Abstract
The degradation state of some biological traces recovered from the crime scene requires
the amplification of very short fragments to attain a useful mitochondrial (mt)DNA
sequence. We have previously introduced two mini-multiplex assays that amplify 10
overlapping control region (CR) fragments in two separate multiplex PCRs, which brought
successful CR consensus sequences from even highly degraded DNA extracts. This procedure
requires a total of 20 sequencing reactions per sample, which is laborious and cost
intensive. For only moderately degraded samples that we encounter more frequently
with typical mtDNA casework material, we developed two new multiplex assays that use
a subset of the mini-amplicon primers but embrace larger fragments (midis) and require
only 10 sequencing reactions to build a double-stranded CR consensus sequence. We
used a preceding mtDNA quantitation step by real-time PCR with two different target
fragments (143 and 283 bp) that roughly correspond to the average fragment sizes of the different multiplex
approaches to estimate size-dependent mtDNA quantities and to aid the choice of the
appropriate PCR multiplexes with respect to quality of the results and required costs.
Keywords
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References
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Article info
Publication history
Published online: March 02, 2009
Identification
Copyright
© 2009 Elsevier Ireland Ltd. Published by Elsevier Inc. All rights reserved.
