URMC / Labs / Kielkopf Lab / Lab Protocols
Lab Protocols
General
Nanodrop UV/VIS Spectrophotometer
Lab Key Code
Washing Glassware - Manual
Washing Glassware - Dishwasher
Using Autoclave
Protein Purification and Biochemistry Protocols
Care for Akta Purifiers
Akta Fraction Collector Racks
Remote Connection to Akta Pures
E. coli Large Scale Induction of Protein Expression
Lysis of E. coli by Sonication
Lysis of E. coli by Avestin press
Protein Purification Stock Buffers
Bacterial Cell Resuspension Buffers
Protein Purification Running Buffers
Purify a Protein with a GST Tag
Purify a Protein with a Histidine Tag
Purify a Protein with Acidic pI
Purify a Protein with Basic pI
Purify a Protein with Thrombin Cleavage Site
Selenomethionine Protein Purification
A280 Protein Concentration Determination
Dialysis Tubing
Laemmli Method for SDS-PAGE Gels and Buffers
Tricine Gels
Prepare Sigma glutathione agarose
Expression Test
GST Pull-Down
GST Competition Pull-Down
PreScission Protease Purification and Use
TEV Protease Purification and Use
U2AF65 DLink Purification
SF1 Purification
U2AF65 UHM Purification
Nucleic Acid Protocols
DNA Desalting 2g SepPak
DNA Desalting 200mg SepPak
Nitrocellulose Filter Binding
Electrophoretic Mobility Shift Assays
Adjusting PhosphoImager Image Quality
SYBR Gold Staining of Nucleic Acids
Thermodynamic Protocols
BIAcore T100 for kinetic or affinity measurements
Isothermal Titration Calorimetry using Kielkopf VP-ITC
Isothermal Titration Calorimetry using Wedekind PEAQ-ITC
Circular Dichroism Protocol: Jasco in SBBF
Circular Dichroism Protocol: Prior Aviv
Fluoromax-3 Polarizer Alignment
Fluorescence Anisotropy RNA Binding Assay
Using Fluoromax-4 Fluorimeter
Equations for Fitting Fluorescence Titration Assay
Electrophoretic Mobility Shift Assay
Amino Acid Analysis to Confirm Concentration for ITC
Effect of Interdomain Linker on Kd
Structural Biology Protocols
Vitrobot for cryoEM Grid Preparation
Dynamic Light Scattering (DLS)
Small Angle X-ray Data Collection at ALS (with Greg Hura's help)
Scaling Pr functions calculated from SAXS data
Running MONSA on SAXS data
Bruker CCD login information
Crystal Photos
Crystal Flash-Cooling Tips
Remote Data Collection at SSRL
Prepare SeMet Protein
Sazaki Table of f' and f"
Molecular Biology Protocols
Making LB for Plates and Liquid Media
Antibiotics Guide
Bacterial Transformation
Single site mutagenesis
PCR Primer Design for Subcloning
PCR
Restriction Enzyme Digest of DNA Insert and Vector
Gel Purifying Your PCR Product
Analyzing Plasmid DNA by Gel Electrophoresis
Photographing DNA Gels
Ligation
Colony PCR
Overnight Cultures of Successful Transformants
DNA prep for sequencing
DNA sequencing at UR
Freezer stocks of transformed bacteria
Making Competent Cells
Tissue Culture and Related Protocols
Endotoxin-free Maxipreps
Tissue Culture Guide
Quantitative Real-Time Reverse Transcription-PCR
Immunoblot Protocol
Mary Pulvino’s coIP videos (courtesy of Justin Galardi)
Co-ImmunoPrecipitation
In Vitro Splicing Assay
Quantifying RT-PCR images