EMD-2162 Experiments and Validation

Structure of bacteriophage phi29 prohead particle with a double insertion mutation (U92,U93) in the pRNA

Single particle reconstruction
Overview of EMD-2162
Sample name: Bacteriophage phi29 prohead particle with a double insertion mutation (U92,U93) in the pRNA
Organism: Bacillus phage phi29
Related EM entry by publication: EMD-5451

Map parameters

Minimum density: -2.999
Maximum density: 6.227
Average density: 0.00
Standard deviation: 0.957
Recommended contour level: 1.50 (author)

Sample information

Sample name: Bacteriophage phi29 prohead particle with a double insertion mutation (U92,U93) in the pRNA
Virus: Bacillus phage phi29

Validation information

Experimental information

 

Image processing

Applied symmetry: C5
Software: EMAN
Number of particles: 2134
Reconstruction protocol: Model-based projection methods for alignment and Fourier methods for reconstruction
CTF correction: Each micrograph
Details: C5 symmetry was imposed in all stages of the reconstruction, with the 5-fold axis along Z. The final map was low-pass filtered at 14 angstrom units The particles were selected using the model-based automatic particle selection implemented in EMAN, followed by manual deletion of bad particles and manual selection of missed particles

Imaging

Session
Microscope model: JEOL 2200FS
Electron source: FIELD EMISSION GUN
Electron dose (e/A**2): 20.0
Energy filter: JEOL
Nominal CS (mm): 2.00
Nominal magnification: 60000.0
Defocus max (nm): 3,000.00
Defocus min (nm): 640.00
Holder model: GATAN LIQUID NITROGEN
Acquisition
Number of images: 109

Specimen

Preparation
Specimen state: particle
Specimen concentration (mg/mL): 0.10
Buffer: 25 mM Tris-HCl, 5 mM MgCl2, 50 mM NaCl
Specimen support: Quantifoil holey carbon on top of200 mesh copper grid, plasma cleaned
Vitrification
Apparatus: HOMEMADE PLUNGER
Cryogen: ETHANE
Protocol: Blot grid from behind the sample for ~4 seconds before plunging.