DOI: 10.22002/m7q4s-rp331
title: Vibrio cholerae SAD2095 induced

description: SAD2095 (T2SS and T4 competence pili) = rpoS*, pilA S81C, lacZ::lacIq, comEA-mCherry, ΔluxO, Ptac-tfoX, Ptac- recA, ∆TCP::ZeoR, ∆MSHA::CarbR, ∆CTX::KanR, ∆pilT::TmRAnkur: "For the taxonomy ID - unfortunately the strain we sent does not have a taxonomy ID. So, you can use the generic one for V. cholerae which is "666". The strain we are working with is related to N16961, but is certainly not the same strain, so not sure if you would want to indicate that somehow? Just for reference - the strain we sent is E7946 (although it is obviously heavily modified with a number of mutations). "
description: <strong>Tilt Series Date:</strong> 2018-07-18</p>
<strong>Data Taken By:</strong> Sara Weaver</p>
<strong>Tilt Series Settings:</strong>  Single Axis, tilt range: (-60.0°, 60.0°), step: 1°, constant angular increment, dosage: 180.0 eV/Å²,  defocus: -6.0 μm,  magnification: 27500x. </p>
<strong>Microscope:</strong> Caltech Polara</p>
<strong>Acquisition Software:</strong> SerialEM</p>
<strong>Upload Method:</strong> pipeline</p>
<strong>Processing Software Used:</strong> Raptor</p>
<strong>Collaborators and Roles:</strong> Strain from Ankur B Dalia ankdalia@indiana.edu, PhD, Assistant Professor, Indiana University. Cells grown, data collected by Sara Weaver</p>
<strong>Purification / Growth Conditions / Treatment:</strong> Streaked stock on a plate. Grew up a single colony in 3 mL LB with shaking at 37˚C. The next day, subcultured 50uL of this overnight culture into 3mL of fresh LB+100uM IPTG + 20mM MgCl2 + 10mM CaCl2 and grow at 30˚C shaking for ~4 hours.</p>
<strong>Sample Preparation:</strong> spun gold (100 µL 10 nm gold + 25 µL 5% BSA) at 15,000rpm for 20 minutes, removed supernatant.  Resuspended gold beads in 20 µL liquid culture.  Vitrobot RT and 95% humidity, 3µL of gold/sample slurry on carbon/copper Quantifoil Finder grids R2/2 (EmiTech glow discharge 50 mA 120 s)  Blot force -6, blot time 3 s, no waiting time, no drain time.  Plunged into ethane/propane mix.</p>
  swj2018-07-18-11_slicer19668.jpg: green + orange - T4P competence pili ? .  pink - competence pilus basal body? .  blue - flagellar filament swj2018-07-18-11_slicer19669.jpg: green  - T4P competence pili ? .  pink - competence pilus basal body? swj2018-07-18-11_slicer19670.jpg: green  - T4P competence pili ? .  pink - competence pilus basal body? .  blue - flagellar filament swj2018-07-18-11_slicer19671.jpg: basal body? swj2018-07-18-11_slicer19672.jpg: membrane neck? swj2018-07-18-11_slicer19673.jpg: green - budding vesicle? .  pink - object on extracellular surface? swj2018-07-18-11_slicer19674.jpg: densities in periplasm swj2018-07-18-11_slicer19675.jpg: densities in periplasm swj2018-07-18-11_slicer19676.jpg: densities in periplasm
description: Files available via S3 at https://sdsc.osn.xsede.org/ini210004tommorrell/tomography_archive/swj2018-07-18-11/rawdata/</p>20180718_tiltseries_11.mrc,  Tilt Series (Pixel Size 0.39 nm), 2.8 GB  
        
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        20180718_tiltseries_11_part98_3.rec,  Reconstruction (Pixel Size 1.56 nm), 444.5 MB  
        
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