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Title:Zernike Phase Contrast electron cryo-tomography of cyanophage Syn5 assembly intermediates
Authors:Dai W, Fu C, Raytcheva D, Flanagan J, Khant HA, Liu X, Rochat RH, Haase-Pettingell C, Piret J, Ludtke SJ, Nagayama K, Schmid MF, King JA, Chiu W
Sample:Cyanophage Syn5 assembly intermediate inside cyanobacteria Synechococcus WH8109 cells
Method:Subtomogram averaging (52 angstroms resolution)
Red flagLatest update:2013-11-06
Sample
Sample name: Cyanophage Syn5 assembly intermediate inside cyanobacteria Synechococcus WH8109 cells
Components:
ID Type Name Exp. MW (MDa) Theo. MW (MDa) Oligomeric details Recombinant expression Synthetic Organism UniProt identifier GO identifier InterPro identifier Virus identifier Details
1virusSynechococcus phage Syn5Synechococcus phage Syn5DNA-containing capsid with tail and horn - assembly intermediate
Experiment
Specimen state: Particle
Specimen preparation:
pHSpecimen conc.DetailsStainingSpecimen support details
8.0 mg/mLartificial sea water200 mesh Quantifoil holey grids, glow discharged
Vitrification:
Cryogen nameHumidityTemp.Instr.MethodTime resolvedDetails
ETHANE95%85 KFEI VITROBOT MARK IIIBlot for 3 seconds before plunging ms
Imaging:
MicroscopeVoltageIllumination modeImaging modeCsDefocus min.Defocus max.Nominal mag.Calibrated mag.Electron sourceDetectorDetector distanceAstigmatism
JEOL 2200FS200 kVFLOOD BEAMBRIGHT FIELD4.1 mm0 nm1000 nm2500025000FIELD EMISSION GUNGATAN ULTRASCAN 4000 (4k x 4k)1200 mmObjective lens astigmatism was corrected at 100,000 times magnification

Specimen holderHolder modelTilt min.Tilt max.Energy filterEnergy windowTemp.Temp. min.Temp. max.Beam tiltElectron doseOther detailsDate
GATAN LIQUID NITROGEN-60°60°in-column GIF energy filter15-20 eV103 K93 K105 K mrad1 e/Å201-JUN-2011
Processing
Protocol:back projection
Software:IMOD
Number of subtomogram averaged:96
Number of class averages:1
Imposed symmetry:C1
Resolution by author:52 Å
Resolution method:FSC 0.143
Processing details:The subtomograms were extracted from infected cells.
Scanned images:
Num. imagesSampling sizeOD rangeQuant. bit numberOther detailsScanner
42 μm/pixel16
Fitting:
PDBProtocolTarget crit.SoftwareB valueFitting spacePDB chainDetails