Chromosome-specific DNA Repeat Probes
Adolf Baumgartner 1, Jingly Fung Weier 1 and Heinz-Ulrich G. Weier 1*
1 Department of Obstetrics, Gynecology and Reproductive Sciences, University of California, San Francisco, California (AB,JFW), and Life Sciences Division, University of California, E.O. Lawrence Berkeley National Laboratory, Berkeley, California (AB,JFW,H-UGW)
* To whom correspondence should be addressed. E-mail: ulliweier{at}hotmail.com.
Submitted on March 20, 2006
Accepted on 21 July 2006
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Abstract |
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In research as well as clinical applications, fluorescence in situ hybridization (FISH) has gained increasing popularity as a highly sensitive technique to study cytogenetic changes. Today, hundreds of commercially available DNA probes serve the basic needs of the biomedical research community. Widespread applications, however, are often limited by the lack of appropriately labeled, specific nucleic acid probes. We describe two approaches for an expeditious preparation of chromosome-specific DNAs and the subsequent probe labeling with reporter molecules of choice. The described techniques allow the preparation of highly specific DNA repeat probes suitable for enumeration of chromosomes in interphase cell nuclei or tissue sections. In addition, there is no need for chromosome enrichment by flow cytometry and sorting or molecular cloning. Our PCR-based method uses either bacterial artificial chromosomes (BAC) or human genomic DNA as templates with alpha satellite specific primers. Here, we demonstrate the production of fluorochrome labeled DNA repeat probes specific for human chromosomes 17 and 18 in just a few days without the need for highly specialized equipment and without the limitation to only a few fluorochrome labels.
Key Words:
chromosome enumeration, DNA repeats, DNA probes, fluorescence in situ hybridization (FISH), chromosomes 17 and 18
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