Sample information
Sample name: TBP-TFIIA-TFIIB-DNA-Pol II-TFIIF-TFIIE (open)
Nucleic acid:
Core promoter DNA
Proteins:
TATA-box-binding protein, General transcription factor IIA, General transcription factor IIB, General transcription factor IIF, General transcription factor IIE, DNA-directed RNA polymerase II
Validation information
Experimental information
Image processing
Applied
symmetry: C1
Software: EMAN2, SPARX
Number of
particles: 53505
Reconstruction
protocol: Projection matching
CTF
correction: whole micrograph
Details: Image processing was performed in the Appion processing environment. 3D reconstruction was performed using EMAN2 and SPARX libraries. Final map was filtered to local resolution using the blocres function in Bsoft package.
Imaging
Session
Microscope
model: FEI TECNAI F20
Electron
source: FIELD EMISSION GUN
Electron
dose (e/A**2): 20.0
Nominal
CS (mm):
2.20
Nominal
magnification: 100000.0
Defocus
max (nm):
2,400.00
Defocus
min (nm):
1,200.00
Holder
model: GATAN LIQUID NITROGEN
Details:
Data acquired using Leginon
Acquisition
Number
of images: 2800
Specimen
Preparation
Specimen
state: particle
Specimen
concentration (mg/mL):
0.09
Buffer:
10 mM HEPES, 5% glycerol, 10 mM MgCl2, 50 mM KCl, 1 mM DTT, 0.05% NP-40
Specimen
support: 400-mesh C-flats with thin carbon support over 4um holes with 2um spacing (Protochips Inc.)
Vitrification
Apparatus:
FEI VITROBOT MARK II
Cryogen:
ETHANE
Humidity:
100
Protocol:
Incubation on grid for 5 minutes before blotting 4 seconds at -1 offset.