KLH Dataset II » History » Revision 30
Revision 29 (Eric Hou, 07/23/2010 02:35 PM) → Revision 30/34 (Eric Hou, 07/23/2010 02:41 PM)
h1. Public Data Sets: KLH2
*KLH Dataset II:* 655 defocus pairs of micrographs, coordinate centers of ~10000 side-view particles, a preliminary 3D reconstruction.
h2. 1. Images.
Images were acquired with a FEI Tecnai F20 equipped with a 2Kx2K CCD Tietz camera, as defocal pairs at a nominal magnification of 62,000 x and a voltage of 120 KeV, using the Leginon system ("Potter et al., 1999":http://www.ncbi.nlm.nih.gov/pubmed/10406132 ; "Carragher et al., 2000":http://www.ncbi.nlm.nih.gov/pubmed/11121305). The first image (named *.001.mrc) is acquired at near to focus (NTF) conditions (-0.6 to -1.5 µm) and the second image (named *.002.mrc) at farther from focus (FFF) conditions (-3µm). The time interval between the two exposures is approximately 20s due to the time required to read out the digital image from the camera. The pixel size is 2.2Å at the specimen scale and the accumulated dose for each high magnification image area was ~10 e/Ų.
Below is an example of a defocus pair. Click on the images to see at full scale.
p=. (a) !fig1a.jpg!:http://emg.nysbc.org/prtl_data/klh/klh_10k/fig1a.jpg (b) !fig1b.jpg!:http://emg.nysbc.org/prtl_data/klh/klh_10k/fig1b.jpg
(a) Near to focus - NTF image. (b) Far from focus - FFF image.
p=. *Figure 1: An example pair of high magnification images of KLH.*
*Downloading High Magnification Images:* Image files are provided in "MRC":http://emg.nysbc.org/prtl_data/mrc_specification.htm format and as JPEG files
Download entire set of images using a tool called [[dbemwiz|Dbemwiz]], and use section name "02oct09a".
We suggest using a tool called "em2em":http://www.imagescience.de/em2em, to convert MRC files to other formats.
h2. 2. Positions of the Picked Particles in the Images
Particles are picked in the FFF images and these coordinates are used to select particle in the NTF images. Phase correlation is used to align the two images to calculate the offset between the FFF coordinates and the NTF coordinates.
p=. (a) !fig3a.jpg!:http://emg.nysbc.org/prtl_data/klh/klh_10k/fig3a.jpg (b) !fig3b.jpg!:http://emg.nysbc.org/prtl_data/klh/klh_10k/fig3b.jpg
(a) The NTF image. (b) The FFF image.
p=. *Figure 3: An example pair of images outlined with particles selected by Selexon. Each "+" indicate a detected particle.*
h2. 2.1 Automated Picking
The method described in ("Zhu (Zhu et al., 2003":http://www.ncbi.nlm.nih.gov/pubmed/14643198) 2003) was used to select 11309 side-view particles from the FFF images. Particle coordinates are provide below:
* "Coordinates":http://emg.nysbc.org/prtl_data/klh/klh_10k/selexon.pik.tar of 11309 side-view particles selected in the FFF images.
h2. 3. 3D Reconstructed Map
A 3D reconstruction was generated using automatically particles using D5 point-group symmetry and several cycles of refinement.
* "Coordinates":http://emg.nysbc.org/prtl_data/klh/klh_10k/prtl_in_3dmap.tar of 10997 side-view particles went into the map.