Abstract
Primed in situ labeling (PRINS) can be used to localize single-copy genes and unique sequences. Using a modified PRINS method that incorporates multiple primers for the same sequence, single-step annealing and extension, anti-Taq DNA polymerase antibody, and stringent washing, we localized the human SRY gene to Yp11.31-p11.32 in chromosome preparations in situ. Locus-specific oligonucleotide probes (i.e., PRINS primers) were annealed to chromosomal DNA fixed on glass slides and extended in the presence of the four trinucleotide precursors, biotin-16-2’-deoxyuridine 5’-triphosphate, Tris-HCl, KCl, MgCl2, bovine serum albumin, and Taq DNA polymerase. After reaction with avidin-conjugated fluorophores, the resulting signals could be visualized by fluorescence microscopy in metaphase spreads and in interphase nuclei. This method could prove useful for unique sequences in general.
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References
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Wachtel, S.S., Tharapel, A.T. (2006). PRINS for the Detection of Unique Sequences. In: Pellestor, F. (eds) PRINS and In Situ PCR Protocols. Methods in Molecular Biology™, vol 334. Humana Press. https://doi.org/10.1385/1-59745-068-5:33
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DOI: https://doi.org/10.1385/1-59745-068-5:33
Publisher Name: Humana Press
Print ISBN: 978-1-58829-549-1
Online ISBN: 978-1-59745-068-3
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