Summary
Since its discovery by Koch in 1989, primed in situ labeling (PRINS) reaction provides an alternative approach for direct detection of human chromosomes. The multiple color (multi)-PRINS technique can simultaneously and specifically display different chromosomes with different colors in the same metaphase or interphase nucleus by using sequential labeling of different chromosome targets. We developed a triple-PRINS reaction on uncultured amniotic cells by omitting the blocking step and taking advantage of mixing two fluorochromes (fluorescein and rhodamine) to create a third color for simultaneous detection in the same amniocytes of three different chromosome targets, e.g., chromosomes 18, X, and Y. Fluorescent signals corresponding to chromosomes 18, X, and Y were shown as yellow, red, and green color spots, respectively. Multi-PRINS is as accurate and reliable as multicolor fluorescent in situ hybridization (multi-FISH) for the detection of aneuploidies involving chromosomes 18, X, and Y. Furthermore, multi-PRINS represents a faster and more cost-effective alternative to FISH for prenatal testing of aneuploidy in uncultured amniocytes.
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References
Cremer, T., Lichter, P., Borden, J., Ward, D. C., and Manuelidis, L. (1988) Detection of chromosome aberrations in metaphase and interphase tumor cells by in situ hybridization using chromosome-specific library probes. Hum. Genet. 80,235–246.
Eiben, B., Trawicki, W., Hammans, W., Goebel, R., Pruggmayer, M., and Epplen, J. T. (1999) Rapid prenatal diagnosis of aneuploidies in uncultured amniocytes by fluorescence in situ hybridization . Evaluation of >3,000 cases. Fetal Diagn. Ther. 14, 193–197.
Koch, J. E., Kolvraa, S., Petersen, K. B., Gregersen, N., and Bolund, L. (1989) Oligonucleotide-priming methods for the chromosome-specific labelling of alpha satellite DNA in situ. Chromosoma 98, 259–265.
Pellestor, F., Girardet, A., Lefort, G., Andreo, B., and Charlieu, J. P. (1995) Selection of chromosome-specific primers and their use in simple and double PRINS techniques for rapid in situ identification of human chromosomes. Cytogenet. Cell Genet. 70, 138–142.
Koch, J., Hindkjaer, J., Kolvraa, S., and Bolund, L. (1995) Construction of a panel of chromosome-specific oligonucleotide probes (PRINS-primers) useful for the identification of individual human chromosomes in situ. Cytogenet. Cell Genet. 71, 142–147.
Speel, E. J., Lawson, D., Hopman, A. H., and Gosden, J. (1995) Multi-PRINS: multiple sequential oligonucleotide primed in situ DNA synthesis reactions label specific chromosomes and produce bands. Hum. Genet. 95, 29–33.
Hindkjaer, J., Koch, J., Brandt, C., Kolvraa, S., and Bolund, L. (1996) Primed in situ labeling (PRINS). A fast method for in situ labeling of nucleic acids. Mol. Biotechnol. 6, 201–211.
Cinti, C., Stuppia, L., and Maraldi, N. M. (2002) Combined use of PRINS and FISH in the study of the dystrophin gene. Am. J . Med . Genet . 107, 115–118.
Kadandale, J. S., Wachtel, S. S., Tunca, Y., Wilroy, R. S., Jr., Martens, P. R., and Tharapel, A. T. (2000) Localization of SRY by primed in situ labeling in XX and XY sex reversal. Am. J. Med. Genet. 95, 71–74.
Kadandale, J. S., Wachtel, S. S., Tunca, Y., Martens, P. R., Wilroy, R. S., and Tharapel, A. T. (2002) Deletion of RBM and DAZ in azoospermia: evaluation by PRINS. Am. J. Med. Genet. 107, 105–108.
Tharapel, A. T., Kadandale, J. S., Martens, P. R., Wachtel, S. S., and Wilroy, R. S. Jr. (2002) Prader Willi/Angelman and DiGeorge/velocardiofacial syndrome deletions: diagnosis by primed in situ labeling (PRINS). Am. J . Med. Genet. 107, 119–122.
