54. Third-Strand Binding Probes for Duplex DNA in Particles of Varying Size 

Marion D. Johnson III and Jacques R. Fresco 
Princeton University, Department of Molecular Biology, Princeton, New Jersey 
mjohnson@molbio.princeton.edu, jrfresco@princeton.edu 

Third-strand oligonucleotide binding to native double-stranded DNA in some free form or in chromatin via triple helix formation provides a sequence specific way to attach ligands, including cytochemically detectable ones, to DNA-containing structures. Last year we described the development of an in situ methodology for identifying metaphase chromosomes. We particularly demonstrated binding of a highly specific fluorescent third-strand probe directed to a 16 base pair DNA -satellite target of human chromosome 17. We continue to exploit this type of interaction for various purposes of relevance to the Human Genome Project. 

Some of our recent efforts have been directed at: 

1. Developing probes specific for the a-satellite regions of human chromosomes X and 16. Such probes expand our capacity to cytogentically identify and isolate individual human chromosomes. 

2. Exploiting such third strand probes to greatly facilitate isolation by flow sorting of individual human chromosomes and possibly chromosomes of other species. 

3. Developing fluorescent-labeled probes to identify accessible multicopy sequences within the Drosophila histone gene cluster. These probes are being employed to locate such target sequences in Drosophila ovarioles. 

To assure these ends, we have also investigated the consequences of DNA structure and size on the mechanism and equilibria of probe binding.