Visualization of BrdU-labelled DNA in cyanobacterial cells by Hilbert differential contrast transmission electron microscopy.
We have attempted to observe the native shape of DNA in rapidly frozen whole cyanobacterial cells through 5-bromo-2-deoxyuridine (BrdU) incorporation and visualization with a Hilbert differential contrast transmission electron microscopy (HDC TEM). The incorporation of BrdU into the DNA of Synechococcus elongatus PCC 7942 was confirmed with fluorescently labelled anti-BrdU antibodies and through EDX analysis of ultra-thin sections. HDC TEM observed cells that had incorporated BrdU into their DNA exhibited electron dense areas at the location corresponding to fluorescently labelled BrdU. Since various strings and strands were observed in high contrast with the HDC TEM, we conclude that the method promises to allow us to identify and understand bulk structural changes of the in vivo DNA and the nucleoid through observation at high resolution.
Nitta
K
K
Terabase Inc., Myodaiji-cho, Okazaki 444-8787, Japan.
Nagayama
K
K
Danev
R
R
Kaneko
Y
Y
D001973:Bromodeoxyuridine / Q000737:chemistry* / Q000378:metabolism; D004269:DNA, Bacterial / Q000737:chemistry* / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D007053:Ice; D046529:Microscopy, Electron, Transmission / Q000379:methods*; D008856:Microscopy, Fluorescence; D046940:Synechococcus / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D014961:X-Ray Diffraction
D004269:DNA, Bacterial; D007053:Ice; D001973:Bromodeoxyuridine