Three-Dimensional Reconstruction of Chromosomes Using Electron Tomography
The evolvement of preparative methods in structural studies has always been as important as the development of sophisticated equipment. Software development is also a significant part for three-dimensional (3D) structural studies using electron tomography methods (ETMs). Advanced computing makes amenable procedures that relatively recently were only visionary, such as the 3D reconstruction of chromosomes with ETM. Morphological guidelines and beauty are occasionally the only standard for a method to be acceptable in the realms of preparative as well as software development. Bulk isolation of metaphase chromosomes using acetic acid is such an apparent accomplishment in preparative methods. Our ETM with maximum entropy and, more so, the ongoing development toward fully automatic alignments, are contributions in the software line. Furthermore, whole mounting of chromosomes on holey-carbon grids makes it possible to use even yesterday’s 80-kV transmission electron microscope with a standard goniometer to collect tilt series. These advances in preparing whole-mount metaphase chromosomes enable laboratories that do not have access to a medium- or high-voltage transmission electron microscope to study complex structures like chromosomes in 3D using today’s desktop computers.
Key WordsWhole-mounted chromosomes electron microscopy 3D reconstruction stereo pairs electron tomography maximum entropy tilt-series automatic alignment
- 1.Frank, J. (ed.) (1992) Electron Tomography: Three-Dimensional Imaging With the Transmission Electron Microscopy. Plenum Press, New York.Google Scholar
- 4.IMOD (2005). Available at: URL: http://bio3d.colorado.edu/imod/. Accessed May 29, 2006.
- 5.Engelhardt, P., Meriläinen, J., Zhao, F., Uchiyama, S., Fukui, K., and Lehto, V.-P. (2006) Whole-mount immunoelectron tomography of chromosomes and cells, in Electron Microscopy Methods and Protocols, 2nd ed (Kuo, J., ed.), Humana, Totowa, NJ, pp. 387–405.Google Scholar
- 11.BOB (2005) Available at: http://www.ahpcrc.org/software/bob/. Accessed May 30, 2006.
- 12.AnimaBob (2005) Available at: http://www.borg.umn.edu/~grant/AnimaBob/. Accessed May 30, 2006.
- 13.Chimera (2005) Available at: http://www.cgl.ucsf.edu/chimera/. Accessed May 30, 2006.
- 14.VMD (2005) Available at: http://www.ks.uiuc.edu/Research/vmd/. Accessed May 30, 2006.
- 15.SITUS (2005) Available at: http://situs.biomachina.org/. Accessed May 30, 2006.
- 16.Vis5D (2005) Available at: http://www.ssec.wisc.edu/~billh/vis5d.html. Accessed May 30, 2006.
- 17.Sullivan, A. (2005) 3-Deep. IEEE Spectrum, INT. April 22–27.Google Scholar