Abstract
In the multiple-color primed in situ labeling (multi-PRINS) technique, using ddNTPs between two PRINS reactions can block the free 3’-end generated in the previous PRINS reaction, thus avoiding the next PRINS reaction, using it as a primer to perform spurious elongation at nondesired sites. However, by omitting the blocking step and taking advantage of the color mixing, we developed a simple and rapid multi-PRINS technique to simultaneously detect three chromosomes in the same cell. With this protocol, one can create a third color using the two most common forms of labeled dUTP (biotin- and digoxigenin-labeled dUTP) and two fluorochromes (fluorescein and rhodamine). The signals at the centromeres of three different chromosomes displayed perfect yellow, red, and green colors, respectively. The entire procedure could be completed in less than 90 min because the blocking step was omitted. This protocol is practical and efficient for multi-PRINS so that even more than three chromosome targets could be detected in the same cell.
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Acknowledgments
The authors are grateful to Dr. Kieron Legge for careful review of the manuscript and to Mr. Marc Bronsard for his excellent technical help. This study was supported by a grant from the Canadian Institute for Health Research to Régen Drouin. Macoura Gadji is a student scholar of the Public Health Minister (MSP) and the National Center of Blood Transfusion (CNTS) of Senegal. Macoura Gadji was the recipient of a studentship from the The Foundation for Research into Children’s Diseases. Régen Drouin holds the Canada Research Chair in Genetics, Mutagenesis and Cancer.
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Yan, J., Gadji, M., Krabchi, K., Drouin, R. (2006). New Rapid Multicolor PRINS Protocol. In: Pellestor, F. (eds) PRINS and In Situ PCR Protocols. Methods in Molecular Biology™, vol 334. Humana Press. https://doi.org/10.1385/1-59745-068-5:3
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DOI: https://doi.org/10.1385/1-59745-068-5:3
Publisher Name: Humana Press
Print ISBN: 978-1-58829-549-1
Online ISBN: 978-1-59745-068-3
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