Dye shift: a neglected source of genotyping error in molecular ecology

Molecular ecologists must be vigilant in detecting and accounting for genotyping error, yet potential errors stemming from dye-induced mobility shift (dye shift) may be frequently neglected and largely unknown to researchers who employ 3-primer systems with automated genotyping. When left uncorrecte...

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Bibliographic Details
Published in:Molecular ecology resources 2011-05, Vol.11 (3), p.514-520
Main Authors: SUTTON, JOLENE T, ROBERTSON, BRUCE C, JAMIESON, IAN G
Format: Article
Language:English
Subjects:
allele size
alleles
Diagnostic Errors
Dyes
ecologists
ecology
Ecology - methods
Electrophoresis - methods
Fluorescent Dyes - metabolism
fluorescent label
Genotype
genotyping
loci
M13
microsatellite
Microsatellite Repeats
mobility shift
Molecular Biology - methods
quality control
researchers
Staining and Labeling - methods
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Summary:Molecular ecologists must be vigilant in detecting and accounting for genotyping error, yet potential errors stemming from dye-induced mobility shift (dye shift) may be frequently neglected and largely unknown to researchers who employ 3-primer systems with automated genotyping. When left uncorrected, dye shift can lead to mis-scoring alleles and even to falsely calling new alleles if different dyes are used to genotype the same locus in subsequent reactions. When we used four different fluorophore labels from a standard dye set to genotype the same set of loci, differences in the resulting size estimates for a single allele ranged from 2.07 bp to 3.68 bp. The strongest effects were associated with the fluorophore PET, and relative degree of dye shift was inversely related to locus size. We found little evidence in the literature that dye shift is regularly accounted for in 3-primer studies, despite knowledge of this phenomenon existing for over a decade. However, we did find some references to erroneous standard correction factors for the same set of dyes that we tested. We thus reiterate the need for strict quality control when attempting to reduce possible sources of genotyping error, and in cases where different dyes are applied to a single locus, perhaps mistakenly, we strongly discourage researchers from assuming generic correction patterns.
ISSN:1755-0998
1755-098X
1755-0998
DOI:10.1111/j.1755-0998.2011.02981.x