A413T cooverexpression

Description

Technical info

Tilt Series Date: 2013-08-30

Data Taken By: Ariane Briegel

Species / Specimen: Escherichia coli

Strain: RP 437

Tilt Series Settings: Single Axis, tilt range: (-60.0°, 60.0°), step: 1°, constant angular increment, dosage: 160.0 eV/Ų, defocus: -10.0 μm, magnification: 34000x.

Microscope: Caltech Polara

Acquisition Software: UCSFTomo

Upload Method: pipeline

Processing Software Used: Raptor

Collaborators and Roles: Laura Kiessling

Purification / Growth Conditions / Treatment: Grew UU1607/pJC3/pPM25 overnight in 5mL TB + 3.8µl Cm + 5µl CarbDiluted back 1:50 into 200mL TB + 152µl Cm + 200µl Carb + 50µl 1M IPTG + 200µl Na-salicylateGrew ~2 hours to mid-log phaseSpun down 10' at 5,000 x gResuspended in 300µl lysis buffer + 0.1µl PMSF + 0.75 µl 1,10-phenanthrolineAdded 33.3µl (2mg/ml) fresh lysozyme solution (20mg/ml in 10mM Tris-HCl, pH 8.0)Incubated 45' at 37°CAdded 1.67µl 1M MgCl2, 0.2µl 1M CaCl2, 0.33µl DNase IIncubated 30' at 37°CSpun down 4000 g 5 min Resuspended in activity buffer (cold room) without glycerol

aba2013-08-30-32_slicer9546.jpg: Escherichia coli . vesicle aba2013-08-30-32_slicer9547.jpg: Escherichia coli . vesicle

Attached Files

A413tCoover0033.mrc, Tilt Series (Pixel Size 0.32 nm), 3.4 GB

A413tCoover0033_full.rec, Reconstruction (Pixel Size 1.28 nm), 712.7 MB