Pellestor, F., Girardet, A., Coignet, L., Andreo, B., and Charlieu, J. P. (1996) Assessment of aneuploidy for chromosomes 8, 9, 13, 16, and 21 in human sperm by using primed in situ labeling technique. Am. J. Hum. Genet. 58,797–802.
Gosden, J. R. and Lawson, D. (1997) Multiple sequential oligonucleotide primed in situ DNA syntheses (MULTI-PRINS). In: Gosden, J. R. (ed.). PRINS and in situ PCR protocols. Humana Press, Totowa, NJ. pp 39–44.
Hindkjaer, J., Koch, J., Terkelsen, C., Brandt, C. A., Kolvraa, S., and Bolund, L. (1994) Fast, sensitive multicolor detection of nucleic acids in situ by PRimed IN Situ labeling (PRINS). Cytogenet. Cell Genet. 66, 152–154.
Mennicke, K., Yang, J., Hinrichs, F., Muller, A., Diercks, P., and Schwinger, E. (2003) Validation of primed in situ labeling for interphase analysis of chromosomes 18, X, and Y in uncultured amniocytes. Fetal Diagn. Ther. 18, 114–121.
Yan, J., Bronsard, M., and Drouin, R. (2001) Creating a new color by omission of 3’ end blocking step for simultaneous detection of different chromosomes in multi-PRINS technique. Chromosoma 109, 565–570.
Yan, J., Marceau, D., and Drouin, R. (2001) Tetrasomy 8 is associated with a major cellular proliferative advantage and a poor prognosis. Two cases of myeloid hematologic disorders and review of the literature. Cancer Genet. Cytogenet. 125, 14–20.
Krabchi, K., Gros-Louis, F., Yan, J., et al. (2001) Quantification of all fetal nucleated cells in maternal blood between the 18th and 22nd weeks of pregnancy using molecular cytogenetic techniques. Clin. Genet. 60, 145–150.
Krabchi, K., Gadji, M., Yan, J., and Drouin, R. (2006) Dual-color primed in situ labeling (PRINS) for in situ detection of fetal cells in maternal blood. Methods Mol. Biol. 334, 141–149.
Gadji, M., Krabchi, K., and Drouin, R. (2005) Simultaneous identification of chromosomes 18, X and Y in uncultured amniocytes by using multi-primed in situ labelling technique. Clin. Genet. 68, 15–22.
Yan, J., Gadji, M., Krabchi, K., and Drouin, R. (2006) New rapid multicolor PRINS protocol. Methods Mol. Biol. 334, 3–13.
Pellestor, F., Anahory, T., Andreo, B., et al. (2004) Fast multicolor primed in situ protocol for chromosome identification in isolated cells may be used for human oocytes and polar bodies. Fertil. Steril. 81, 408–415.
Spurbeck, J. L., Zinsmeister, A. R., Meyer, K. J., and Jalal, S. M. (1996) Dynamics of chromosome spreading. Am. J. Med. Genet. 61, 387–393.
Acknowledgments
We are grateful to Marc Bronsard for technical help. This study was supported by a grant from the Canadian Institute for Health Research to Régen Drouin. M.G. was the recipient of a studentship from the “The Foundation for Research into Children’s Diseases”. J.Y. is a research scholar (junior I level) of the “Fonds de la Recherche en Santé du Québec” program. R.D. holds the Canada Research Chair in Genetics, Mutagenesis and Cancer.
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Gadji, M., Krabchi, K., Yan, J., Drouin, R. (2008). Application of Multi-PRINS to Simultaneously Identify Chromosomes 18, X, and Y in Prenatal Diagnosis. In: Hahn, S., Jackson, L.G. (eds) Prenatal Diagnosis. Methods in Molecular Biology™, vol 444. Humana Press. https://doi.org/10.1007/978-1-59745-066-9_4
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DOI: https://doi.org/10.1007/978-1-59745-066-9_4
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