phosphorus-radioisotopes and Colorectal-Neoplasms--Hereditary-Nonpolyposis

phosphorus-radioisotopes has been researched along with Colorectal-Neoplasms--Hereditary-Nonpolyposis* in 1 studies

Other Studies

1 other study(ies) available for phosphorus-radioisotopes and Colorectal-Neoplasms--Hereditary-Nonpolyposis

ArticleYear
Optimized PCR labeling in mutational and microsatellite analysis.
    Clinical chemistry, 1998, Volume: 44, Issue:7

    To optimize the labeling and visualization of PCR products we tested different variables, including deoxynucleotide concentration and ratio, dilution of labeled product, number of PCR cycles, and use of one-step or nested labeling protocols. Labeling was achieved using a fixed amount of labeled dATP, whose relative specific activity was varied by adding increasing amounts of cold dATP. Optimal PCR-labeling intensity was reached at dATP concentrations between 0.9 and 7.0 micromol/L, with a peak at 1.8 micromol/L. This concentration corresponded to an optimal ratio between the increase in specific activity and the decrease in DNA yield. Nucleotide imbalances >1:2 were not advantageous. Mutational analysis by single-strand conformational polymorphism (SSCP) was used to validate PCR-labeling protocols. The limiting nucleotide concentrations did not affect SSCP. Clear SSCP patterns were obtained using DNA templates of different sizes derived from several genes. SSCP patterns obtained using one-step or nested PCR-labeling protocols were equivalent and were visualized after overnight exposure, using [alpha35S]dATP as the label. Dilutions of labeled products ranging between 1:10 and 1:2.5 influenced SSCP patterns, and the lowest dilution tested produced better-defined and more-intense signals. Optimized SSCP conditions allowed the detection of novel and previously characterized nucleotide variants. Clear microsatellite typing was also obtained using optimized protocols and [alpha35S]dATP as the label.

    Topics: Colorectal Neoplasms, Hereditary Nonpolyposis; Deoxyadenine Nucleotides; DNA Mutational Analysis; DNA, Neoplasm; Humans; Microsatellite Repeats; Nucleotides; Phosphorus Radioisotopes; Polymerase Chain Reaction; Polymorphism, Single-Stranded Conformational; Reproducibility of Results; Sulfur Radioisotopes

1998