Comparative analysis of human DNA variations by fluorescence-based sequencing of PCR products.
Automated, direct cycle sequencing of purified double-stranded PCR products using Taq polymerase and fluorescently labeled dideoxynucleotide terminators provides a robust and highly reproducible method for identifying DNA sequence variations in sequence-tagged sites. We describe a simple and sensitive strategy that reliably detects the presence of DNA variations when sequencing traces from several different individuals are compared. We also demonstrate the use of this strategy to estimate allele frequencies of single nucleotide substitutions in a population. Taken together, this approach provides an automated method for conducting rapid population studies of candidate gene regions that are in linkage or association with a specific disease and for comparative evolutionary analysis of selected regions of the human genome.
Kwok
P Y
PY
Department of Molecular Biotechnology, University of Washington, Seattle.
Carlson
C
C
Yager
T D
TD
Ankener
W
W
Nickerson
D A
DA
D000483:Alleles; D001331:Automation; D001483:Base Sequence; D004247:DNA / Q000235:genetics*; D004259:DNA-Directed DNA Polymerase; D005470:Fluorometry; D005787:Gene Frequency; D014644:Genetic Variation; D006801:Humans; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D015203:Reproducibility of Results; D017422:Sequence Analysis, DNA / Q000379:methods*; D019914:Taq Polymerase
D004247:DNA; D019914:Taq Polymerase; D004259:DNA-Directed DNA Polymerase