [PDF] (KF)
  • Clark Lab [HTML] [PDF] (KF)
  • Agarose Gel Documentation [HTML] [PDF] (KF)
  • Using NIH Image Software [HTML] [PDF] (MJ)

Immunoassays
  • Competition ELISA [HTML] [PDF] (PLC)
  • Indirect ELISA (MSE)

 

  • Plate Reader User Guide [HTML] [PDF] (MSE)
  • 96-well Plate Template [PDF] (PLC)

 
  • Immunoprecipitation [HTML] [PDF] (PLC)
  • Western Blotting [HTML] [PDF] (MSE)
  

Spectroscopy
  • Absorbance Measurements with the Baker Lab UV/Vis

 

  • Collecting Fluorescence Spectra with the Nowak Lab SLM8100 [HTML] [PDF] (EK)

 

  • Collecting CD Spectra with the Aviv [HTML] [PDF] (MJ)
  • Collecting Fluorescence Spectra with the PTI Fluorimeter [HTML] [PDF] (MJ)
  • Cleaning Cuvettes [HTML] [PDF] (KF)

Molecular Imaging & Figure Making
  • Creating Images with Swiss PDBViewer [HTML] [PDF] (PLC/EK)
  • Rendering PDBViewer Images with POV-Ray [HTML] [PDF] (PLC)

 

  • Panel Poster Printing [HTML] [PDF] (KF)
    

Miscellaneous
  • Refolding Tailspike (adapted from R. Seckler lab) [HTML] [PDF] (KF)

 

  • Unfolding and Refolding P.69 Pertactin (MJ)

 

  • Mike's Buffer Recipes [PDF] (MSE)
  • Radiolabeling Pulse/Chase Setup [HTML] [PDF] (PLC)

 

  • Partial Proteolytic Digestion with Proteinase K [HTML] [PDF] (KW)
  • Ultracentrifuge Accidents:
    MIT (1/99)
    Cornell (3/99)
University of Notre Dame Homepage Department of Chemistry & Biochemistry Homepage Molecular Biosciences Graduate Program Brochure Clark Lab Homepage