Atomic force microscopy of DNA-colloidal gold and DNA-protein complexes

Date
1993
Authors
Dobbs, Drena
Niu, Luming
Shaiu, Wen-Ling
Vesenka, James
Henderson, Eric
Larson, Drena
Henderson, Eric
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Zoology and Genetics
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Zoology and Genetics
Abstract

The atomic force microscope (AFM)1 is capable of imaging and manipulating nucleic acids in solution and in air29' 13 We are developing methods for random and site-specific labeling of individual DNA molecules to facilitate manipulation of fragments excised in the AFM and for localization of specific DNA domains, such as protein binding sites and origins of replication. One successful method was to incorporate biotinylated nucleotides at random internal locations or specifically at the ends of linearized DNA molecules in vitro. Following complex formation with Snm diameter streptavidin-gold conjugates, chromatographic purification and passive adsorption of the complexes to mica, the biotinylated domains were easily localized in the AFM by virtue of the distinctive size and shape of the streptavidin-gold complex. In many cases unconjugated streptavidin (i.e., lacking gold) was also observed attached to the biotinylated DNA. A second approach to site-specific labeling of DNA for imaging in the AFM was to react DNA with restriction enzymes having sequence-specific binding properties. Like the unconjugated streptavidin-DNA complexes, these enzyme-DNA complexes were visible without attached colloidal gold. Efforts to image DNA labeled in vivo using bromodeoxyuridine (BrdU) and anti-BrdU antibodies are ongoing.

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Luming Niu ; Wenling Shaiu ; James Vesenka ; Drena D. Larson ; Eric Henderson; Atomic force microscopy of DNA-colloidal gold and DNA-protein complexes. Proc. SPIE 1891 (1993); doi:10.1117/12.146706.

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