Approach to genotyping errors caused by nontemplated nucleotide addition by Taq DNA polymerase.
Thermostable DNA polymerases can catalyze nontemplated addition of a nucleotide to the 3' end of amplification products. This presents a potential source of error in genotyping studies employing Taq DNA polymerase to amplify microsatellite loci. Although the activity is marker specific, experimental variation is often seen in the degree of modification. Consequently, for a given microsatellite marker, an allele may be inconsistently identified as either the unmodified or modified amplification product. Full automation of high-throughput genotyping has been hampered by the need for manual editing of data because of this source of allele misidentification. In this study we estimate a 1% to 3% error rate attributable to nontemplated nucleotide addition in the ABI PRISM genotyping system. We present a PCR-based strategy to minimize this source of error.
Smith
J R
JR
National Center for Human Genome Research, National Institutes of Health, Bethesda, Maryland 20892, USA. jsmith@nchgr.nih.gov
Carpten
J D
JD
Brownstein
M J
MJ
Ghosh
S
S
Magnuson
V L
VL
Gilbert
D A
DA
Trent
J M
JM
Collins
F S
FS
D000483:Alleles; D016477:Artifacts; D001331:Automation; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D018900:Dinucleotide Repeats / Q000235:genetics*; D005819:Genetic Markers; D005838:Genotype; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D019914:Taq Polymerase
D005819:Genetic Markers; D019914:Taq Polymerase; D004259:DNA-Directed DNA Polymerase