PRINS for the detection of gene deletions in cancer.
The chromosomal regions 13q14 and 17p13 often are found rearranged in hematopoietic tumors in humans, but the rearrangements can be subtle and can escape detection on gross cytogenetic analysis. For example, submicroscopic perturbations of the RB1 and p53 tumor suppressor genes, located in 13q14 and 17p13, respectively, frequently occur in leukemias; this has been confirmed by molecular methods such as fluorescence in situ hybridization (FISH). Our group modified the primed in situ labeling (PRINS) method to study RB1 and p53 in cultured bone marrow cells from leukemia patients with known deletions within the 13q14 or 17p13 regions. Locus-specific oligonucleotide probes ("PRINS primers") were annealed to chromosomal DNA on glass slides and extended in the presence of the four trinucleotide precursors, biotin-16-dUTP, and Taq DNA polymerase. After addition of avidin-conjugated fluorophores, the resulting signals could be visualized by fluorescence microscopy in metaphase spreads and interphase nuclei of controls but were absent in the corresponding preparations from patients. The results of these and similar studies suggest that with further development, PRINS might be used as a convenient and rapid alternative to FISH in the delineation of deletions involving single genes or unique sequences.
Tharapel
Avirachan T
AT
Department of Pediatrics, University of Tennessee, Memphis, TN, USA.
Wachtel
Stephen S
SS
D001710:Biotin / Q000378:metabolism; D001854:Bone Marrow Cells; D002478:Cells, Cultured; D004273:DNA, Neoplasm / Q000032:analysis* / Q000235:genetics; D017353:Gene Deletion; D006801:Humans; D008856:Microscopy, Fluorescence; D009369:Neoplasms / Q000235:genetics*; D020401:Primed In Situ Labeling / Q000379:methods*
D004273:DNA, Neoplasm; D001710:Biotin