JV5443

Description

Technical info

Tilt Series Date: 2016-08-09

Data Taken By: Debnath Ghosal

Species / Specimen: Legionella pneumophila

Strain: Philadelphia-1_Lp02

Tilt Series Settings: Single Axis, tilt range: (-60.0°, 60.0°), step: 1.50°, constant angular increment, dosage: 100.0 eV/Ų, defocus: -6.0 μm, magnification: 26000x.

Microscope: Janelia Titan

Acquisition Software: UCSFTomo

Upload Method: pipeline

Processing Software Used: Raptor

Collaborators and Roles: Strain sent to us by Joseph P. Vogel, Washington University School of Medicine. Data collected and processed by Debnath Ghosal at Janelia Krios.

Purification / Growth Conditions / Treatment: Cells were grown in AYE liquid culture with following supplements: ferric nitrate and cysteine hydrochloride (no thymidine). Scrape cells off a plate, dissolve in the media, adjust the cell density to 0.6 and make a dilution series. Start O/N culture with all of them and freeze cells next day from the one in early stationary phase that is OD ~ 3.0.

Sample Preparation: 8 grids were frozen for the JV5443 strain at OD~3.2. Vitrobot (MK iv) settings: 100% humidity, 4µl sample, Blot force 6, blot time 3 sec. Wait time 0 sec. Drain time 1 sec. Ethane/propane mix. Grid: Quantifoil Cu 2/2 200. Add gold to cells prior to freezing: 1ul of gold to 4 ul of cells, freeze.

Attached Files

JV5443a0084.mrc, Tilt Series (Pixel Size 0.42 nm), 2.3 GB

JV5443a0084_full.rec, Reconstruction (Pixel Size 1.68 nm), 445.4 MB

dga2016-08-09-31_slicer16939.png, , 414.2 kB