EMD-2192 Experiments and Validation

human RNA polymerase II in complex with B2 81-131 RNA

Single particle reconstruction
Overview of EMD-2192
Sample name: human RNA polymerase II in complex with B2 81-131 RNA
Organisms: Homo sapiens, Mus musculus
Related EM entries by publication: EMD-2190, EMD-2191, EMD-2193, EMD-2194, EMD-2189, EMD-2188

Map parameters

Minimum density: -0.06
Maximum density: 0.22
Average density: 0
Standard deviation: 0.02
Recommended contour level: 0.07 (author)

Sample information

Sample name: human RNA polymerase II in complex with B2 81-131 RNA
Nucleic acid: B2 81-131 RNA
Proteins: Rpb12, Rpb11, Rpb10, Rpb9, Rpb8, Rpb7, Rpb6, Rpb5, Rpb4, Rpb3, Rpb2, Rpb1

Validation information

Experimental information

 

Image processing

Applied symmetry: C1
Software: EMAN2, FREALIGN
Number of particles: 31423
Reconstruction protocol: projection matching
CTF correction: whole micrograph
Details: Projection matching using a low-pass filtered model of apo Pol II as the starting model

Imaging

Session
Microscope model: FEI TECNAI 20
Electron source: FIELD EMISSION GUN
Electron dose (e/A**2): 20.0
Nominal CS (mm): 2.2
Nominal magnification: 50000.0
Defocus max (nm): 5,000
Defocus min (nm): 2,000
Holder model: GATAN LIQUID NITROGEN
Acquisition
Number of images: 256
Scanner model: NIKON SUPER COOLSCAN 9000
Sampling interval (μm): 12.699999999999999

Specimen

Preparation
Specimen state: particle
Specimen concentration (mg/mL): 0.03
Buffer: 10 mM Tris pH 7.9, 10 mM HEPES pH 8.0, 4 mM MgCl2, 50 mM KCl, 0.05% NP-40, 1 mM DTT, 0.1% trehalose
Specimen support: 400 mesh copper grids covered with a holey carbon film, thin carbon film floated on top
Vitrification
Apparatus: FEI VITROBOT MARK II
Cryogen: ETHANE
Humidity: 100
Protocol: 6 sec blot, offset -2