Sequencing homopolymer tracts and repetitive elements.
We investigated the use of Taq dye primer and Taq terminator sequencing chemistry to optimize the quality of sequence data obtained from templates containing homopolymer tracts and repetitive elements. In direct side-by-side comparisons using the Applied Biosystems Model 373A Fluorescent Sequencer, the Taq terminator sequencing chemistry gave much cleaner and more consistent results on long homopolymer tracts and dinucleotide repeats. We also investigated various thermal cycling conditions and determined that higher annealing temperatures and longer denaturation times improved the ability to sequence through these problem templates.
Robbins
C M
CM
National Center for Human Genome Research, National Institutes of Health, Bethesda, MD 20892-4470, USA.
Hsu
E
E
Gillevet
P M
PM
D001483:Base Sequence; D004396:Coloring Agents; D004247:DNA / Q000737:chemistry*; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D005784:Gene Amplification; D016045:Human Genome Project; D008969:Molecular Sequence Data; D012091:Repetitive Sequences, Nucleic Acid / Q000235:genetics*; D016415:Sequence Alignment / Q000379:methods*; D017422:Sequence Analysis, DNA / Q000379:methods*; D019914:Taq Polymerase; D013696:Temperature; D013698:Templates, Genetic
D004396:Coloring Agents; D004247:DNA; D019914:Taq Polymerase; D004259:DNA-Directed DNA Polymerase